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.