Genomic characterization of antimalarial drug resistance markers in plasmodium falciparum using selective whole genome amplification and the nomad Nanopore sequencing in Huye, Gisagara, and Kirehe districts

Abstract

Malaria remains a major public health challenge, with increasing cases and emerging drug resistance threatening global control efforts. This study evaluates and compares two DNA amplification strategies selective whole genome amplification (sWGA) and MVP within the nomadic sequencing (NOMAD) nanopore sequencing protocol to enhance genomic surveillance of Plasmodium falciparum in Rwanda. Our findings demonstrate that sWGA produces significantly higher sequencing reads compared to MVP, although it does not markedly improve genome coverage or depth. The NOMAD protocol delivers robust performance across multiple key resistance and vaccine target genes, maintaining high sequencing depth and coverage even with varying DNA input levels from clinical field samples. Molecular surveillance identified mutations in crucial resistance markers including pfkelch13, pfcrt, pfmdr1, pfdhfr, and pfdhps, providing actionable insights into regional parasite populations and drug resistance dynamics. This work underscores the practical utility of rapid, field-deployable nanopore sequencing combined with selective amplification methods to support real-time malaria control efforts. Strategic use of sWGA should balance increased throughput benefits against higher costs and longer processing times, while MVP may suit rapid, low-bias applications. Continued optimization and nationwide scale-up of this protocol, alongside sustained molecular surveillance and functional validation of emerging mutations, will strengthen Rwanda’s capacity for adaptive malaria control and elimination