Dihydrofolate Reductase I164L Mutation in Plasmodium falciparum, Madagascar

rates of gar-ganey (Anas querquedula) staging in the Camargue, southern France, during spring migration. North American ducks and gulls to H5N1 highly pathogenic avian infl uenza viruses.ment of multiple sublineages of H5N1 infl uenza virus in Asia: Implications for pandemic control. To the Editor: Malaria remains a major public health problem and a primary cause of illness in Madagas-car (1). Since 2005, the National Malaria Control Program has revised its treatment policy and replaced chlo-roquine (CQ) with artesunate plus amodiaquine as fi rst-line therapy for uncomplicated malaria and CQ with sulfadoxine-pyrimethamine (SP) for prevention of malaria during pregnancy. The latter choice was partially supported by high effectiveness of SP and absence of pyrimethamine resistance in Madagascar, in contrast to proximal African countries such as the Comoros Islands (2,3). Analysis of the molecular basis of antimalarial drug resistance has demonstrated that mutations in the dihydrofolate reductase (dhfr) and dihydropteroate synthase genes are associated with development of SP resistance. It has been assumed that pyrimethamine resistance conferred by multiple mutations arose through stepwise selection of the S108N single mutant (except for the A16V/ S108T allele). This single-point mutation decreases the sensitivity of dhfr to pyrimethamine in vitro by ≈10× (4). Subsequent mutations, such as N51I and C59R, cause additional decreases in the sensitivity of dhfr to py-rimethamine. Parasites with a triple-mutant allele (51I/59R/108N) are less sensitive to pyrimethamine in vitro, and patients infected with these parasites have a high probability of not responding to SP treatment (5). Addition of I164L to 51I/59R/108N creates a quadruple-mutant allele and decreases the sensitivity of dhfr by ≈1,000× (4), eliminating the clinical effectiveness of SP, as observed in Southeast Asia and South America. However, the situation in Africa seems to be different because most studies conducted since the mid 1990s have shown the quadruple mutant to be rare, even in areas of intensive pyrimethamine use (6). Increasing SP resistance is principally a result of rapid selection for parasites that carry a triple-mutant allele that arose in Southeast Asia and has spread widely in Africa (7,8). In 2006, blood samples were obtained from 114 children 6 months to 15 years of age enrolled in a clinical trial monitoring the effi cacy of SP in treatment of uncomplicated Plasmodi-um falciparum malaria. The dhfr gene from pretreatment samples was se-quenced at the Genomics Platform of the Pasteur Institute in Paris, France. Four (3%) samples contained the 108N single-mutant allele, …