Agricultural practices drive multifactorial pyrethroid resistance in Anopheles coluzzii populations from southern Benin.

BACKGROUND

The rapid spread of insecticide resistance threatens the effectiveness of malaria vector control across sub-Saharan Africa. In southern Benin, the respective contributions of agricultural pesticide use and urban vector control interventions to the selection of resistant Anopheles populations remain poorly documented.

METHODS

Pyrethroid resistance was investigated in Anopheles coluzzii populations collected in southern Benin from two zones: two agricultural areas (Sèmè-Kpodji and Abomey-Calavi) and one non-agricultural urban area (Yenawa). WHO tube bioassays were performed using deltamethrin (0.05%) and alpha-cypermethrin (0.05%). Molecular species was conducted using PCR-SINE200, whereas the kdr-West (L1014F) mutation was genotyped by PCR. The activities of cytochrome P450 monooxygenases, esterases, and glutathione S-transferases (GSTs) were quantified, while the expression levels of CYP6Z1, CYP9K1, and GSTE2 were measured by RT-qPCR.

RESULTS

All populations exhibited confirmed resistance to both pyrethroids, with mortality rates ranging from 78% to 82% for deltamethrin and from 36% to 89% for alpha-cypermethrin. Resistance to alpha-cypermethrin was markedly greater in agricultural areas, particularly in Sèmè-Kpodji (36%) and Abomey-Calavi (52%), than in Yenawa (89%). Molecular analyses identified Anopheles coluzzii as the sole species detected (100%). The resistant kdr-west allele occurred at high frequencies, ranging from 73.3% to 93.3%, and no susceptible homozygotes were observed. Biochemical assays revealed significantly elevated CYP450, GST, and esterase activities, especially in Sèmè-Kpodji and Yenawa, indicating the contribution of metabolic detoxification mechanisms. In contrast, RT‒qPCR revealed no overexpression of CYP6Z1, CYP9K1, or GSTE2 relative to the susceptible N'Gousso strain (fold-change values of 0.26-0.58, 0.06-0.21, and 0.03-0.06, respectively), despite significant inter-population differences in transcript levels.

CONCLUSIONS

Pyrethroid resistance in Anopheles coluzzii from southern Benin is widespread and multifactorial, involving both the near-fixation of the kdr-west mutation and increased detoxification enzyme activities. The stronger phenotypic resistance observed in agricultural areas highlights the role of agricultural pesticide exposure, whereas elevated kdr resistance markers at urban sites indicate additional selection pressure from public health and domestic insecticide use. These findings underscore the urgent need for integrated resistance management strategies that combine agricultural pesticide regulation, continuous resistance surveillance, and deployment of next-generation vector control tools.