BackgroundEstablishing the extent, geographical distribution and mechanisms of insecticide resistance in malaria vectors is a prerequisite for resistance management. Here, we report a widespread distribution of insecticide resistance in the major malaria vector An. funestus across Uganda and western Kenya under the control of metabolic resistance mechanisms.Methodology/Principal FindingsFemale An. funestus collected throughout Uganda and western Kenya exhibited a Plasmodium infection rate between 4.2 to 10.4%. Widespread resistance against both type I (permethrin) and II (deltamethrin) pyrethroids and DDT was observed across Uganda and western Kenya. All populations remain highly susceptible to carbamate, organophosphate and dieldrin insecticides. Knockdown resistance plays no role in the pyrethroid and DDT resistance as no kdr mutation associated with resistance was detected despite the presence of a F1021C replacement. Additionally, no signature of selection was observed on the sodium channel gene. Synergist assays and qRT-PCR indicated that metabolic resistance plays a major role notably through elevated expression of cytochrome P450s. DDT resistance mechanisms differ from West Africa as the L119F-GSTe2 mutation only explains a small proportion of the genetic variance to DDT resistance.ConclusionThe extensive distribution of pyrethroid and DDT resistance in East African An. funestus populations represents a challenge to the control of this vector. However, the observed carbamate and organophosphate susceptibility offers alternative solutions for resistance management.

Pyrethroid resistance in malaria vector, An. funestus is increasingly reported across Africa, threatening the sustainability of pyrethroid-based control interventions, including long lasting insecticidal nets (LLINs). Managing this problem requires understanding of the molecular basis of the resistance from different regions of the continent, to establish whether it is being driven by a single or independent selective events. Here, using a genome-wide transcription profiling of pyrethroid resistant populations from southern (Malawi), East (Uganda), and West Africa (Benin), we investigated the molecular basis of resistance, revealing strong differences between the different African regions. The duplicated cytochrome P450 genes (CYP6P9a and CYP6P9b) which were highly overexpressed in southern Africa are not the most upregulated in other regions, where other genes are more overexpressed, including GSTe2 in West (Benin) and CYP9K1 in East (Uganda). The lack of directional selection on both CYP6P9a and CYP6P9b in Uganda in contrast to southern Africa further supports the limited role of these genes outside southern Africa. However, other genes such as the P450 CYP9J11 are commonly overexpressed in all countries across Africa. Here, CYP9J11 is functionally characterized and shown to confer resistance to pyrethroids and moderate cross-resistance to carbamates (bendiocarb). The consistent overexpression of GSTe2 in Benin is coupled with a role of allelic variation at this gene as GAL4-UAS transgenic expression in Drosophila flies showed that the resistant 119F allele is highly efficient in conferring both DDT and permethrin resistance than the L119. The heterogeneity in the molecular basis of resistance and cross-resistance to insecticides in An. funestus populations throughout sub-Saharan African should be taken into account in designing resistance management strategies.

BackgroundAlthough the An. funestus group conceals one of the major malaria vectors in Africa, little is known about the dynamics of members of this group across the continent. Here, we investigated the species composition, infection rate and susceptibility to insecticides of this species group in Uganda.MethodsIndoor resting blood-fed Anopheles adult female mosquitoes were collected from 3 districts in Uganda. Mosquitoes morphologically belonging to the An. funestus group were identified to species by PCR. The sporozoite infection rates were determined by TaqMan and a nested PCR. Susceptibility to major insecticides was assessed using WHO bioassays. The potential role of four candidate resistance genes was assessed using qRT-PCR.ResultsAn. funestus s.s. and An. parensis, were the only members of the An. funestus group identified. Both species were sympatric in Masindi (North-West), whereas only An. parensis was present in Mityana (Central) and Ntungamo (South-West). The Plasmodium falciparum infection detected in An. parensis (4.2%) by TaqMan could not be confirmed by nested PCR, whereas the 5.3% infection in An. funestus s.s. was confirmed. An. parensis was susceptible to most insecticides, however, a moderate resistance was observed against deltamethrin and DDT. In the sympatric population of Masindi, resistance was observed to pyrethroids (permethrin and deltamethrin) and DDT, but all the resistant mosquitoes belonged to An. funestus s.s. No significant over-expression was observed for the four P450 candidate genes CYP6M7, CYP9K1, CYP6P9 and CYP6AA4 between deltamethrin resistant and control An. parensis. However, when compared with the susceptible FANG An. funestus s.s strain, the CYP9K1 is significantly over-expressed in An. parensis (15-fold change; P < 0.001), suggesting it could play a role in the deltamethrin resistance.ConclusionThe contrasting infection rates and insecticide susceptibility profiles of both species highlights the importance of accurate species identification for successful vector control programs.