甜菜夜蛾抗氯氟氰菊酯品系相对适合度、抗性生化机理及抗性遗传方式

比较了甜菜夜蛾Spodoptera exigua 抗氯氟氰菊酯品系和敏感品系的繁殖和生长发育特征。结果表明：抗性品系幼虫发育历期延长、蛹重减轻、化蛹率和产卵量降低,抗性品系的适合度为0.61,抗性品系在繁殖和生长发育上存在明显的生存劣势。用两品系3龄幼虫分别测定胡椒基丁醚（PBO）、增效磷SV₁）、脱叶磷（DEF）和顺丁烯二酸二乙酯（DEM）对氯氟氰菊酯的增效作用,抗性品系增效倍数与敏感品系增效倍数之比分别为14.1、14.8、2.3和2.3倍,胡椒基丁醚和增效磷对氯氟氰菊酯增效作用最明显,表明多功能氧化酶参与了甜菜夜蛾对氯氟氰菊酯的抗性。抗性品系3龄幼虫酯酶和谷胱甘肽S-转移酶的活性分别为敏感品系的1.05倍和0.91倍, 抗性品系5龄幼虫多功能氧化酶O-脱甲基活性为敏感品系的1.05倍,两品系间3种酶的活性差异不显著,表明甜菜夜蛾对氯氟氰菊酯的抗性与酯酶、谷胱甘肽S-转移酶及多功能氧化酶O-脱甲基酶活性无关。用剂量对数死亡机率值回归线分析法研究甜菜夜蛾对氯氟氰菊酯的抗性遗传规律,表明甜菜夜蛾对氯氟氰菊酯的抗性为常染色体遗传、多基因控制;正、反交后代的显性度分别为0.61和0.43,抗性遗传为不完全显性。     

斜纹夜蛾抗药性监测及茚虫威对其解毒代谢酶的影响

【目的】斜纹夜蛾Spodoptera litura (Fabricius)是主要的农业害虫之一。本研究旨在明确该害虫在湖南省5个主要蔬菜种植区的抗药性水平,并探讨该害虫对茚虫威的抗性与解毒代谢酶活性之间的关系,为斜纹夜蛾有效防控及抗性治理提供依据。【方法】采用浸叶法测定了2014-2016年湖南5地斜纹夜蛾田间种群对10种杀虫剂的抗性水平;将斜纹夜蛾敏感种群3龄幼虫在死亡率40%~70%的选择压下用茚虫威进行汰选,比较了斜纹夜蛾敏感种群和抗茚虫威种群的羧酸酯酶、谷胱甘肽S-转移酶和多功能氧化酶对硝基苯甲醚O-脱甲基活性。【结果】湖南5地斜纹夜蛾田间种群对有机磷类杀虫剂产生了26.9~220.2倍的抗性,对氨基甲酸酯类杀虫剂产生了68.3~890.8倍的抗性,对拟除虫菊酯类杀虫剂产生了21.0~267.2倍的抗性,对相对较新型杀虫剂（甲维盐、阿维菌素、茚虫威和溴虫腈）产生了5.2~53.4倍的抗性。经茚虫威汰选后第14代[抗性倍数(resistance ratio, RR)=26.43]斜纹夜蛾羧酸酯酶、谷胱甘肽S-转移酶和对硝基苯甲醚O-脱甲基酶活性分别上升2.86, 1.01和1.83倍。【结论】斜纹夜蛾对多种药剂产生了不同水平的抗性,斜纹夜蛾幼虫羧酸酯酶和对硝基苯甲醚O-脱甲基活性增强可能是斜纹夜蛾对茚虫威的抗性上升的重要因素。     

二斑叶螨对阿维菌素、哒螨灵和甲氰菊酯的抗性选育及其解毒酶活力变化

在室内模拟田间药剂的选择压力,用阿维菌素、哒螨灵和甲氰菊酯对二斑叶螨Tetranychuc urticae逐代处理,以选育其抗性种群。选育至12代,对阿维菌素抗性增长到6.72倍,对哒螨灵抗性增长到12.1倍,对甲氰菊酯抗性增长到19.9倍。酶抑制剂和离体酶活性的测定结果表明,阿维菌素抗性种群的多功能氧化酶和谷胱甘肽S-转移酶的活性均有所提高;二斑叶螨对哒螨灵的抗性可能与多功能氧化酶、羧酸酯酶的活性增强有关;而羧酸酯酶、多功能氧化酶和谷胱甘肽S-转移酶活性的增强可能是二斑叶螨对甲氰菊酯产生抗性的主要原因。     

