A single E205D allele of a key P450CYP6P3is driving metabolic pyrethroid resistance in the major African malaria vectorAnopheles gambiae

Abstract

Deciphering the molecular drivers of insecticide resistance is paramount to extend the effectiveness of malaria vector control tools. Here, we demonstrated that the E205D amino acid change in a key metabolic resistance P450CYP6P3drives pyrethroid resistance in the major malaria vector,Anopheles gambiae. Spatio-temporal whole genome Poolseq analyses in Cameroon detected a major P450-linked locus on chromosome 2R beside the sodium channel locus.In vitrometabolism assays with recombinantly expressedCYP6P3protein revealed that the catalytic efficiency of 205D was 2.5 times higher than E205 with α-cypermethrin. Similar patterns were observed for permethrin. Overexpression of the 205D allele in transgenic flies confers higher more pyrethroids and carbamates resistance, compared to controls. A DNA-based assay further supported that theCYP6P3-205D variant strongly correlates with pyrethroid resistance in field populations (OR=26.4; P<0.0001) and that it reduces the efficacy of pyrethroid-only LLINs with homozygote RR genotype exhibiting significantly higher survival following PermaNet 3.0 exposure compared to the SS genotype (OR: 6.1, p = 0.0113). Furthermore, theCYP6P3-E205D combines with thekdrtarget-site resistance mechanisms to worsen the loss of bednet efficacy. The 205D mutation is now predominant in West and Central Africa but less abundant or absent in East and South Africa with signs of introgression withAn. coluzziiin Ghana. This study highlights the importance of P450-based resistance and designs field-applicable tools to easily track the spread of metabolic resistance and assess its impact on control interventions.One Sentence Summary:The major obstacle to malaria control and elimination is the spread of parasite resistance to anti-malarial drugs, and mosquito resistance to insecticides. In this study, we identified a key point mutation E205D in the metabolic geneCYP6P3(cytochrome P450) conferring resistance to pyrethroids by enhancing the breakdown of insecticides used for bednets impregnation. DNA-based assays were then designed and used to determine the spread of the resistance across Africa and demonstrate that theCYP6P3-205D allele works together with the knockdown resistance in the voltage-gated Sodium channel to reduce the efficacy of insecticide-treated bednets.

Copy number variants underlie the major selective sweeps in insecticide resistance genes inAnopheles arabiensisfrom Tanzania

“…While a CNV was found in the cluster, it did not include Cyp6aa1 and was not associated with resistance to deltamethrin. In Cameroon, a non-CNV haplotype has been shown to be associated with pyrethroid resistance (Kengne-Ouafo et al, 2024), while two large signals of selection are found around the same gene cluster, in the absence of any CNVs ( Anopheles gambiae 1000 Genomes Consortium et al, 2017). While these haplotypes have not yet been phenotypically tested, we believe it very likely that it is associated with deltamethrin resistance, given the consistent results coming out of our study and the wider literature.…”

Parallel Evolution in Mosquito Vectors—A Duplicated Esterase Locus is Associated With Resistance to Pirimiphos-methyl in Anopheles gambiae

“…More recently, whole-genome sequencing combined with population genetics has revealed the full extent of positive selection acting upon the wild genomes of Anopheles populations ( Miles et al 2017 ; Clarkson et al 2020 ). These data have revealed a complex genetic architecture of insecticide resistance, including the involvement of amino acid mutations, copy number variants (CNVs), and cis-regulatory variation ( Lucas et al 2019 ; Clarkson et al 2021 ; Dyer et al 2023 ; Lucas et al 2023 ; Kengne-Ouafo et al 2024 ).…”