Pyrethroids resistance and distribution of kdr mutations (L1014F and N1575Y) in Anopheles gambiae s.l. along the south-north transect of Benin

BACKGROUND: Pyrethroid resistance in Anopheles gambiae s.l. compromises malaria vector control in sub-Saharan Africa. In Benin, resistance has been documented for over two decades, but the geographic distribution of emerging mutations such as N1575Y remains poorly characterized. This study investigated pyrethroid susceptibility, kdr mutations, and metabolic resistance mechanisms across twelve communes spanning a south-north transect of the country. METHODS: Larvae were collected from twelve sites between January 2024 and January 2025, reared to adults, and tested for susceptibility to deltamethrin (0.05%) and alphacypermethrin (0.05%) using WHO tube assays. Molecular species were identified by PCR, kdr mutations (L1014F, N1575Y) detected by TaqMan genotyping, and enzymatic activities (oxidases, esterases, GSTs) quantified through biochemical assays. RESULTS: All populations showed high resistance to pyrethroids, with mortality consistently below 60%. An. coluzzii predominated (58.2%), followed by An. gambiae s.s. (39.2%) and An. arabiensis (2.8%). The L1014F mutation was highly prevalent across all communes (50-100%), while N1575Y was detected in six communes at frequencies ranging from 1 to 10%. Detoxification enzymes were significantly overexpressed compared to the Kisumu reference strain, confirming the contribution of metabolic resistance. CONCLUSIONS: This large-scale survey confirms widespread pyrethroid resistance in An. gambiae s.l. across Benin, driven by both kdr mutations and metabolic mechanisms. The heterogeneous distribution of N1575Y highlights localized selection pressures and its emerging role in resistance intensification. These findings emphasize the urgent need to deploy PBO-based or dual-active ingredient LLINs, strengthen molecular surveillance, and implement integrated resistance management strategies to sustain malaria control in Benin.