Nationwide assessment of pyrethroid resistance escalation and investigation of its molecular basis in Anopheles funestus from Cameroon

Abstract

<title>Abstract</title> <bold>Background</bold>: Malaria prevention still relies greatly on vector control interventions. However, increasing levels of resistance to pyrethroids across Africa have significantly reduced the efficacy of pyrethroid-based interventions leading to a stagnation and/or an increase of malaria burden. Consequently, it is imperative to nation-widely characterize the extent and molecular basis of this resistance. <bold>Methods</bold>: This study was conducted from 2020 to 2021 in a South-North transect across Cameroon to assess patterns of resistance to insecticides in <italic>Anopheles funestus,</italic> establish its impact on efficacy of insecticide-treated nets and investigate underlying molecular mechanisms. Taqman assay and nested PCR were used to determine <italic>Plasmodium </italic>sporozoite prevalence. Genotyping of known genetic markers as well as species identification was done using PCR technique while quantitative PCR was used to determine gene expression. <bold>Results: </bold>High pyrethroid resistance intensity was noticed in all sites with mortalities ranging from 80-93.9%, 84.9-96.7% and 82% for permethrin, deltamethrin and alphacypermethrin at 10x concentration respectively. This high level of resistance led to dramatic inefficacy of pyrethroid-only nets with 0-17% mortality recorded 24-hour post exposure while PBO-based nets displayed optimal efficacy. The L119F-<italic>GSTe2</italic> allele was higher in the South (56-68%) compared to the North (20-37%) meanwhile the P450-linked 4.3kb structural variant was fixed contrasting with the absence of the <italic>CYP6P9a/b-R</italic>, 6.5kb insertion and N485I-<italic>Ace1</italic> alleles. Furthermore, the L119F-<italic>GSTe2</italic>allele confers significant ability to mosquito to survive permethrin. In addition, the <italic>CYP325A, CYP6P5, CYP6P9a/b, and </italic>the<italic> Carb2514 </italic>were the most overexpressed genes in pyrethroid resistant mosquitoes. However, no further association was noticed between these alleles/genes and increasing doses of pyrethroids. <bold>Conclusion</bold>: This study confirms the escalation of pyrethroid resistance across Cameroon and the inefficacy of pyrethroid-only nets and highlights genes potentially implicated in the aggravation of insecticide resistance.