小菜蛾有机磷抗性季节性变化及毒理机制研究

Seasonal dynamics of the resistance to organophosphorus insecticides and its biochemical mechanism in Plutella xylostella (L.)

1998～ 2 0 0 3年期间的田间监测结果显示 ,福州地区小菜蛾 Plutella xylostella对甲胺磷和敌敌畏的抗性水平呈季节性变化 ,每年春秋季的抗性水平显著高于夏季 7月期间的抗性水平。与 2 5℃ 2 d的饲养条件相比 ,35℃ 2 d和 5℃ 30 d可使小菜蛾羧酸酯酶 (Car E)和谷胱甘肽 - S-转移酶 (GST)活力显著抑制 ,35℃ 2 d还可使小菜蛾乙酰胆碱酯酶 (ACh E)活力显著抑制 ,但 5℃4 d对小菜蛾 ACh E、Car E和 GST活力影响差异不显著。当反应温度均为 2 5℃时 ,在 35℃ 2 d、2 5℃ 2 d和 5℃ 30 d下饲养的小菜蛾 ACh E对甲胺磷和克百威的敏感性 (K i值 )没有明显变化 ,但酶与杀虫剂在 37℃的反应温度下 ,甲胺磷、敌敌畏和克百威对在 2 5℃下饲养的小菜蛾 ACh E的抑制作用显著增高。上述结果表明环境温度会影响小菜蛾的解毒代谢及杀虫剂对 ACh E的抑制作用。探讨了环境温度的季节性变化对田间小菜蛾有机磷抗性水平影响的毒理学机制。

The resistance to methamidophos and dichlorvos in the field populations of Plutella xylostella (Lepidoptera: Yponomeutidae), collected from Fuzhou, Fujian, China from October 1998 to January 2003, was determined using a leaf-dipping method with 3rd instar larvae. P. xylostella's resistance to insecticides, methamidophos and dichlorvos, was significantly higher in autumn and spring than in summer. Apparently, seasonal variations of the insecticide resistance existed in the field population of Plutella xylostella. Therefore, the effects of temperature on the activities of acetylcholinesterase (AChE), carboxyesterase (CarE) and glutathione-S-transferase (GST), and the bimolecular rate constant ( Ki ) of AChE in the larvae of P. xylostella were investigated.The AChE activity was determined with acethythiochloine (ATCh) iodide as a substrate in the presence of 5,5′-dithiobis (2-nitrobenzoic acid) (DTNB) in the pH 7.8 phosphate buffer at 25 ℃. The CarE activity was measured with 1-naphthyl acetate (α-NA) as a substrate in the pH 7.8 phosphate buffer containing eserine at 37 ℃. The GST activity was measured with 1-chloro-2,4-dinitrobenzene (CDNB) as a substrate in the pH 7.4 phosphate buffer at 25 ℃. To determine the Ki of AChE, AChE solutions were mixed with insecticides and incubated at various constant temperatures in a water bath. At different time periods, the incubated solutions were sampled for residual enzymatic activities in an ATCh solution. All experiments were carried out in triplicate, in the presence or absence of the insecticides. In order to study the temperature effects of AChE, CarE and GST on the enzymatic activities, P. xylostella larvae were reared in an environmental chamber at 35 ℃ and 25 ℃ for 2 days, and at 5 ℃ for 4 and 30 days. Then, the 4th instar larva was used for the determination of AChE, CarE and GST activities. The biochemical analysis for the larvae with the different rearing temperatures was conducted simultaneously. In comparison with the larvae reared at 25 ℃ for 2 days, the activities of CarE and GST were significantly inhibited when reared at 35 ℃ for 2 days or 5 ℃ for 30 days. The AChE activity was also considerably inhibited when reared at 35 ℃ for 2 days. There was no apparent activity inhibition on the three enzymes observed when the P. xylostella larvae were reared at 5 ℃ for 4 days. P. xylostella larvae were also reared in an environmental chamber at 35 ℃ and 25 ℃ for 2 days and 5 ℃ for 30 days to study the rearing temperature on the Ki value of AChE. The 4th instar larva was subsequently used for the Ki determination. The enzyme and insecticide mixtures were incubated at 25 ℃. No significant effect on the Ki of AChE to the methamidophos and carbofuran in P. xylostella larvae was found. Under the constant 25 ℃ rearing temperature, the incubating temperature was found to significantly affect the Ki of AChE of P. xylostella larvae to the insecticides. The Ki values to methamidophos, dichlorvos and carbofuran at 37 ℃ were 1.33, 1.60 and 1.75 times higher than those incubated at 25 ℃, respectively. The results suggested that the environmental temperature could affect the detoxifying metabolism and the inhibition of AChE in P. xylostella . The effects of the seasonal variation of temperature on the resistance to organophosphates and its toxicological mechanism in P. xylostella were discussed.