This study addresses the evolutionary dynamics of antimalarial drug resistance after changes in drug use. We show that chloroquine resistance in Plasmodium falciparum from French Guiana was lost after sustained drug removal, whereas the resistance marker PfCRT K76T remained fixed in the parasite population. This phenotypic reversion was caused by the acquisition of a single additional C350R substitution in PfCRT. This genetic change also impaired susceptibility to piperaquine, suggesting that piperaquine pressure drove the expansion of this allele. These findings have important implications for understanding drug resistance evolution when standard resistance alleles reach fixation and can lose their utility as markers because of adaptive changes at other amino acid positions or loci in the genome.