甜菜夜蛾田间种群对甲氨基阿维菌素苯甲酸盐和高效氯氰菊酯的代谢抗性机制

甜菜夜蛾Spodoptera exigua(Hübner)云南晋宁、上海奉贤和江苏六合种群对甲氨基阿维菌素苯甲酸盐抗性为45～437倍,对高效氯氰菊酯抗性为211～555倍,对其它药剂抗性不明显。这3个田间种群3龄幼虫多功能氧化酶、谷胱甘肽S-转移酶和酯酶的活力分别为室内敏感种群的2.7～8.4倍、1.9～8.6倍和1.6～5.7倍。多功能氧化酶抑制剂PBO和酯酶抑制剂DEF对甲氨基阿维菌素苯甲酸盐的增效比为1.2～4.3和1.3～7.7;PBO和DEF对高效氯氰菊酯的增效比为1.8～58和3.6～245;谷胱甘肽S-转移酶抑制剂DEM对这2种药剂均无增效作用。上述结果表明,解毒代谢增强可能是甜菜夜蛾田间种群对甲氨基阿维菌素苯甲酸盐和高效氯氰菊酯的重要抗性机理,与酯酶和多功能氧化酶活性升高有关,与谷胱甘肽S-转移酶活性升高无关。本文的研究结果还表明,对于代谢抗性机理复杂的多抗性田间种群,根据不同解毒酶抑制剂对药剂的增效作用判断不同解毒代谢酶在抗性形成中的作用更加可靠。     

桔全爪螨的抗药性选育及其解毒酶活力变化

在室内模拟田间药剂的选择压力,用齐墩螨素和甲氰菊酯对桔全爪螨Panonychus citri 逐代处理,以选育其抗药性种群。选育至12代,对齐墩螨素抗性增长到7.30倍,对甲氰菊酯抗性增长到17.11倍。增效剂和离体酶活性的测定结果表明,抗性种群的乙酰胆碱酯酶和多功能氧化酶的活性都提高了;推测羧酸酯酶、乙酰胆碱酯酶、多功能氧化酶和谷胱甘肽-S-转移酶比活力的增加是桔全爪螨对甲氰菊酯产生抗性的主要原因。     

甜菜夜蛾抗高效氯氟氰菊酯近等基因系的构建

将甜菜夜蛾Spodoptera exigua抗性种群雄成虫与敏感种群雌成虫杂交,杂交后代再自交后,用高效氯氟氰菊酯杀死敏感和部分杂合个体的剂量(50 μg/mL)处理4龄幼虫,对1天后存活的幼虫再用250 μg/mL汰选,将存活幼虫发育成的雄成虫与敏感种群的雌成虫回交,再经自交和选育,如此循环6次,获得了甜菜夜蛾抗高效氯氟氰菊酯近等基因系。对该基因系构建过程中各代幼虫的抗性水平和相关生物学特性进行了监测和比较,结果表明获得的抗高效氯氟氰菊酯近等基因系抗性种群的抗药性仍保持较高水平;酯酶同工酶的电泳分析表明抗高效氯氟氰菊酯近等基因系抗性种群与敏感种群图谱相近,而与亲本抗性种群之间存在明显差异。     

小菜蛾对定虫隆的抗性生化机制初探

以定虫隆汰选的抗性指数为23.78倍抗性种群(CH-R),并以上海田间种群(SH-R,对定虫隆的抗性指数为9.16倍)作为参比种群,研究小菜蛾Plutella xylostlla(L.)对定虫隆的抗性机制,结果表明：CH—R种群多功能氧化酶O-脱甲基活力比相对敏感种群(SZ-S)提高1倍,而SH-R种群多功能氧化酶O-脱甲基活力虽有所提高,但与SZ-S种群相比差异并不显著;CH-R和SH-R种群羧酸酯酶比活力均明显高于Sz-S种群,分别为SZ-S种群的4.61和2.18倍,且CH-R种群的Km仅为SZ-S种群的1/20;酸性磷酸酯酶和碱性磷酸酯酶比活力种群间无明显差异;CH-R种群的几丁质酶活力降低48%,酚氧化酶活力降低60%。说明小菜蛾对定虫隆的抗性机制具有多因子性,多功能氧化酶解毒代谢能力的提高可能是主导抗性机制之一,羧酸酯酶、几丁质酶和酚氧化酶也参与了小菜蛾对定虫隆的抗性。     

