The prevalence of molecular markers of drug resistance in Plasmodium vivax from the border regions of Thailand in 2008 and 2014

Abstract

The prevalence of Plasmodium vivax is increasing in the border regions of Thailand; one potential problem confounding the control of malaria in these regions is the emergence and spread of drug resistance. The aim of this study was to determine the genetic diversity in genes potentially linked to drug resistance in P. vivax parasites isolated from four different border regions of Thailand; Thai-Myanmar (Tak, Mae Hong Son and Prachuap Khiri Khan Provinces), and Thai-Cambodian borders (Chanthaburi Province). Isolates were collected from 345 P. vivax patients in 2008 and 2014, and parasite DNA extracted and subjected to nucleotide sequencing at five putative drug-resistance loci (Pvdhfr, Pvdhps, Pvmdr1, Pvcrt-o and Pvk12). The prevalence of mutations in Pvdhfr, Pvdhps and Pvmdr1 were markedly different between the Thai-Myanmar and Thai-Cambodian border areas and also varied between sampling times. All isolates carried the Pvdhfr (58R and 117N/T) mutation, however, whereas the quadruple mutant allele (I₅₇R₅₈M₆₁T₁₁₇) was the most prevalent (69.6%) in the Thai-Myanmar border region, the double mutant allele (F₅₇R₅₈T₆₁N₁₁₇) was at fixation on the Thai-Cambodian border (100%). The most prevalent genotypes of Pvdhps and Pvmdr1 were the double mutant (S₃₈₂G₃₈₃K₅₁₂G₅₅₃) (65.1%) and single mutant (M₉₅₈Y₉₇₆F₁₀₇₆) (46.5%) alleles, respectively on the Thai-Myanmar border while the single Pvdhps mutant (S₃₈₂G₃₈₃K₅₁₂A₅₅₃) (52.7%) and the triple Pvmdr1 mutant (M₉₅₈F₉₇₆L₁₀₇₆) (81%) alleles were dominant on the Thai-Cambodian border. No mutations were observed in the Pvcrt-o gene in either region. Novel mutations in the Pvk12 gene, the P. vivax orthologue of PfK13, linked to artemisinin resistance in Plasmodium falciparum, were observed with three nonsynonymous and three synonymous mutations in six isolates (3.3%).

Keywords: Antimalarial drugs; Drug-resistant mutations; Genetic diversity; Plasmodium vivax.

Graphical abstract

Fig. 1

Prevalence of alleles of the Pvdhfr gene in P. vivax samples collected at four malaria endemic districts of Thailand. The small and big circle represent year 2008 and 2014, respectively. (The two numbers followed each province name represent the numbers of isolates sequenced in 2008, 2014) (bold with underline letters indicate mutant amino acids).

Fig. 2

Prevalence of alleles of the Pvdhps gene in P. vivax samples collected at four malaria endemic districts of Thailand. The small and big circle represent year 2008 and 2014, respectively. (The two numbers followed each province name represent the numbers of isolates sequenced in 2008, 2014) (bold with underline letters indicate mutant amino acids).

Fig. 3

Prevalence of alleles of the Pvmdr1 gene in P. vivax samples collected at four malaria endemic districts of Thailand. The small and big circle represent year 2008 and 2014, respectively. (The two numbers followed each province name represent the numbers of isolates sequenced in 2008, 2014) (bold with underline letters indicate mutant amino acids).

Fig. 4

Distribution and prevalence of tandem repeat variants Pvdhfr collected from Thai-Myanmar and Thai-Cambodian borders. (A) Sequences alignment of Type 1 (wild type, accession no. X98123) and Type 2 amino-acid repeat regions. Dashes (−) represent tandem repeat deletions between amino acid position 82 and 109. Bold underlined letters indicate the tandem repeat. (B) Prevalences of two tandem repeat types obtained from P. vivax isolates from the Thai-Myanmar border (black) and Thai-Cambodian border (blue). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)