# Investigating the potential role of metabolic resistance genes in conferring cross-resistance to pyrethroids and polycyclic aromatic hydrocarbon pollutants in the major malaria vector Anopheles coluzzii

## TL;DR

This study shows that certain mosquito genes linked to insecticide resistance can also break down environmental pollutants, potentially increasing resistance in the wild.

## Contribution

The study identifies specific P450 enzymes in malaria mosquitoes that metabolize PAHs, linking environmental pollutants to insecticide resistance.

## Key findings

- Microsomes from resistant mosquitoes significantly depleted fluorene and fluoranthene.
- CYP6P3 and CYP6Z3 showed high fluorene metabolism, while CYP6Z2 and CYP6Z3 metabolized fluoranthene.
- No naphthalene metabolism was observed with the tested P450s.

## Abstract

Polycyclic aromatic hydrocarbons (PAHs) are a class of ubiquitous and recalcitrant environmental pollutants generated from petroleum activities and/or biological conversion of organic materials. Environmental exposure of mosquitoes to these pollutants can potentially select resistance to insecticides used in public health for vector control. To understand the cross-resistance potentials between PAHs and pyrethroid insecticides, microsomal fractions prepared from Anopheles coluzzii mosquitoes obtained from agricultural sites and a laboratory susceptible strain, Ngousso, were tested with three major PAHs - fluorene, fluoranthene and naphthalene. Recombinant P450s previously associated with pyrethroid resistance in Anopheles gambiae (CYPs 6M2, 6Z2, 6Z3, 9J5, 6P3, 6P4, 6P5, CYP9K1) and Anopheles funestus CYP6P9a were also used to investigate metabolism of the above PAHs alongside the microsome.

Microsomes prepared from pyrethroid-resistant Anopheles coluzzii significantly (P = 0.001, r = 0.99) depleted fluorene and fluoranthene with percentage depletions of 73%±0.5 and 43%0.0 ± 2.2, respectively. A Steady-state kinetic study demonstrated that the microsome has a high affinity for fluorene with a Km and turnover of 58.69 µM ± 20.47​ and 37.016 min-1 ± 3.67, respectively. On the other hand, significant metabolism of fluorene up to 47.9%±2.3 (P = 0.001, r = 0.99) and 52.8%±0.8 (P = 0.001, r = 0.97) depletions were observed with recombinant CYP6P3 and CYP6Z3, respectively. Other P450s showed little to no metabolism with fluorene. CYP6Z2 and CYP6Z3 metabolised fluoranthene with percentage depletions of 50.4%±4.9 (P = 0.003, r = 0.96) and 60.3% ±5.3 (P = 0.002, r = 0.84), respectively. However, no metabolism of naphthalene was observed with all the recombinant P450s used in this study.

This study demonstrates that P450 monooxygenases from the malaria vectors can metabolise PAHs, highlighting the potential of these environmental pollutants selecting the P450s, driving insecticide resistance in field populations of major malaria vectors.