Molecular Surveillance of Plasmodium falciparum Drug Resistance Markers in Clinical Samples from Botswana

Abstract

Drug-resistant Plasmodium falciparum is a major threat to global malaria control and elimination efforts. In Botswana, a southern African country approaching malaria elimination, P. falciparum molecular data are not available. Parasites were assessed through pollymerase chain reaction (PCR) for confirmation of positive rapid diagnostic tests, multiplicity of infection (MOI), and drug resistance markers among isolates from clinical uncomplicated malaria cases collected at health facilities. Of 211 dried blood spot samples selected for the study, 186 (88.2%) were PCR positive for P. falciparum. The mean MOI based on MSP1 genotyping was 2.3 and was not associated with age. A high prevalence of wild-type parasites for pfcrt and pfmdr1 was found, with a haplotype frequency (K76/N86) of 88.8% and 17.7% of the isolates having two copies of the pfmdr1 gene. For pfATPase6, all the parasites carried the wild-type S769 allele. Sequencing showed no evidence of non-synonymous mutations associated with reduced artemisinin derivative sensitivity in the P. falciparum k13 gene. In conclusion, we found that P. falciparum parasites in Botswana were mostly wild type for the drug resistance markers evaluated. Yet, there was a high rate of a molecular marker associated to reduced sensitivity to lumefantrine. Our results indicate the need for systematic drug efficacy surveillance to complement malaria elimination efforts.

Figure 1.

Size variant allele frequency for each allelic family observed in the analyzed samples and number of samples harboring each MSP1 allelic family. For K1, the mean size ± SE of PCR fragments was 247 base pairs (bp) ± 4 bp, K-S D statistic = 0.18, P = n.s.; for MAD20, the mean size ± SE was 222 bp ± 5 bp, K-S D statistic = 0.22, P = n.s.; and for RO33, the mean size ± SE was 165 bp ± 3 bp, K-S D statistic = 0.23, P = n.s. This figure appears in color at www.ajtmh.org.