Reference genes are used as internal controls in gene expression studies, but their expression levels vary according to tissue types and experimental

Cytochrome P450 monooxygenases (P450s) are involved in metabolic resistance to insecticides and require NADPH cytochrome P450 reductase (CPR) to transfer electrons when they catalyze oxidation reactions. The carmine spider mite, Tetranychus cinnabarinus is an important pest mite of crop and vegetable plants worldwide and its resistance to acaricides has quickly developed. However, the role of CPR on the formation of acaricide-resistance in T. cinnabarinus is still unclear. In this study, a full-length cDNA encoding CPR was cloned and characterized from T. cinnabarinus (designated TcCPR). TcCPR expression was detectable in all developmental stages of T. cinnabarinus, but it’s much lower in eggs. TcCPR was up-regulated and more inducible with fenpropathrin treatment in the fenpropathrin-resistant (FeR) strain compared with the susceptible SS strain. Feeding of double-strand RNA was effective in silencing the transcription of TcCPR in T. cinnabarinus, which resulted in decreasing the activity of P450s and increasing the susceptibility to fenpropathrin in the FeR strain but not in the susceptible strain. The current results provide first evidence that the down-regulation of TcCPR contributed to an increase of the susceptibility to fenpropathrin in resistant mites. TcCPR could be considered as a novel target for the development of new pesticides.

Uridine diphosphate (UDP)‐glycosyltransferases (UGTs) are widely distributed within living organisms and share roles in biotransformation of various lipophilic endo‐ and xenobiotics with activated UDP sugars. In this study, it was found that the activity of UGTs in abamectin‐resistant (AbR) strain was significantly higher (2.35‐fold) than that in susceptible strain (SS) ofTetranychus cinnabarinus. Further analysis showed that 5‐nitrouracil, the inhibitor of UGTs, could enhance the lethal effect of abamectin on mites. From the previous microarray results, we found an UGT gene (UGT201D3) overexpressed in AbR strain. Quantitative PCR analysis showed thatUGT201D3was highly expressed and more inducible with abamectin exposure in the AbR strain. After silencing the transcription ofUGT201D3, the activity of UGTs was decreased and the susceptibility to abamectin was increased in AbR strain whereas it was not in SS. Furthermore,UGT201D3gene was then successfully expressed inEscherichia coli. The recombinant UGT201D3 exhibited α‐naphthol activity (2.81 ± 0.43 nmol/mg protein/min), and the enzyme activity could be inhibited by abamectin (inhibitory concentration at 50%: 57.50 ± 3.54 μmol/L). High‐performance liquid chromatography analysis demonstrated that the recombinant UGT201D3 could effectively deplete abamectin (15.77% ± 3.72%) incubating with 150 μg protein for 6 h. These results provided direct evidence thatUGT201D3was involved in abamectin resistance inT. cinnabarinus.