Molecular Surveillance of Artemisinin-Based Combination Therapies Resistance in Plasmodium falciparum Parasites from Bioko Island, Equatorial Guinea.

Artemisinin-based combination therapies (ACTs) resistance has emerged and could be diffusing in Africa. As an offshore island on the African continent, the island of Bioko in Equatorial Guinea is considered severely affected and resistant to drug-resistant Plasmodium falciparum malaria. However, the spatial and temporal distribution remain unclear. Molecular monitoring targeting the , , , and genes was conducted to provide insight into the impact of current antimalarial drug resistance on the island. Furthermore, polymorphic characteristics, haplotype network, and the effect of natural selection of the gene were evaluated. A total of 152 Plasmodium falciparum samples (collected from 2017 to 2019) were analyzed for copy number variation of the gene and , , and mutations. Statistical analysis of sequences was performed following different evolutionary models using 96 Bioko sequences and 1322 global sequences. The results showed that the prevalence of and mutations was 73.68%, 78.29%, and 75.66%, respectively. Large proportions of isolates with multiple copies of were observed (67.86%). In Bioko parasites, the genetic diversity of was low, and purifying selection was suggested by Tajima's D test (-1.644, > 0.05) and the dN/dS test (-0.0004438, > 0.05). The extended haplotype homozygosity analysis revealed that _K189T, although most frequent in Africa, has not yet conferred a selective advantage for parasitic survival. The results suggested that the implementation of continuous drug monitoring on Bioko Island is an essential measure. Malaria, one of the tropical parasitic diseases with a high transmission rate in Bioko Island, Equatorial Guinea, especially caused by P. falciparum is highly prevalent in this region and is commonly treated locally with ACTs. The declining antimalarial susceptibility of artemisinin-based drugs suggested that resistance to artemisinin and its derivatives is developing in P. falciparum. Copy number variants in and genetic polymorphisms in , , and can be used as risk assessment indicators to track the development and spread of drug resistance. This study reported for the first time the molecular surveillance of , , and genes in Bioko Island from 2017 to 2019 to assess the possible risk of local drug-resistant P. falciparum.