Genetic architecture driving plasmodium falciparum fitness and resistance in Kigali city, district of Nyarugenge.

Abstract

Malaria remains a major global health concern, disproportionately affecting sub-Saharan Africa, which accounts for over 90% of the disease’s burden in terms of cases and mortality(P. Venkatesan, 2025). Rwanda has made remarkable progress in malaria control, significantly reducing disease incidence and mortality over the past two decades. However, an outbreak has been observed since 2024 and new hotspots have emerged, including the City of Kigali. To reinforce malaria control strategies, it is important to perform molecular surveillance of drug resistance and parasite strain diversity across the country. This study investigated the genetic architecture driving Plasmodium falciparum fitness and antimalarial drug resistance in Nyarugenge District, Kigali City, Rwanda. A total of 167 P. falciparum samples were successfully sequenced. Among them, 64.67% carried nonsynonymous mutations in the Pfk13 gene, 58.08% in pfmdr1, and 4.79% in pfcrt. Several mutations detected were either validated by the World Health Organization (WHO) or considered candidate markers of drug resistance by the same organization, including Pfk13 R561H, A675V, P574L ; pfmdr1 Y184F; and pfcrt K76T. The most prevalent mutations were pfmdr1 Y184F (58.08%) and Pfk13 R561H (41%), often found in association with Pfk13 K189T. Notably, a strong correlation was observed between R561H and K189T, suggesting that K189T may function as a compensatory mutation. Strain genotyping using the pfmsp2 gene revealed three major strain types circulating in the study area: 60% belonged to Pf3D7_13_v3, 32% to Pf3D7_5_v3, and 8% to Pf3D7_7_v3. Sequencing cost analysis indicated that Oxford Nanopore Technology (ONT) is more costeffective for multi-gene analysis per sample, while Sanger sequencing remains more economical for targeting single genes under 1000 base pairs. Finally, the study highlights the critical role of the Cullin 3 (CUL3) E3 ubiquitin ligase complex in protein degradation, particularly its interaction with the BTB/POZ domain of the P. falciparum K13 protein, a key focus for understanding the mechanisms underlying artemisinin resistance