Molecular surveillance of antifolate drug resistance markers in Plasmodium vivax from Khyber Pakhtunkhwa province, northwest Pakistan

Abstract

The emergence and spread of antimalarial drug resistance pose significant challenges in the fight against malaria. Mutations in dihydrofolate reductase (DHFR) and dihydropteroate synthetase (DHPS) in Plasmodium vivax are associated with sulfadoxine-pyrimethamine (SP) drug resistance. This study assessed SP resistance status in P. vivax isolates collected in Khyber Pakhtunkhwa province, Pakistan, by analyzing mutations in pvdhfr and pvdhps. Both genes were successfully amplified concurrently from 112 Pakistan P. vivax isolates. Sequence analysis of pvdhfr indicated that mutations F57L, S58R, and S117N were present with frequencies of 0.9 %, 31.3 %, and 46.4 %, respectively. The predominant wild-type haplotype F57S58T61S117 was identified in 51.8 % of samples, whereas mutant haplotypes were also detected: F57R58T61N117 (29.5 %), F57S58T61N117 (16.9 %), F57R58T61S117 (0.9 %), and L57R58T61S117 (0.9 %). In pvdhps, the sole mutation A383 G was found at a low frequency of 1.8 %, leading to a mutant haplotype S382G383K512A553V585. The integrated analysis of pvdhfr and pvdhps haplotypes showed that the wild-type haplotype was the most prevalent (50.9 %), followed by mutant haplotypes F57R58T61N117/ S382A383K512A553V585 (28.6 %) and F57S58T61N117/S382A383K512A553V585 (16.9 %). These findings indicate a relatively low level of antifolate resistance in Pakistan P. vivax isolates, suggesting that Pakistan P. vivax may still be amenable to SP treatment. Nevertheless, the persistence of similar mutation rates and patterns associated with SP resistance in the Pakistan pvdhfr and pvdhps populations, despite the absence of current SP pressure, underscores the importance of ongoing monitoring of SP resistance in the Pakistan P. vivax population. © 2025 Elsevier B.V.