Molecular epidemiology of malaria in Cameroon.: XIV.: Plasmodium falciparum chloroquine resistance transporter (PFCRT) gene sequences of isolates before and after chloroquine treatment

Laboratory studies have strongly suggested that the gene coding for Plasmodium falciparum chloroquine resistance transporter (PFCRT) may play a determinant role in chloroquine resistance. A clinical study in Mali also found evidence for selection of the key PFCRT amino acid substitution, Lys76Thr, in patients who fail to respond to chloroquine treatment. To test the hypothesis that in vivo selection of mutant PFCRT alleles occurs after chloroquine treatment, PFCRT and merozoite surface antigen 2 (msa-2) polymorphisms were compared between 61 pretreatment and posttreatment paired samples from children with either clinical or parasitologic failure. There were six wild-type PFCRT alleles, 44 mutant alleles, and 11 mixed alleles among pretreatment isolates. All posttreatment parasites had mutant PFCRT alleles. Recrudescence accounted for 42 of 61 posttreatment infections, while 19 posttreatment infections were due to new infection (including all isolates with Lys-76 before treatment and Thr-76 after treatment). Seven pretreatment isolates with mixed PFCRT alleles had only Thr-76 on recrudescence, providing a direct evidence for in vivo selection for mutant PFCRT. Although the presence of mutant PFCRT alleles in pretreatment isolates is not predictive of chloroquine treatment failure, our data support the hypothesis that in vivo selection for recrudescent parasites carrying mutant PFCRT alleles occurs. These results may have important implications for the future surveillance of chloroquine resistance by the use of molecular markers.