RESISTANCE BIOMARKERS ASSESSMENT OF PLASMODIUM FALCIPARUM IN SCHOOL AGED CHILDREN

Abstract

Efficient antimalarials against Plasmodium falciparum have resulted in a substantial reduction in global malaria deaths. The ongoing spread of P. falciparum resistance to antimalarial drugs poses a very major challenge in malaria control, particularly within the waning immune systems of young children. Recently, resistance to artemisinin has been shown to be associated with mutations in the Kelch 13 propeller gene (Pfk13) of Plasmodium falciparum in South Asia. Functional Genomic Studies on Plasmodium falciparum Chloroquine Resistance Transporter Gene (Pfcrt) and Mutations Associated with the Chloroquine (CQ) Resistance A total of 280 archived samples were obtained from primary school children during a period covering May 2022 - November 2022, in the city Rawalpindi, Pakistan. MightyPrep extract genomic DNA Of the samples tested, 67 were positive for P. falciparum (24% prevalence rate). This resulted in 67 samples that were used for amplification of the molecular marker resistance to Pfcrt by PCR. The amplicons 
were purified after PCR amplification and sequenced using Sanger Sequencing. Point mutation identification of the sequence data were performed using BioEdit software.BioEdit Sequence Alignment Editor (Ibis Therapeutics, Carlsbad, CA) analysis of amino acid changes in 14 samples among falciparum isolates showed the following alterations in particular positions: F76C, Y66H, L70A, Y58C, T59V, V65I P67L, T81L, Y60S, Y66S, P67T and I71F. This included newly reported mutations at 76 and a subsequent amino acid substitution in one of the Pfcrt gold standard biomarker genes. Furthermore, new changes in amino acid were detected, and the prevalence of concern for Pfcrt increased from 0% to 5.0%. Upon genetic similarities present in all our samples, a phylogenetic evolutionary relationship was constructed. This demonstrates persistent P. falciparum resistance to Pfcrt and the need for strong ongoing monitoring and surveillance. Acknowledging the expanded prevalence of mutant Pfcrt genes resistant to chloroquine and ongoing emergence, it is critical that antimalarial drugs be repurposed now for future use as partner therapies.