Functional genetic validation of key genes conferring insecticide resistance in the major African malaria vector, <i>Anopheles gambiae</i>

Abstract

Resistance in <i>Anopheles gambiae</i> to members of all 4 major classes (pyrethroids, carbamates, organochlorines, and organophosphates) of public health insecticides limits effective control of malaria transmission in Africa. Increase in expression of detoxifying enzymes has been associated with insecticide resistance, but their direct functional validation in <i>An. gambiae</i> is still lacking. Here, we perform transgenic analysis using the GAL4/UAS system to examine insecticide resistance phenotypes conferred by increased expression of the 3 genes-<i>Cyp6m2</i>, <i>Cyp6p3</i>, and <i>Gste2</i>-most often found up-regulated in resistant <i>An. gambiae</i> We report evidence in <i>An. gambiae</i> that organophosphate and organochlorine resistance is conferred by overexpression of GSTE2 in a broad tissue profile. Pyrethroid and carbamate resistance is bestowed by similar <i>Cyp6p3</i> overexpression, and <i>Cyp6m2</i> confers only pyrethroid resistance when overexpressed in the same tissues. Conversely, such <i>Cyp6m2</i> overexpression increases susceptibility to the organophosphate malathion, presumably due to conversion to the more toxic metabolite, malaoxon. No resistant phenotypes are conferred when either <i>Cyp6</i> gene overexpression is restricted to the midgut or oenocytes, indicating that neither tissue is involved in insecticide resistance mediated by the candidate P450s examined. Validation of genes conferring resistance provides markers to guide control strategies, and the observed negative cross-resistance due to <i>Cyp6m2</i> gives credence to proposed dual-insecticide strategies to overcome pyrethroid resistance. These transgenic <i>An. gambiae</i>-resistant lines are being used to test the "resistance-breaking" efficacy of active compounds early in their development.