Prevalence of mutations in the Plasmodium falciparum chloroquine resistance transporter, PfCRT, and association with ex vivo susceptibility to common anti-malarial drugs against African Plasmodium falciparum isolates

Abstract

Background: The Plasmodium falciparum chloroquine transporter gene (pfcrt) is known to be involved in chloroquine and amodiaquine resistance, and more particularly the mutations on the loci 72 to 76 localized within the second exon. Additionally, new mutations (T93S, H97Y, C101F, F145I, M343L, C350R and G353V) were recently shown to be associated with in vitro reduced susceptibility to piperaquine in Asian or South American P. falciparum strains. However, very few data are available on the prevalence of these mutations and their effect on parasite susceptibility to anti-malarial drugs, and more particularly piperaquine in Africa.

Methods: A molecular investigation of these mutations was performed in 602 African P. falciparum parasites collected between 2017 and 2018 on malaria patients hospitalized in France after a travel in African countries. Associations between genotypes and in vitro susceptibilities to piperaquine and standard antimalarial drugs were assessed.

Results: None of the mutations, previously described as associated with piperaquine resistance, was found in the 602 P. falciparum African isolates. The K76T mutation is associated with resistance to chloroquine (p < 0.0002) and desethylamodiaquine (p < 0.002) in Africa. The K76T mutation is not associated with in vitro reduced susceptibility to piperaquine. The mutation I356T, identified in 54.7% (n = 326) of the African isolates, was significantly associated with reduced susceptibility to quinine (p < 0.02) and increased susceptibility to mefloquine (p < 0.04). The K76T and I356T mutations were significantly associated in West African isolates (p = 0.008).

Conclusion: None of the mutations in pfcrt found to be associated with piperaquine reduced susceptibility in Asia or South America (T93S, H97Y, C101F, F145I, M343L C350R and G353V) were found in the 602 African isolates including the three isolates with reduced susceptibility to piperaquine. The K76T mutation, involved in resistance to chloroquine and amodiaquine, and the I356T mutation were not associated with in vitro reduced susceptibility to piperaquine. Differences in mefloquine susceptibility between I356 and 356T isolates were, while statistically different, minimal. Further analyses are needed with a more important sample size from the same geographic area to confirm the role of the I356T mutation on quinine susceptibility.

Keywords: Africa; Antimalarial drug; I356T; In vitro; Malaria; Molecular marker; Plasmodium falciparum; Resistance; pfcrt.

Fig. 1

Dot plot of the IC₅₀ values distribution of P. falciparum isolates from West Africa and Central Africa for chloroquine (CQ), quinine (QN), desethylamodiaquine (DQ), lumefantrine (LMF), mefloquine (MQ), piperaquine (PPQ), dihydroartemisinin (DHA), artesunate (AS) and pyronaridine (PND) according to I356T. Each dot represents the log10(IC₅₀) for each isolate, red dots represent wild type I356 isolates (I), blue dots mutant 356T isolates (T) and yellow dots the log10(median IC₅₀) value for each drug. The y-axis represents the log10(IC₅₀) in nM. * Significant differences were found between wild type (I356) and mutant (356T) in West Africa (QN: p = 0.001, MQ: p = 0.01, PND: p = 0.005) and in Central Africa (QN: p = 0.02, MQ: p = 0.04, CQ: p = 0.01)