Abstract
Insecticide resistance in Anopheles mosquitoes threatens the effectiveness of key malaria control tools such as insecticide-treated nets (ITNs) and indoor residual spraying (IRS) in Ethiopia. Genomic analysis is essential to model known and novel molecular markers of insecticide resistance for effective resistance management. This study investigated insecticide resistance genes using whole-genome sequencing in a major malaria vector, Anopheles arabiensis sampled across the whole regions of Ethiopia and found high geographic and temporal variability in genes associated with insecticide resistance. The Vgsc-L995F target-site substitution in the voltage-gated sodium channel gene was highly prevalent in northern Ethiopia but less common at other sites. Metabolic modes of resistance in western Ethiopia were indicated by the high frequencies of copy number variants observed at the cytochrome P450 cluster Cyp6aa/p and the carboxylesterase Coeae2-7g. Frequencies of genetic markers associated with molecular target sites and metabolic resistance were generally lower in the Central Rift Valley. However, copy number variants (CNVs) at Gste2 and Cyp9k1 were observed at high frequency. We observed seasonal shifts in both target-site and metabolic marker frequencies, including increasing frequencies of Vgsc-L995F and several cytochrome P450 variants during the major transmission season. These patterns were specific to each location. Findings indicate that molecular insecticide resistance arises from a complex interplay of factors, including malaria control interventions, agricultural practices, human behavior, and possibly vector behavior. Selection scans revealed signals of selection on chromosome 2L, centered on the Coejhe1-5e genes in Werkamba, northernmost Ethiopia. Additional signals were detected on chromosome 3L (~ 20 Mb), near genes that may regulate detoxification pathways, including those associated with the ubiquitin-proteasome system in Asossa, western Ethiopia. Findings highlight the importance of integrating genomic surveillance of resistance markers into entomological monitoring to strengthen insecticide resistance management. They also underscore the need to investigate lesser-known sources of adaptive change that may have significant consequences for vector control.