Establishment of tetra-primer ARMS-PCR technique for detecting exo-E415G SNP in Plasmodium falciparum isolates from Central Highlands of Vietnam

Abstract

Drug resistance in Plasmodium falciparum parasites is associated with the single nucleotide polymorphism
(SNP) exo-E415G, which is considered as a marker to reduce the susceptibility to artimisinin and leads
to treatment failure. This study aims to establish a procedure based on tetra-primer ARMS PCR technique
to detect the SNP exo-E415G in malaria patients who experienced treatment failure with artemisinin. DNA sample from P. falciparum 3D7 isolate was used to optimise the procedure based on tetra-primer ARMS PCR technique to detect exo-E415G SNP. The method was then applied to screen for exo-E415G SNP on 123 blood samples from patients who experienced treatment failure with artemisinin in 3 provinces (Gia Lai, Dak Nong, Dak Lak) from Central Highlands of Vietnam. A DNA fragment (749 bp) containing the exo-E415G SNP was amplified using primer pairs E415G-Fc and E415G-Rc. Specific primers for the wildtype alen (A alen) and the  mutant alen (G alen) amplified fragments with the length of 316 and 474 bp, respectively. The genotyping results were consistent with Sanger sequencing. The tetra-primer ARMS PCR technique was used for genotyping exo-E415G SNPon 123 patients who failed with artemisinin treatment, the frequency of exo-E415G alen was observed at 78% (96/123). In conclusion, the simple tetra-primer ARMS PCR technique was established to detect the SNP Exo-E415G. The prevalence of the mutant alen Exo-E415G was 78% in the Central Highlands of Vietnam.