【作者】 贺艳萍；

【导师】 马恩波；

【作者基本信息】 山西大学 ， 环境科学， 2004， 博士

【摘要】 蝗灾是世界性的农业灾害，20世纪90年代以来，受异常气候及生态环境恶化等因素的影响，我国蝗虫发生逐年加重，特别是近年北方地区持续高温干旱，蝗虫孳生环境变化，以东亚飞蝗为主的蝗灾发生面积扩大，暴发频率提高，对我国农业生产构成严重威胁。我国目前控制蝗虫危害的手段主要是采用化学药剂来降低虫口密度，化学防治虽然具有快速、高效、使用灵活等优点，但是随着农药品种及数量的增加和无限制地使用农药，会导致引起蝗虫的抗药性、农药的效能降低、农药生产成本增加等一系列问题。东亚飞蝗某些自然种群对常用杀虫剂马拉硫磷的抗性已经引起了植保部门有关人员的注意。 自从1908年首次发现美国的梨圆蚧对石硫合剂产生抗药性以来，至今至少有600多种昆虫及螨已产生了抗药性。害虫对杀虫剂产生抗性的主要机制有代谢抗性和靶标抗性。代谢抗性是指对杀虫剂起水解或隔离作用的酶发生改变，从而对杀虫剂起到解毒作用，昆虫主要解毒酶系有酯酶、谷胱甘肽-S-转移酶、细胞色素P450单加氧酶等，这三种酶系任何一种的组成部分发生改变均会引起害虫抗性的改变；靶标抗性是指由于杀虫剂作用靶标敏感度降低而产生的抗性。杀虫剂的作用靶标主要有：有机磷和氨基甲酸酯杀虫剂的作用靶标—乙酰胆碱酯酶；DDT和拟除虫菊酯类杀虫剂的作用靶标—神经轴突钠离子通道；环戊二烯类和吡唑类等杀虫剂的作用靶标—γ-氨基丁酸（GABA）受体-氯离子通道复合体。这两种抗性机理在果蝇、家蝇、棉铃虫、小菜蛾、蚊类等害虫中已经有深入的研究，为其它害虫的抗性机理研究提供了丰富的资料。 本文选取我国东亚飞蝗河北黄骅、河北平山、天津北大港和山东无棣四个杀虫剂选择压力不同的东亚飞蝗自然种群进行有机磷抗性相关酶的生化特性研究。河北黄骅、天津北大港和山东无棣三个蝗区生态特征相似，均靠近渤海湾，属于滨海蝗区，河北平山蝗区位于岗南水库库区，属于河泛蝗区。其中河北黄骅和天津北大港两个蝗区为我国重点蝗区，这两个东亚飞蝗种群的农药选择压力较大，河北平山和山东无棣种群较上述两个种群农药选择压力小。通过马拉硫磷对这四个东亚飞蝗种群的生物测定，结果表明，山东无棣和河北平山种群对马拉硫磷的敏感性较高，而天津北大港和河北黄骅种群的较低。天津北大港、河北黄骅及河北平山三种群对马拉硫磷的LD₅₀值分别是山东无棣种群的1.96、1.8及1.02倍，这说明，我国重点蝗区杀虫剂的选择压力已经造成当地东亚飞蝗种群对常用杀虫剂的敏感性下降。 对上述四个东亚飞蝗种群酷酶的生化特性进行了比较研究。东亚飞蝗中酷酶活性主要集中在胸部和腹部，水解a^NA的酷酶活性在雌性东亚飞蝗中分别有46.1%和36.1%集中在胸部和腹部，在雄性中分别为42.7%和36%。酉旨酶动力学研究表明，东亚飞蝗中酷酶对a一A、a一B、p一A及p一A等四种模式底物均有水解活性，其中a一NA为最适底物，而对p一A的水解活性较低。用对氧磷、马拉氧磷、西维因及毒扁豆碱等四种抑制剂对这四个东亚飞蝗种群的酷酶进行体外抑制实验，结果表明，四个种群中大部分酷酶属于B一型。酉旨酶活性在四个种群中有明显差异，其中山东无棣种群对a一A、a一B及p一NA三种底物的水解活性均为最低水平，在雌性种群中，河北黄骥、河北平山及天津北大港三个种群水解a一A酉旨酶的活性分别是山东无棣种群的2.09、1.28和1.24倍;水解a一NB酉旨酶的活性分别是山东无棣种群的6.30、3.79和1.61倍;水解p一A酉旨酶的活性分别是山东无棣种群的2.06、1.21和1.29倍。在雄性种群中，河北黄弊、河北平山及天津北大港三个种群水解a一A酉旨酶的活性分别是山东无棣种群的1 .71、1.53和1.51倍;水解a一NB酉旨酶的活性分别是山东无棣种群的4.71、3.34和1.55倍;水解p一A醋酶的活性分别是山东无棣种群的1.82、1.43和1.49倍。四个种群醋酶活性大小的分布统计表明，四个种群酷酶活性在个体中的分布有明显差异，河北平山种群和山东无棣种群水解三种底物的醋酶活性在低水平有分布，而河北黄弊种群则有较多个体的醋酶活性分布在较高的水平，天津北大港种群个体酷酶活性一致性较高，这可能与我们的取样有关。根据醋酶非变性聚丙烯酞胺凝胶电泳染色后，河北黄弊和天津北大港两个种群表现出相似的带型，在慢迁移率区有两条明显染色较深的带，而河北平山和山东无棣两种群中，慢迁移率区只可分辨出一条带，其它迁移较快的酷酶在四个种群间没有明显的差异，推测，天津北大港种群酷酶活性可能与河北黄弊种群处于同一水平，且酷酶活性的增加可能是由这两个迁移较慢的醋酶产生。结合四个种群对马拉硫磷的敏感性差异，分析认为，河北黄弊和天津北大港两种群中酷酶活性的提高与马拉硫磷敏感性降低有关。对江苏沛县、河南郑州、山西永济和山西临琦四个东亚飞蝗种群的64个个体（雌雄各半）以a一NA为底物进行醋酶活性检测，结果表明，山西临椅和河南郑州两个种群醋酶活性处于较低水平，而江苏沛县和山西永济两个种群醋酶活性水平较高，它们之间的酷酶活性差异与该地区蝗虫防治的用药背景相关。提出，酉旨酶活性可以更多还原

