Development a fluorescent 96-well assay for dual detection of esterase- and GST-mediated pyrethroid resistance in mosquito

Investigator: Bruce D. Hammock, UC Davis 

Resistance to pyrethroid insecticides involves two primary mechanisms of action: (i) target site insensitivity and (ii) elevation in the activity of detoxification enzymes such as esterases, glutathione Stransferases (GSTs), and cytochrome P450s. Rapid, population-level detection of these resistance mechanisms is critical for maintaining the effectiveness of pyrethroid insecticides. We have developed (with partial funding from a previous MRF grant) a series of pyrethroid-like, fluorescent substrates for the detection ofesterase and GST activities. Here, we propose to test and optimize these fluorescent substrates with authentic and purified esterases and GSTs from mosquito. We will then use these fluorescent substrates as the basis for a rapid, sensitive, and easy to use assay for the simultaneous detection of elevated esterase and/or GST activity in pyrethroid resistance in mosquitoes. The ultimate goal is to develop a simple 96-well format assay that MVCDs can use for the detection of elevated esterase and/or GST activity from the homogenate of a single mosquito.