Evaluating the Genetic Landscape of (Plasmodium falciparum) PfKelch13 Gene Polymorphisms in Côte d'Ivoire Following a Decade of Artemisinin-based Combination Therapy

Introduction: Artemisinin-based combination therapies (ACTs) are the mainstay of malaria treatment globally. However, their effectiveness is threatened by the emergence of resistance in Plasmodium falciparum (P.f), particularly in Southeast Asia (SEA). Specific mutations within the pfKelch13 gene, such as Cys-580-Tyr, Arg-539-Thr, Tyr-493-His, and Ile-543-Thr, have been strongly linked to delayed parasite clearance following ACT treatment. This study aimed to investigate polymorphisms within the pfKelch13 gene (also known as the K13-propeller or K13 gene) in four regions of Côte d'Ivoire. Côte d'Ivoire experiences year-round malaria transmission and has utilized ACTs for over a decade. Methods: From September 2013 to July 2014, samples were collected from patients residing in Abidjan (south), Ayamé (southeast), Man (west), and Korhogo (north) who presented with microscopically confirmed uncomplicated malaria. Following Plasmodium falciparum DNA extraction, nested PCR was employed to amplify an 849 bp fragment of the pfKelch13 gene. Amplicons were subsequently sequenced and analyzed using BioEdit software. Results: 531 DNA sequences were analyzed including 301 (58.7%) from Abidjan, 61(11.5%) from Ayamé, 93 (17.5%) from Man and 76 (13.4%) from Korhogo. Only 20 isolates carrying 22 mutations were observed including 6 non-synonymous single-nucleotide polymorphisms (nsSNP): 4 in Abidjan (Asp-535-Met; Ala-578-Ser; Phe-583-Ser and Ile-601-Thr) and 2 in Korhogo (Asp-559-Asn and Val-510-Met). Only synonymous SNP (sSNP) were identified in the two other Towns. The proportion of mutated pfK13 sequences is 3.8% (20/531). Conclusion: The identification of non-synonymous mutations in this study underscores the importance of heightened surveillance for potential ACT resistance in Plasmodium falciparum within Côte d'Ivoire. Combining in vitro assays, such as the Ring-stage Survival Assay, with molecular testing will be crucial for definitively determining the phenotypic impact of these mutations on parasite susceptibility to ACTs.