High prevalence of Pfdhfr–Pfdhps quadruple mutations associated with sulfadoxine–pyrimethamine resistance in Plasmodium falciparum isolates from Bioko Island, Equatorial Guinea

Abstract

BackgroundSulfadoxine–pyrimethamine (SP) is recommended for intermittent preventive treatment of malaria in Africa. However, increasing SP resistance (SPR) affects the therapeutic efficacy of the SP. As molecular markers, Pfdhfr (dihydrofolate reductase) and Pfdhps (dihydropteroate synthase) genes are widely used for SPR surveillance. This study aimed to assess the prevalence of Pfdhfr and Pfdhps genes mutations and haplotypes in Plasmodium falciparum isolates collected from Bioko Island, Equatorial Guinea (EG).MethodsIn total, 180 samples were collected in 2013–2014. The single nucleotide polymorphisms (SNPs) of the Pfdhfr and Pfdhps genes were identified with nested PCR and Sanger sequencing. The genotypes and linkage disequilibrium (LD) tests were also analysed.ResultsSequences of Pfdhfr and Pfdhps genes were obtained from 92.78% (167/180) and 87.78% (158/180) of the samples, respectively. For Pfdhfr, 97.60% (163/167), 87.43% (146/167) and 97.01% (162/167) of the samples carried N51I, C59R and S108N mutant alleles, respectively. The prevalence of the Pfdhps S436A, A437G, K540E, A581G, and A613S mutations were observed in 20.25% (32/158), 90.51% (143/158), 5.06% (8/158), 0.63% (1/158), and 3.16% (5/158) of the samples, respectively. In total, 3 unique haplotypes at the Pfdhfr locus and 8 haplotypes at the Pfdhps locus were identified. A triple mutation (CIRNI) in Pfdhfr was the most prevalent haplotype (86.83%), and a single mutant haplotype (SGKAA; 62.66%) was predominant in Pfdhps. A total of 130 isolates with 12 unique haplotypes were found in the Pfdhfr and Pfdhps combined haplotypes, 65.38% (85/130) of them carried quadruple allele combinations (CIRNI-SGKAA), whereas only one isolate (0.77%, 1/130) was found to carry the wild-type (CNCSI-SAKAA). For LD analysis, the Pfdhfr N51I was significantly associated with the Pfdhps A437G (P < 0.05).ConclusionBioko Island possesses a high prevalence of the Pfdhfr triple mutation (CIRNI) and Pfdhps single mutation (SGKAA), which will undermine the pharmaceutical effect of SP for malaria treatment strategies. To avoid an increase in SPR, continuous molecular monitoring and additional control efforts are urgently needed in Bioko Island, Equatorial Guinea.

Prevalence of PfDHFR and PfDHPS Mutations Associated with Drug Resistance Among Pregnant Women Receiving IPTp-SP at Msambweni County Referral Hospital, Kwale County, Kenya

“…Findings from this study demonstrated very high prevalence of SNPs at the two important genes that confer SP resistance. The high prevalence of 89% observed at PfDHFR51I in this study was similarly to (85%-100%) documented in western Kenya and elsewhere (10,(21)(22)(23)(24)29), but contrary to the 21% reported in India (26). Although a study in DRC reported a conservative prevalence of 60% (23) at 59R, other studies found slightly higher prevalence (87%-98%) at this codon (10,21,22,24,26,29), compared to 78% demonstrated in this study.…”

“…The high prevalence of 89% observed at PfDHFR51I in this study was similarly to (85%-100%) documented in western Kenya and elsewhere (10,(21)(22)(23)(24)29), but contrary to the 21% reported in India (26). Although a study in DRC reported a conservative prevalence of 60% (23) at 59R, other studies found slightly higher prevalence (87%-98%) at this codon (10,21,22,24,26,29), compared to 78% demonstrated in this study. In agreement with our study, investigations elsewhere found a prevalence of ≥97% at 108N (10,(21)(22)(23)(24)26,27,29).…”

“…Although a study in DRC reported a conservative prevalence of 60% (23) at 59R, other studies found slightly higher prevalence (87%-98%) at this codon (10,21,22,24,26,29), compared to 78% demonstrated in this study. In agreement with our study, investigations elsewhere found a prevalence of ≥97% at 108N (10,(21)(22)(23)(24)26,27,29). In contrast to average prevalence (66%-68%) reported in India and Gabon (24,26), prevalence of >90% at PfDHPS437G was reported in this study, western Kenya and parts of central Africa.…”

“…Similar to our findings, other studies have reported high prevalence (78%-97%) of quintuple PfDHFR/PfDHPS haplotype mutations in western Kenya (10,21). Studies elsewhere have documented quadruple mutations (65%) in Equatorial Guinea (22) and 48% in DRC (23), and triple mutations (92%) in Gabon (24) and 61-71% in Burkina Faso (25) as the most prevalent. Similar to India, where double mutation was the most prevalent (26,27), a study in Brazil reported double mutation as most prevalent but did not find quintuple or quadruple mutations (28).…”

Efficacy of Artemisinin-lumefantrine for the treatment of uncomplicated malaria after more than a decade of its use in Kenya

“…The instant and extensive increase of anti-malarial drug resistance are hindering the progress of malaria control [12,44]. In this study, we confirm a high cure rate and efficacy (100% ACPR) of the nationally recommended AL for the treatment of uncomplicated malaria since its introduction in Kenya over a decade ago.…”