isrctn.org Identifier: ISRCTN37517549.

Polymorphisms in the Plasmodium falciparum pfmdr1 gene were assayed in pretreatment samples and in samples from patients reinfected following therapy with artemether-lumefantrine. The pfmdr1 alleles 86N, 184F, and 1246D significantly increased in prevalence after treatment. All samples had a single pfmdr1 copy. Treatment with artemether-lumefantrine selects for polymorphisms that may alter antimalarial drug response.Due to widespread resistance of Plasmodium falciparum to drugs such as chloroquine and sulfadoxine-pyrimethamine, artemisinin combination therapy (ACT) is currently advocated in Africa as a means of improving treatment efficacy and slowing the spread of resistance. The rationale behind ACT is to rapidly reduce the parasite burden with a short-acting artemisinin compound, leaving a longer-acting partner drug to eliminate the remaining parasites, thus reducing the chance of selection of drug-resistant parasites. In Southeast Asia, where the risk of reinfection following therapy is low, the combination of artesunate and mefloquine has been effective at slowing the spread of resistance (10). It is unclear whether ACT will be as successful in preventing the selection of resistant parasites in Africa, where parasite transmission rates are generally much higher. In a recent study from Tanzania, treatment with the widely advocated ACT artemether-lumefantrine (AL) was associated with selection of newly infecting parasites containing the pfmdr1 86N allele (16), which has been associated with decreased in vitro sensitivity to artemisinins and lumefantrine (4, 5).We recently completed a clinical trial including AL at a rural site in Uganda with extremely high transmission intensity (1). Briefly, children aged 1 to 10 years with uncomplicated falciparum malaria received directly observed therapy and were followed for 28 days. Molecular 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 the day of failure (late clinical failure or late parasitological failure) were analyzed for polymorphisms in merozoite surface protein 1 (MSP-1) and MSP-2 using nested PCR as previously described (2). An outcome was defined as recrudescence if all MSP-1 and MSP-2 gene alleles present at the time of failure were present at the time of treatment initiation and as a new infection otherwise. To test the hypothesis that AL selects for polymorphisms in the pfmdr1 gene, we compared the prevalences of key alleles and pfmdr1 copy numbers between pretreatment isolates and new isolates following treatment with AL.Polymorphisms studied were pfmdr1 N86Y, Y184F, N1042D, and D1246Y. Alleles were identified using nested PCR and restriction fragment length polymorphism methods, and copy number was assessed by TaqMan quantitative PCR as previously described (4, 12). Single (3D7)-and three-copy (W2mef) standards were used as positive controls. Reactions were done in quadruplicate and repeated for a cha...

Based on the high PPV and NPV, HRP2-based RDTs are likely to be the best diagnostic choice for areas with medium-to-high malaria transmission rates in Africa.

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