Field-evolved resistance to neonicotinoids in the mosquito, Anopheles gambiae, is associated with mutations of nicotinic acetylcholine receptor subunits combined with cytochrome P450-mediated detoxification

New insecticides prequalified for malaria control interventions include modulators of nicotinic acetylcholine receptors that act selectively on different subunits leading to variable sensitivity among arthropods. This study aimed to investigate the molecular mechanisms underlying contrasting susceptibility to neonicotinoids observed in wild populations of two mosquito sibling species. Bioassays and a synergist test with piperonyl butoxide revealed that the sister taxa, Anopheles gambiae and An. coluzzii, from Yaounde, Cameroon, both have the potential to develop resistance to acetamiprid through cytochrome P450-mediated detoxification. However, contrary to An. coluzzii, An. gambiae populations are evolving cross-resistance to several active ingredients facilitated by mutations of nicotinic acetylcholine receptors (nAChRs). We sequenced coding regions on the β1 and α6 nAChR subunits where variants associated with resistance to neonicotinoids or to spinosyns have been found in agricultural pests and detected no mutation in An. coluzzii. By contrast, six nucleotide substitutions including an amino acid change in one of the loops that modulate ligand binding and affect sensitivity were present in the resistant species, An. gambiae. Allele frequency distributions were consistent with the spread of beneficial mutations that likely reduce the affinity of An. gambiae nAChRs for synthetic modulators. Our findings provide critical information for the application and resistance management of nAChR modulators in malaria prevention.