Discovery of possible molecular factors underlying spinosad resistance in Leptinotarsa decemlineata

Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae), or Colorado potato beetle (CPB), is an insect pest known for the extensive damage it does to potato plants and its ability to gain insecticide resistance via diverse molecular mechanisms. It is hypothesized that members of two enzyme superfamilies, cytochrome P450s (CYPs) and glutathione s-transferases (GSTs), underlie this resistance and are modulated in CPB following exposure to insecticides including spinosad. This project addresses this hypothesis by quantifying, using qRT-PCR, the expression of transcripts coding for CYPs and GSTs in response to low (0.5 µg) and high (2 µg) spinosad doses applied for 4 or 24 hours on adult CPBs. An RNAi-based approach, which decrease expression of select transcripts, is used to assess the effect of such reduction on insect viability. Results show, following exposure to a low dose of spinosad for 4 hours, increased levels of CYP6a23 (P < 0.05) and CYP4g15 (n.s.) transcripts, as well as reduced levels of CYP12a5 (P < 0.05) transcripts, when compared with control. Exposure to high dose of spinosad for 4 hours caused the down-regulation of CYP6a13, CYP6d4 (P < 0.05), GST1 and GST-T (P < 0.01) transcripts. Following dsRNA-based knockdown of CYP6a23 and CYP4g15, viability was quantified in L. decemlineata, with the latter target causing significant mortality in dsRNA- versus saline-injected insects. In conclusion, modulation of various CYP- and GST-associated transcript targets was demonstrated in spinosad-exposed L. decemlineata and this information aligns with the development of RNAi-based approaches aimed at novel molecular targets to control this potato pest.