Validation of PfSNP-LAMP-Lateral Flow Dipstick for Detection of Single Nucleotide Polymorphism Associated with Pyrimethamine Resistance in Plasmodium falciparum.

Suganya Yongkiettrakul,Darin Kongkasuriyachai,Namfon Nawattanapaibool,Fassou René Kolié,Chayanut Suansomjit,Jetsumon Sattabongkot,Sureemas Buates,Saradee Warit,Niwat Kangwanrangsan,Wang Nguitragool

doi:10.3390/diagnostics10110948

Abstract

The loop-mediated isothermal amplification coupled with lateral flow dipstick (PfSNP-LAMP-LFD) was recently developed to detect single nucleotide polymorphism (AAT → ATT), corresponding to substitution of asparagine to isoleucine at amino acid position 51 in the P. falciparum dhfr-ts gene associated with antifolate resistance. In this present study, the PfSNP-LAMP-LFD was validated on 128 clinical malaria samples of broad ranged parasite densities (10 to 87,634 parasites per microliter of blood). The results showed 100% accuracy for the detection of single nucleotide polymorphism for N51I mutation. Indeed, the high prevalence of N51I in the Pfdhfr-ts gene detected in the clinical samples is in line with reports of widespread antifolate resistant P. falciparum in Thailand. The relationship between enzyme choice and reaction time was observed to have an effect on PfSNP-LAMP-LFD specificity; however, the method yielded consistent results once the conditions have been optimized. The results demonstrate that PfSNP-LAMP-LFD is a simple method with sufficient sensitivity and specificity to be deployed in routine surveillance of antifolate resistance molecular marker and inform antimalarial management policy.

Highlights

The global malaria morbidity and mortality have decreased substantially over the past decades, yet malaria still remains a significant global health threat
The primary objective of this study was to validate the PfSNP-Loop-mediated isothermal amplification (LAMP)-lateral flow dipstick (LFD) using clinical samples while exploring the robustness of the method when performed under different conditions
Our demonstrated that the Pf single nucleotide polymorphisms (SNPs)-LAMP-LFD could accurately distinguish SNP (AAT → ATT, N51I)

Summary

Introduction

The global malaria morbidity and mortality have decreased substantially over the past decades, yet malaria still remains a significant global health threat. The increasing threats of multidrug-resistant malaria have raised the urgency to accelerate the global malaria elimination agenda. Diagnostics 2020, 10, 948 is the hotspot for multidrug-resistant malaria, where resistant parasites have emerged and spread across the globe. Health Organization (WHO) to be used as seasonal malaria chemoprevention (SMC) in children under. SP blocks the enzymes in the folate synthesis pathway of P. falciparum, the dihydropteroate synthetase (DHPS), and the dihydrofolate reductase (DHFR), respectively [4,5]. Pyrimethamine resistance arose from specific point mutations resulting in amino acid substitutions in the DHFR at positions N51I, C59R, S108N/T, and I164L [6,7,8,9,10,11,12]

Objectives

Methods

Results

Conclusion