Four-year monitoring of the malaria vector Anopheles funestus in central-west Cameroon reveals an escalation of pyrethroid resistance combined with high malaria transmission

Abstract Background Insecticide resistance presents a critical obstacle to malaria vector control, necessitating ongoing surveillance to guide control strategy. Despite widespread resistance in central Africa, the temporal adaptive changes driving resistance at both phenotypic and genetic levels remain uncharacterised. This study provides a comprehensive, four-year (2020–2023) assessment of Anopheles funestus s.s. in Mibellon, Cameroon, examining sporozoite infection rates and changes in insecticide resistance relative to 2015–2018 data. Methods Susceptibility profile, resistance intensity and cone assays were conducted following the WHO protocols. Sporozoite infection was detected in the mosquito head/thorax by TaqMan assay, confirmed by nested-PCR. Gene expression was assessed using RT-qPCR while insecticide resistance markers were genotyped using allele-specific PCR and LNA. Results Plasmodium sporozoite infection rates ranged from 4 to 21% with the predominance of P. falciparum while P. malariae and P. ovale contributed often as mixed infections. ​Pyrethroid resistance significantly increased over time, with mortalities decreasing from 77.7% in 2015 to 23.2% in 2023 for permethrin and 46.6% in 2016 to just 8.5% in 2023 for deltamethrin, while full susceptibility was noted for organophosphates.​ Worryingly, high intensity of resistance was recorded for all pyrethroids. Partial recovery of susceptibility with PBO suggests other resistance mechanisms beside P450-based metabolic resistance. PBO-based nets yielded high efficacy which decreases slightly over time contrasting with complete loss in efficacy of pyrethroid-only nets. Monitoring the genetic markers revealed a rapid selection of G454A- CYP9K1 and 4.3 kb SV alleles, which increased considerably and reaching high frequency during the same period in which phenotypic resistance intensified. Other resistance markers (A296S- rdl and L119F- GSTe2 ) varied slightly in frequency while the N485I- Ace1 , 6.5 kb SV, and CYP6P9a / b _R alleles were absent throughout the years. Consistent overexpression of CYP9K1 and CYP6P9a/b genes in pyrethroid-resistant mosquitoes highlights their potential role in resistance intensification. Conclusion The high infection rate and co-circulation of three Plasmodium species coupled with intense pyrethroid resistance pose a serious menace to malaria control in this region. To address these complex challenges, current vector control strategies should prioritize the deployment of PBO-based nets and organophosphates for IRS. Continuous vector and parasite surveillance should guide the choice of future interventions to accelerate progress towards malaria elimination Clinical trial Not applicable