【Abstract】 The swarming locusts represent a group of prominent worldwide agriculture pests. Due to the extended high temperature, long-lasting drought and the consequent alternations in locust breeding environment the destructive outbreaks of locust had been increasing in China since 1990s, especially in the north, which seriously threaten the agriculture production. So far the chemical controls are still the major measures taken in locust abatement programs in China. In spite of the effectiveness and convenience, the frequent use of insecticides both in quantity and variety has caused a series of problems like locust resistance development, increased control difficulty and rising cost. Recently, the Chinese plant protection agencies reported the growing hardships in controlling some field populations of oriental migratory locust with organophosphate (OP) compounds.Up to now there are more than 600 arthropod species with documented resistance to one or more insecticides and/or acaricides since resistance phenomenon was first described in San Jose scale to lime-sulfur in Washington in 1908. Enhanced metabolic detoxification and reduced sensitivity of insecticide target-sites are the two major mechanisms in resistance development, involving three primary metabolic enzymes of esterases, glutathione S-transferases and cytochrome P450 polysubstrate monooxygenases. Resistance would be developed when the activities of any of these three enzymes are increased. The three insecticide target-sites are acetylcholinesterase (AChE) for organophosphate and carbamate insecticides, voltage-gated sodium channels: knockdown resistance (kdr or kdr-type) for DDT & pyrethroids and y-aminobutyric acid (GABA) receptors for cyclodienes. The resistance mechanisms have been particularly studied in fruit fly, housefly, cotton bollworm, diamond back moth and mosquitoes, which have rendered abundant references for the research in this field.This dissertation studied biochemical characters of enzymes associated to OP resistance in field populations of Locusta migratoria manilensis (Meyen) from Huanghua-Hebei, Pingshan-Hebei, Beidagang-Tianjin and Wudi-Shandong, where the insecticide selection pressures are considered different. Huanghua-Hebei, Beidagang-Tinjin and Wudi-Shandong are close to Bohai Bay, classified in sea-coast locust area with the similar ecological characteristics. Pingshan-Hebei, on the other hand is at the verge of Gangnan reservoir, classified in river-flood area. The two areas are important in locust control in China because of the frequent massive outbreaks of Locusta migratoria manilensis (Meyen). The selection pressure of OP was found higher in Huanghua-Hebei and Beidagang-Tianjin than in Pingshan-Hebei and Wudi-Shandong. LD50 values for malathion of Huanghua-Hebei and Beidagang-Tianjin are higher than those of Pingshan-Hebei and Wudi-Shandong. LD50 values for malathion of Beidagang-Tianjin, Huanghua-Hebei andPingshan-Hebei were each 1.96-, 1.8- and 1.02-fold higher than that of Wudi-Shandong.General esterases of oriental migratory locust collected from the four locations were compared. General esterases were most concentrated in the thorax and abdomen, which showed 46.1% and 36.1% of total esterase activity in females, and 42.7 and 36.0% in males, when using -NA as substrate. Kinetic studies demonstrated that general esterases in four oriental migratory locust populations hydrolyze -NA, -NB, P-NA and p-NA as substrate. Among the four substrates, -NA seemed to be the most favorable substrate for general esterases of the locust. In contrast, p-NA is not a preferred substrate for these esterases, with the lowest Vmax and the highest Km in both Huanghua-Hebei and Pingshan-Hebei populations. Inhibition studies of the esterases using four inhibitors (i. e., paraoxon, malaoxon, eserine, and carbaryl) indicated that most of general esterases in the four populations were B-type. Moreover, significant difference in general esterases activities was observed among four populations. Wudi-Shandong population更多还原