Glutathione S-transferase contributes to the resistance of Megalurothrips usitatus against lambda-cyhalothrin by strengthening its antioxidant defense mechanisms
Description
The damage caused by Megalurothrips usitatus, a common pest, has significantly affected the Chinese vegetable industry The inappropriate application of chemical pesticides has led to the development of high resistance in the M. usitatus to conventional insecticides. Glutathione S-transferase (GSTs), known for its multifunctional properties, contributes to detoxification and antioxidation. It enhances the insects’ adaptability to pesticides by facilitating the elimination of lipid peroxidation (LPO) products resulting from pyrethroid insecticides. The research employed RT-qPCR to identify GSTs genes that exhibited significant expression in response to lambda-cyhalothrin stress. It also quantified changes in antioxidant and apoptosis markers within the M. usitatus under lambda-cyhalothrin exposure. The functional significance of GSTs was validated by assessing alterations in the antioxidant defense system and resistance to lambda-cyhalothrin following the inhibition of GSTs activity. The study’s outcomes indicated that MuGSTS1 was markedly upregulated in response to lambda-cyhalothrin stress (P<0.0001). The GSTs activity was effectively suppressed by the specific inhibitor, diethyl maleate (DEM), achieving an inhibition rate of 64.05%. Following the inhibition of GSTs, the overall antioxidant capacity was reduced by 3.1-fold compared to the control, and the M. usitatus exhibited a 7.91-fold increase in sensitivity to lambda-cyhalothrin. These findings confirm the pivotal role of GSTs in the oxidative stress response of the M. usitatus and their contribution to the development of resistance to lambda-cyhalothrin through enhanced antioxidant defenses, providing valuable insights into insects’ adaptive responses to chemical stressors.
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2.1Biological Assay This research investigates the sensitivity of four M. usitatus populations (IN, SY, WZS, YN) to lambda-cyhalothrin across different geographical areas. The biological assay employed is a modification of the TIBS (Thrips Insecticide Bioassay System) by the leaf-tube film method, as described by Yuan (2023). The insecticide is diluted using 1% Triton X-100 (Sigma, USA) to five concentrations: 5000 mg/L, 2500 mg/L, 500 mg/L, 100 mg/L, and 20 mg/L, with a control group consisting of 1% Triton X-100. Each concentration level is replicated thrice. The 96.3% lambda-cyhalothrin is obtained from Beijing Yingtai Jiahe Biotechnology Co., Ltd. in China. 2.2Analysis of the GSTs Gene Expression Pattern. In accordance with the outcomes of the biological assays, the IN population of M. usitatus was subjected to lambda-cyhalothrin at the concentrations of LC25, LC50, and LC75. After a 48-hour exposure period, samples were rapidly frozen in liquid nitrogen and preserved at -80℃ for future analysis, with three replicates for each treatment condition. GSTs genes exhibiting the most marked differential expression in response to lambda-cyhalothrin stress were identified using RT-qPCR. 2.3Measuring the activities of antioxidant and detoxifying enzymes. The M. usitatus were exposed to lambda-cyhalothrin at concentrations of LC25, LC50, and LC75. After 48 h, samples were collected, and enzyme extracts were prepared to determine the changes in activities of antioxidant and detoxifying enzymes. GSTs activity was evaluated in field populations of M. usitatus from different regions. Control groups (CK) included untreated and indoor populations of M. usitatus. 2.4Determination of oxidative stress indicators The M. usitatus were subjected to lambda-cyhalothrin at concentrations of LC25, LC50, and LC75. The levels of H2O2 (BC3595), total antioxidant capacity (T-AOC, BC1315), and malondialdehyde (MDA, BC0025) content were then determined. All necessary kits were procured from Solarbio Science & Technology Co., Ltd. in Beijing. A control group (Control, CK) consisting of untreated M. usitatus was also included for comparison. 2.5Determination of apoptosis indicators The M. usitatus treated with lambda-cyhalothrin were subjected to measurements of adenosine triphosphate (ATP, JM-011512), cytochrome C content (Cytc, JM-1216602), Caspase-3 (Cas-3) activity (JM-1213802), and glucose-6-phosphate dehydrogenease (G6PD) activity (JM-006002). All necessary kits were obtained from Jiangsu Jingmei Biotechnology Co., Ltd. A control group (Control, CK) consisting of untreated M. usitatus was also included for comparison.