The high level of resistance to azinphosmethyl in both the azinphosmethyl-resistant (AZ-R), near-isogenic strain (136-fold) and the multiply resistant (MA-R) field strain (435-fold) of Colorado potato beetle is complex and due to multiple factors including reduced penetration, enhanced xenobiotic metabolism, and site insensitivity. A slightly reduced penetration rate was observed in the MA-R field strain. This resistance mechanism may be potentiated further by the overall rapid excretion of aziophosmethyl in Colorado potato beetle, as well as by the above-mentioned additional resistance mechanisms. Although no significant differences were noted in monooxygenase activity or in cytochrome P450 content, both resistant strains produced significantly more of the azinphosmethyl metabolites, benzazimide and desmethyl azinphosmethyl-oxon, than the SS strain under both in vivo and in vitro conditions. The AZ-R strain, which is 97% isogenic with the SS strain, showed a significant increase in activity toward chlorodinitrobenzene, indicating glutathione S-transferase involvement. However, there was no increase in azinphosmethyl metabolites in the resistant strains using an in vitro glutathione S-transferase assay. Although not directly responsible for overall resistance, increased activity of glutathione S-transferase(s) in the resistant strains may further metabolize toxic oxidative metabolites of azinphosmethyl which are present at higher levels in the resistant strains than in the SS strain. The AZ-R strain possessed an altered acetylcholinesterase with a reduced substrate affinity for acetylthiocholine. There was a significant reduction (2-fold) in the bimolecular rate constant to azinphosmethyl-oxon in the AZ-R strain compared to that in the susceptible (SS) strain. Differential sensitivity of the acetylcholinesterase activity measured in individual beetles of these two strains to azinphosmethyl-oxon and paraoxon corroborate the presence of an altered acetylcholinesterase. Additionally, this resistant form of the acetylcholinesterase is more sensitive to the inhibitory action of the steroidal glycoalkaloid, tomatine, than the form present in the SS strain. This phenomenon may be at least partly responsible for the reduced fitness associated with these resistant strains. The demonstration of multiple resistant factors used to mitigate the toxicity of azinphosmethyl corroborates the initial findings by us that this resistance was not monofactorial. The wide range of resistance mechanisms employed by CPB is in keeping with its well-recognized status as an agricultural pest that is extremely difficult to control. © 1994 Academic Press. All rights reserved.
