Abstract
BackgroundTo combat malaria transmission, the Ugandan government has embarked upon an ambitious programme of indoor residual spraying (IRS) with a carbamate class insecticide, bendiocarb. In preparation for this campaign, we characterized bendiocarb resistance and associated transcriptional variation among Anopheles gambiae s.s. mosquitoes from two sites in Uganda.ResultsGene expression in two mosquito populations displaying some resistance to bendiocarb (95% and 79% An. gambiae s.l. WHO tube bioassay mortality in Nagongera and Kihihi, respectively) was investigated using whole-genome microarrays. Significant overexpression of several genes encoding salivary gland proteins, including D7r2 and D7r4, was detected in mosquitoes from Nagongera. In Kihihi, D7r4, two detoxification-associated genes (Cyp6m2 and Gstd3) and an epithelial serine protease were among the genes most highly overexpressed in resistant mosquitoes. Following the first round of IRS in Nagongera, bendiocarb-resistant mosquitoes were collected, and real-time quantitative PCR analyses detected significant overexpression of D7r2 and D7r4 in resistant mosquitoes. A single nucleotide polymorphism located in a non-coding transcript downstream of the D7 genes was found at a significantly higher frequency in resistant individuals. In silico modelling of the interaction between D7r4 and bendiocarb demonstrated similarity between the insecticide and serotonin, a known ligand of D7 proteins. A meta-analysis of published microarray studies revealed a recurring association between D7 expression and insecticide resistance across Anopheles species and locations.ConclusionsA whole-genome microarray approach identified an association between novel insecticide resistance candidates and bendiocarb resistance in Uganda. In addition, a single nucleotide polymorphism associated with this resistance mechanism was discovered. The use of such impartial screening methods allows for discovery of resistance candidates that have no previously-ascribed function in insecticide binding or detoxification. Characterizing these novel candidates will broaden our understanding of resistance mechanisms and yield new strategies for combatting widespread insecticide resistance among malaria vectors.
Highlights
To combat malaria transmission, the Ugandan government has embarked upon an ambitious programme of indoor residual spraying (IRS) with a carbamate class insecticide, bendiocarb
Insecticide resistance detection World Health Organization (WHO) tube bioassays were performed between January and May 2014 using An. gambiae s.l. mosquitoes collected as larvae in Nagongera and Kihihi (Fig. 1)
Species identification PCR revealed differing species compositions in the two sites: 68% of mosquitoes in Nagongera (n = 925) were identified as An. gambiae s.s. and 32% were identified as Anopheles arabiensis, while those from Kihihi were exclusively
Summary
The Ugandan government has embarked upon an ambitious programme of indoor residual spraying (IRS) with a carbamate class insecticide, bendiocarb. In preparation for this campaign, we characterized bendiocarb resistance and associated transcriptional variation among Anopheles gambiae s.s. mosquitoes from two sites in Uganda. The investigation described is part of a comprehensive malaria surveillance program conducted since 2011 in three areas of Uganda: Walukuba, Kihihi, and Nagongera [6] During this time, an LLIN distribution campaign has been successful in increasing bed net ownership. This high level of pyrethroid resistance may partially explain why only modest declines in malaria metrics were observed following the universal LLIN distribution campaign
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