甜菜夜蛾对甲氧虫酰肼抗药性选育及其抗性生化机理初探

在室内用甲氧虫酰肼对甜菜夜蛾Spodoptera exigua(Hübner)幼虫进行抗药性选育,经过13代汰选,甜菜夜蛾抗性种群(Met-R)对甲氧虫酰肼的抗性倍数为4.19倍;交互抗性测定发现,Met-R甜菜夜蛾对毒死蜱、高效氯氰菊酯、虫螨腈、虫酰肼、甲胺基阿维菌素苯甲酸盐等药剂的交互抗性比值在1.05～2.07之间,不存在交互抗性。离体酶活性测定结果表明,Met-R甜菜夜蛾5龄幼虫中肠谷胱甘肽S-转移酶比活力是相对敏感种群(SS)甜菜夜蛾的2.44倍,存在显著性差异,说明谷胱甘肽S-转移酶活性增强可能与甜菜夜蛾对甲氧虫酰肼产生抗药性有关;但Met-R甜菜夜蛾全酯酶催化活性与SS的无显著性差异,说明其抗药性可能与酯酶无关。     

甜菜夜蛾不同世代对氯氟氰菊酯抗性减退及多功能氧化酶系活性变化

在室内用人工饲料连续饲养和未用药剂筛选条件下,测定了采自田间对氯氟氰菊酯产生高水平抗性甜菜夜蛾Spodoptera exigua (Hübner)种群的抗药性及其多功能氧化酶系活性的变化情况。用点滴法测定不同世代3龄幼虫抗性结果为：室内F₁代LD₅₀值为 0.9672 μg/头,抗性倍数为4 836.0倍,以后各世代逐渐降低,至F₄₃代LD₅₀值为0.0325μg/头,抗性倍数为162.5倍,抗性水平下降了29.8倍。用浸叶法测定不同世代3龄幼虫抗性结果为：室内F₁代LC₅₀值为185.6 mg/L,抗性倍数为964.7倍,以后各世代也逐渐降低,至F₄₃代LC₅₀值为9.2 mg/L,抗性倍数为47.8倍,抗性水平下降了20.2倍。与敏感品系相比,该田间种群室内饲养至F₄₃代仍处于较高的抗性水平,抗性减退缓慢,很难恢复到敏感水平。测定甜菜夜蛾田间种群室内F₂、F₂₀和F₄₁代及敏感品系5龄幼虫中肠微粒体甲氧试卤灵-O-脱甲基酶、乙氧试卤灵-O-脱乙基酶、芳香基羟基化酶及艾氏剂环氧化酶活性,结果表明：与敏感品系相比,田间种群甲氧试卤灵-O-脱甲基酶和艾氏剂环氧化酶的活性仅F₂代显著较高,F₂₀和F₄₁代差异不显著,乙氧试卤灵-O-脱乙基酶和芳香基羟基化酶的活性F₂、F₂₀和F₄₁代均显著较高。结果提示甜菜夜蛾抗性水平可能与其体内微粒体多功能氧化酶系活性有密切关系。     

棉铃虫抗药性的生理生化机制研究

本文报道了棉铃虫Helicoverpa armigera田间抗性种群对杀虫剂抗药性的生理生化机制。抗性种群(HJ-R)5龄幼虫羧酸酯酶、谷胱甘肽转移酶、多功能氧化酶活力均明显高于相对敏感种群(HD-S)。两种群乙酰胆碱酯酶对杀虫剂敏感性没有显著差异。HJ-R种群的腹神经索对氰戊菊酯表现了2-3倍的神经不敏感性。HJ-R种群对氨基甲酸酯类杀虫剂的抗性主要是由代谢机制引起,其中多功能氧化酶可能起主导作用;对菊酯的抗性是由多功能氧化酶、酯酶、以及神经不敏感性几个因子综合作用的结果。     

