Cytochrome P450 associated with insecticideresistance catalyzes cuticular hydrocarbonproduction in Anopheles gambiae

The role of cuticle changes in insecticide resistance in the major malariavector Anopheles gambiae was assessed. The rate of internalization of14C deltamethrin was significantly slower in a resistant strain than in asusceptible strain. Topical application of an acetone insecticide formu-lation to circumvent lipid-based uptake barriers decreased the resis-tance ratio by ∼50%. Cuticle analysis by electron microscopy andcharacterization of lipid extracts indicated that resistant mosquitoeshad a thicker epicuticular layer and a significant increase in cuticularhydrocarbon (CHC) content (∼29%). However, the CHC profile andrelative distribution were similar in resistant and susceptible insects.The cellular localization and in vitro activity of two P450 enzymes,CYP4G16 and CYP4G17, whose genes are frequently overexpressedin resistant Anopheles mosquitoes, were analyzed. These enzymesare potential orthologs of the CYP4G1/2 enzymes that catalyze thefinal step of CHC biosynthesis in Drosophila and Musca domestica, re-spectively. Immunostaining indicated that both CYP4G16 and CYP4G17are highly abundant in oenocytes, the insect cell type thought to se-crete hydrocarbons. However, an intriguing difference was indicated;CYP4G17 occurs throughout the cell, as expected for a microsomalP450, but CYP4G16 localizes to the periphery of the cell and lies onthe cytoplasmic side of the cell membrane, a unique position for a P450enzyme. CYP4G16 and CYP4G17 were functionally expressed in insectcells. CYP4G16 produced hydrocarbons from a C18 aldehyde substrateand thus has bona fide decarbonylase activity similar to that ofdmCYP4G1/2. The data support the hypothesis that the coevolutionof multiple mechanisms, including cuticular barriers, has occurred inhighly pyrethroid-resistant An. gambiae.

The cuticular hydrocarbons of the Triatoma sordida species subcomplex (Hemiptera: Reduviidae)

The cuticular hydrocarbons of the Triatoma sordida subcomplex (Hemiptera: Reduviidae: Triatominae) were ana-lysed by gas chromatography and their structures identified by mass spectrometry. They comprised mostly n-alkanes and methyl-branched alkanes with one-four methyl substitutions. n-alkanes consisted of a homologous series from C21-C33 and represented 33-45% of the hydrocarbon fraction; n-C29 was the major component. Methyl-branched alkanes showed alkyl chains from C24-C43. High molecular weight dimethyl and trimethylalkanes (from C35-C39) represented most of the methyl-branched fraction. A few tetramethylalkanes were also detected, comprising mostly even-numbered chains. Several components such as odd-numbered 3-methylalkanes, dimethylalkanes and trimethylalkanes of C37 and C39 showed patterns of variation that allowed the differentiation of the species and populations studied. Triatoma guasayana and Triatoma patagonica showed the most distinct hydrocarbon patterns within the subcomplex. The T. sordida populations from Brazil and Argentina showed significantly different hydrocarbon profiles that posed concerns regarding the homogeneity of the species. Triatoma garciabesi had a more complex hydrocarbon pattern, but it shared some similarity with T. sordida. The quantitative and qualitative variations in the cuticular hydrocarbons may help to elucidate the relationships between species and populations of this insect group.