Microsomal cytochrome P450 monooxygenases in the house fly ( Musca domestica L.): Biochemical changes associated with pyrethroid resistance and phenobarbital induction

Abstract

Differences in the microsomal cytochrome P450 monooxygenase system and its inducibility by sodium phenobarbital (PB) were investigated in insecticide-susceptible (S+) and pyrethroid-resistant (LPR) house fly strains. In comparison to the S+ strain, LPR had higher levels of total cytochrome P450s, cytochrome b 5, NADPH-cytochrome c (P450) reductase, and six monooxygenase activities: aryl hydrocarbon hydroxylation (AHH), aldrin epoxidation (AE), 7-ethoxycoumarin O-deethylation (ECOD), p-chloro- N-methylaniline N-demethylation (PCMA), ethoxyresorufin O-deethylation (EROD), and methoxyresorufin O-demethylation (MROD). The largest differences between strains were noted for MROD (64×) and EROD (56×) activities, and these appeared to be correlated with the monooxygenase-mediated pyrethroid resistance in the LPR strain. Comparisons between susceptible and resistant strains by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed that the LPR strain showed a marked increase in the 54,400-Da protein band, suggesting this may be a cytochrome P450 isozyme involved in conferring part of the monooxygenase-mediated pyrethroid resistance. In vivo administration of 0.25% PB in sugar water to S+ flies induced higher levels of three components of the microsomal monooxygenase system, as well as increased levels of all six monooxygenase activities. In contrast, PB treatment had no significant effect on the level of cytochrome P450 or on EROD, AHH, or AE activity in LPR flies. Additionally, PB caused a decrease in the level of cytochrome b 5 and MROD and PCMA activities, while ECOD activity and the level of NADPH-cytochrome c (P450) reductase were significantly increased. SDS-PAGE indicated that induction of S+ flies with PB caused an increase in at least seven proteins with apparent molecular masses of 47,400; 49,400; 51,500; 52,200; 54,400; 56,000; and 57,500 Da. However, no observable difference was noted in PB-induced LPR flies. Differences in monooxygenase activities, in conjunction with the changes in turnover numbers of ECOD and AE activities in S+ flies and ECOD and MROD activities in LPR flies, suggest that PB treatment induced both quantitative and qualitative changes in the S+ and LPR cytochrome P450 monooxygenases.