西花蓟马抗辛硫磷种群的抗性机制及交互抗性

为明确西花蓟马对辛硫磷的抗性风险,研究了西花蓟马抗辛硫磷种群对其他杀虫剂的交互抗性及其对辛硫磷的抗性机制.交互抗性测定结果表明,西花蓟马抗辛硫磷种群对辛硫磷与毒死蜱、高效氯氟氰菊酯和灭多威存在中等水平的交互抗性,对溴虫腈、吡虫啉、甲维盐和多杀菌素存在低水平交互抗性,对啶虫脒和阿维菌素不存在交互抗性.酶抑制剂与辛硫磷的增效剂测定结果表明,胡椒基丁醚(PBO)、三丁基三硫磷酸酯(DEF)和磷酸三苯酯(TPP)对西花蓟马抗辛硫磷种群(XK)、田间种群(BJ)和敏感种群(S)均起到了显著的增效作用(P＜0.05),马来酸二乙酯对西花蓟马抗辛硫磷种群和敏感种群增效作用均不显著,但对田间种群增效作用显著(P＜0.05).生化测定发现:除田间种群西花蓟马乙酰胆碱酯酶活性提高不显著外,西花蓟马抗辛硫磷种群和田间种群的细胞色素P450含量(2.79和1.48倍)、细胞色素b5含量(2.88和1.88倍)及O-脱甲基酶活性(2.60和1.68倍)、羧酸酯酶活性(2.02和1.61倍)和乙酰胆碱酯酶活性(3.10倍)均显著高于敏感种群(P＜0.05);谷胱甘肽-S-转移酶酶活性也有一定程度提高(1.11和1.20倍),但不显著(P＞0.05).表明其体内解毒代谢酶和靶标酶活性提高是西花蓟马对辛硫磷产生抗性的重要原因.     

二斑叶螨对甲氰菊酯的抗性选育及解毒酶活力变化

为了明确二斑叶螨Tetranychus urticae Koch对甲氰菊酯产生抗性的机理,在室内模拟田间药剂的选择压力, 用甲氰菊酯对二斑叶螨敏感品系（S）进行逐代汰选,选育至38代时, 获得了抗性倍数( resistance ratio, RR)为247.35的抗甲氰菊酯品系（Fe-R）。对S和Fe-R解毒酶活性的分析表明,Fe-R₃₈体内羧酸酯酶（carboxylesterase, CarE）、酸性磷酸酯酶(acid phosphatase, ACP)、 碱性磷酸酯酶(alkaline phosphatase, ALP)、谷胱甘肽-S-转移酶(glutathione s-transferase, GSTs)和多功能氧化酶(mixed function oxidase, MFO)较S体内相应酶的活力显著升高（P< 0.05）,其相对比值（R/S）分别为1.822,13.941,3.789,4.262和17.386。此外,筛选至第9,19,25,32代时,除Fe-R₂₅和Fe-R₃₂的MFO活性与S相比有显著性差异（P< 0.05）外,其余解毒酶（CarE,ACP,ALP,GSTs）的活性与S相比均无显著性差异(P>0.05)。筛选至第38代时, 5种解毒酶的活力与S相比均差异显著(P<0.05)。结果说明二斑叶螨Fe-R随着筛选代数的增加（第25代后）,MFO活性的上升可能是二斑叶螨对甲氰菊酯产生抗性的主要原因。     

Stability of the resistance to lambda-cyhalothrin in the ladybird beetle Eriopis connexa

Natural enemies resistant to insecticides are expected to help control the remaining arthropod pests after insecticide application and, hence, prevent crop damage, pest resurgence, and resistance selection. Field-evolved resistance to lambda-cyhalothrin and to other pyrethroids exhibited by the neotropical ladybird beetle Eriopis connexa (Germar) (Coleoptera: Coccinellidae) has been characterized and enhanced under laboratory selection. In this study, we investigated the stability of the resistance and its relationship with detoxification enzyme activity and biological performance, which are important for a biocontrol agent. One subgroup of the resistant population of E. connexa was established without selection pressure (R-UNSEL) during eight generations, and compared to either a susceptible (SUS) or its parental resistant (R-SEL) population. The resistance ratio in R-UNSEL was reduced by 50% compared to R-SEL between the first and fourth generation without selection pressure, but stayed stable afterwards, from the fifth to the eighth generation. Despite eight generations without selection pressure and reduction in the resistance level, the resistance ratio in R-UNSEL was still 39× greater than in the SUS population. The reduced resistance in R-UNSEL correlated to reduction in esterase activity, but the R-UNSEL maintained greater activity than the SUS group. The absence of selection pressure and reduction in enzyme activity in R-UNSEL did not mitigate the adaptive costs, with 2.7× lower egg production compared to SUS females. These findings indicate that resistance to lambda-cyhalothrin in R-UNSEL has already stabilized in the population. Although the resistance ratio and detoxifying enzymes were reduced in R-UNSEL, the impact on fecundity was maintained. Furthermore, the absence of crossing with wild SUS individuals will allow the R-UNSEL offspring to retain the resistance allowing survival to lambda-cyhalothrin even when used at the highest recommended field rate.     

