Co-Expression of a Homologous Cytochrome P450 Reductase Is Required for In Vivo Validation of the Tetranychus urticae CYP392A16-Based Abamectin Resistance in Drosophila

SIMPLE SUMMARY: The two-spotted spider mite, Tetranychus urticae, is one of the most damaging agricultural pests worldwide, feeding on over 1100 plant species and causing extensive damage to several crops. Chemical acaricides remain the most widely used strategy to control this pest. However, T. urticae has developed significant resistance to numerous acaricide compounds, due to certain features of mite biology and extensive acaricide applications that lead to the selection of resistant pests and subsequently the emergence of resistant populations. Several molecular/genetic mechanisms may contribute to these highly resistant phenotypes. Such mechanisms frequently involve expression of P450 detoxification enzymes, which act together with a partner protein named cytochrome P450 reductase (CPR). In this study, we investigated the potential of a mite P450 enzyme, CYP392A16, to confer resistance to the acaricide abamectin in vivo, when expressed in tissues of the model fruit fly Drosophila melanogaster. We confirmed that expression of this enzyme contributes to abamectin resistance in the fruit fly model, but only when a homologous mite CPR is co-expressed. Our findings indicate that the Drosophila model system can be engineered to facilitate validation of the candidate mite P450s, in order to elucidate resistance mechanisms and their underlying interactions. ABSTRACT: Overexpression of the cytochrome P450 monooxygenase CYP392A16 has been previously associated with abamectin resistance using transcriptional analysis in the two-spotted spider mite Tetranychus urticae, an important pest species worldwide; however, this association has not been functionally validated in vivo despite the demonstrated ability of CYP392A16 to metabolize abamectin in vitro. We expressed CYP392A16 in vivo via a Gal4 transcription activator protein/Upstream Activating Sequence (GAL4/UAS) system in Drosophila melanogaster flies, driving expression with detoxification tissue-specific drivers. We demonstrated that CYP392A16 expression confers statistically significant abamectin resistance in toxicity bioassays in Drosophila only when its homologous redox partner, cytochrome P450 reductase (TuCPR), is co-expressed in transgenic flies. Our study shows that the Drosophila model can be further improved, to facilitate the functional analysis of insecticide resistance mechanisms acting alone or in combination.