Functional characterization of CYP6P9a and CYP6P9b from the malaria mosquito Anopheles funestus

To date vector control interventions targeting disease-transmitting mosquitoes still heavily rely on pyrethroids, an insecticide class targeting the voltage-gated sodium channel. Cytochrome P450 monooxygenases (P450/CYP) are known to play a major role in metabolic pyrethroid resistance in mosquitoes. In the laboratory strain FUMOZ-R of the Malaria mosquito Anopheles funestus CYP6P9a and CYP6P9b are upregulated and shown to confer pyrethroid resistance. To restore the susceptibility towards pyrethroids the P450-inhihbitor piperonyl butoxide is commercially used. As we have shown in previous work, other P450-inhibitors such as azole fungicides have the potential to fill up the vector control toolbox.

For the first time we functionally expressed in High Five cells both CYP6P9a and CYP6P9b using a baculovirus expression system. In a fluorescence assay approach, we tested a range of different P450 inhibitors and pyrethroids and observed significant differences of the metabolic activity towards structurally different pyrethroids. Subsequently, we confirmed these findings and identified a key metabolic pathway of pyrethroids by LC/MS. In fluorescent probe inhibitions assays with recombinantly expressed CYP6P9a and CYP6P9b different IC₅₀ values between enzymes were observed towards several P450 inhibitors and pyrethroids.