阿维菌素对小菜蛾的抗性选育及其对解毒酶活性的影响

用阿维菌素对小菜蛾Plutella xylostella (L.)进行了抗性选育,并对选育过程中小菜蛾解毒酶的活性进行了研究。选育从F₀至F₂₁代,抗性缓慢波动上升,达到选育前的122.91倍;F₂₁至F₂₇代,抗性迅速增长,达到选育前的812.73倍,抗性发展趋势呈现S型曲线。随着选育代数的增加,对乙酰胆碱酯酶(AChE)没有明显的影响;羧酸酯酶(CarE)活性,F₂₇是F₀的1.5倍,从F₂₂开始,活性在较高水平上波动;谷胱甘肽转移酶(GST)活性F₂₇是F₀的2.2倍,且从F₁₈开始,活性在较高水平上波动。选育的抗性品系,增效醚对阿维菌素增效6.34倍。     

二化螟抗杀虫单和甲胺磷品系的生化特性

二化螟抗性和敏感品系分别采自浙江和安徽太湖。毒力测定结果表明,抗性品系对杀虫单和甲胺磷分别产生了37.7和52.7倍的抗性。代谢酶活性的测定结果显示,抗性品系多功能氧化酶氧脱甲基活性、氮脱甲基活性分别是敏感品系的3.3和1.34倍,而羧酸酯酶活性和谷胱甘肽转移酶活性两个品系之间没有显著差异。说明多功能氧化酶活性提高可能是二化螟对甲胺磷、杀虫单抗性的一个重要机制。为了研究二化螟可能存在的对沙蚕毒素杀虫剂靶标不敏感机制,采用RT-PCR 等分子生物学技术,分别对5个敏感个体和4抗性个体中克隆杀虫单作用的靶标烟碱型乙酰胆碱受体α1亚基（nAChR α subunit 1）cDNA序列进行了分子克隆。序列比较发现一共存在33个单核苷酸的多态性,其中14个引起了编码氨基酸的改变。但是没有发现与抗性有关的特有的碱基突变。     

Effect of entomopathogenic fungi on detoxification enzyme activity in greater wax moth Galleria mellonella L. (Lepidoptera, Pyralidae) and role of detoxification enzymes in development of insect resistance to entomopathogenic fungi

Fungal infection of insects increases total esterase and glutathione S-transferase activities in the hemolymph. Activities of acid and alkaline phosphatases were similar in the infected and intact insects. Fungal infection increased the resistance of greater wax moth caterpillars to organophosphorus insecticide malathion 1.46 times relative to intact caterpillars. Possible involvement of detoxification enzymes in the development of insect resistance to entomopathogenic fungi and development of complex biological products based on entomopathogenic microorganisms and inhibitors of detoxification enzymes are discussed.     

Host plant-induced changes in detoxification enzymes and susceptibility to pesticides in the twospotted spider mite (Acari: Tetranychidae)

Adult female twospotted spider mites, Tetranychus urticae Koch, reared on lima bean plants were moved to cucumber, maize, or new lima bean plants (the latter being a control) and evaluated after 24 h or 7 d for changes in susceptibility to three pesticides and in levels of related detoxification enzymes. The largest and most consistent changes were observed in mites feeding on cucumber. Susceptibility of mites on cucumber to the synthetic pyrethroids bifenthrin and lambda-cyhalothrin was greater than that of mites reared on lima bean and maize after only 24 h on the plants, and remained higher after 7 d. Mites on cucumber also were more susceptible to the organophosphate dimethoate than were mites on lima bean, but only after 7 d on the host. Susceptibility was inversely related to activities of both general esterase and glutathione S-transferase (GST) in mites on cucumber; general esterase and GST activities were 60 and 25% lower, respectively, than activities of twospotted spider mite on lima bean after 7 d of feeding. Mites on maize were slightly but significantly more susceptible than those on lima bean to bifenthrin, but not to lambda-cyhalothrin, after 7 d and to dimethoate after 24 h but not after 7 d. General esterase and GST activities in twospotted spider mite fed on maize for 24 h were 20 and 16% higher, respectively, than activities in twospotted spider mite on lima bean, but general esterase activity was 30% lower than lima bean-fed mites and GST was not different after 7 d. Thus, plant-induced changes in general esterase activity, perhaps in combination with GST activity, in twospotted spider mite appear to be inversely related to, and possibly responsible for, changes in susceptibility of twospotted spider mite to several pesticides, particularly the synthetic pyrethroids. General esterases appear to play less of a role in the detoxification of the organophosphate insecticide dimethoate.