解毒酶和转运蛋白介导的害虫抗药性分子机制研究进展

害虫抗药性是导致杀虫剂防效降低的一个重要因素,而抗性机制的阐明是害虫抗药性综合治理的基础。研究表明,代谢能力增强是害虫抗药性产生的重要原因,害虫对杀虫剂等外源物质的代谢需要细胞色素P450酶系(P450s)、羧酸酯酶(CarEs)、谷胱甘肽S-转移酶(GSTs)、UDP-葡萄糖醛酸转移酶(UGTs)和ATP结合盒转运蛋白(简称ABC转运蛋白)等解毒酶和转运蛋白的参与。结合近年来对害虫抗药性分子机制的研究进展,本文综述了上述解毒酶和转运蛋白参与杀虫剂抗药性的分子机制,并对害虫抗药性治理的新方法进行了展望。     

有机磷类杀虫剂代谢机制研究进展

文章对有机磷类杀虫剂代谢机制的研究进展以及昆虫对此类杀虫剂的相关代谢抗性机制进行了总结,阐述了有机磷杀虫剂的生物代谢途径及相关代谢酶系。在生物体中,有机磷类杀虫剂主要发生氧化代谢、水解代谢和轭合代谢等反应。其氧化代谢主要在细胞色素P450酶系(P450s)的催化作用下进行,其中,最重要的氧化反应是硫代有机磷酸酯类杀虫剂氧化脱硫形成生物毒性更高的有机磷氧化物的反应,以及氧化脱芳(烷)基化的反应;有机磷杀虫剂及其氧化产物在生物体内还可发生水解代谢反应,在对氧磷酶PON1等磷酸三酯酶的催化作用下,水解生成低毒性或者无毒的代谢物;有机磷杀虫剂的轭合代谢主要是在谷胱甘肽硫转移酶(GSTs)的催化下进行的。昆虫对有机磷类杀虫剂的代谢抗性与昆虫中参与此类杀虫剂代谢的解毒酶的改变密切相关,其中,与有机磷类杀虫剂代谢相关的P450s基因的过量表达和酶活性增强、丝氨酸水解酯酶的过量表达及基因突变、GSTs基因的过量表达等,均可导致铜绿蝇Lucilia cuprina、桃蚜Myzus persicae等昆虫对二嗪磷和马拉硫磷等有机磷类杀虫剂的代谢抗性。明确有机磷类杀虫剂的结构特点、代谢途径以及昆虫对此类杀虫剂的代谢抗性机制,对掌握有机磷类杀虫剂的毒理学机制,安全有效地使用此类杀虫剂,有效治理害虫对有机磷类杀虫剂的抗药性,以及开发生物选择性好的新型有机磷类杀虫剂,均具有重要意义。     

昆虫对拟除虫菊酯类杀虫剂的代谢抗性机制研究进展

随着拟除虫菊酯类杀虫剂在卫生和农业害虫防治中的广泛应用,昆虫对此类杀虫剂产生抗性的报道越来越多。目前已明确昆虫对拟除虫菊酯类杀虫剂的抗性机制包括表皮穿透率下降、靶标抗性以及代谢抗性,其中代谢抗性机制较为普遍,而且其与昆虫对多种杀虫剂的交互抗性关系密切。目前,随着基因组、转录组以及蛋白质组学等新技术的发展及应用,昆虫对拟除虫菊酯类杀虫剂的代谢抗性机制研究也取得了很多新进展。昆虫体内细胞色素P450酶（P450s）、羧酸酯酶（CarE）及谷胱甘肽S-转移酶（GSTs）等重要解毒酶系的改变均与昆虫对拟除虫菊酯类杀虫剂的代谢抗性有关,其中这3类解毒酶的活性及相关基因表达量的变化是昆虫对此类杀虫剂产生代谢抗性的主要原因。明确昆虫对拟除虫菊酯类杀虫剂的代谢抗性机制,对合理使用此类杀虫剂及延缓抗药性的产生均具有重要意义。本文在总结拟除虫菊酯类杀虫剂代谢路径及相关生物酶研究概况的基础上,综述了近年来有关昆虫对此类杀虫剂代谢抗性机制研究的主要进展。     

重大实蝇类昆虫解毒代谢基因的研究进展

实蝇类昆虫种类很多,其中地中海实蝇Ceratitis capitata、橘小实蝇Bactrocera dorsalis、瓜实蝇B. cucurbitae等严重为害多类果蔬,造成巨大经济损失,是国际重要检疫性或入侵性害虫。目前,杀虫剂仍然是防治实蝇类害虫的重要手段,但是多种实蝇因已经产生抗药性而导致防治困难。在昆虫抗药性产生与发展中,解毒代谢家族基因起着十分重要的作用。本文综述了实蝇科重要经济性昆虫包括橘小实蝇、瓜实蝇、油橄榄果实蝇B. oleae、昆士兰果实蝇B. tryoni、辣椒果实蝇B. latifrons、桃果实蝇B. zonata、木瓜果实蝇B. papayae、杨桃果实蝇B. carambolae、柑橘大实蝇B. minax、地中海实蝇、苹绕实蝇Rhagoletis pomonella、雪果绕实蝇R. zephyria和泽兰始实蝇Procecidochares utilis等的细胞色素P450酶、酯酶、谷胱甘肽S转移酶、ABC转运蛋白这4类解毒代谢基因方面的研究进展,为全面深入了解研究实蝇科昆虫应对有毒有害物质的生理和遗传机制以及研发实蝇类害虫化学防治新策略、新技术等提供参...     

二化螟对双酰胺类杀虫剂抗性的分子机制研究进展

新型双酰胺类杀虫剂已广泛用于保障水稻生产,而二化螟作为危害水稻生产的钻蛀性害虫,已经对该类杀虫剂产生了抗性,明确该类杀虫剂抗性分子机制,可为二化螟抗性快速检测和绿色防控提供技术支撑。本文在总结二化螟对双酰胺类杀虫剂抗性现状的基础上,重点综述了近年来有关其抗性分子机制研究的进展,主要包括解毒酶和转运蛋白基因过表达介导的代谢抗性,以及鱼尼丁受体基因突变介导的靶标抗性;指出了该研究领域在抗性分子检测、抗性新基因鉴定、抗性基因调控网络和多重抗性机制等方面存在的问题,并展望了其发展方向,认为：利用高通量测序技术检测害虫种群抗药性;利用多组学技术鉴定新抗性基因及调控网络,以探明多重抗性机制;将反向遗传学工具放射性配基结合及电生理技术深入验证抗性基因功能;需开发靶向抗性基因的dsRNA转基因作物、纳米农药及选择性新型化学杀虫剂,以达到杀虫剂减施增效的目的。     

棉铃虫对化学杀虫剂的抗性现状及分子机制研究进展

棉铃虫Helicoverpa armigera是一种世界性分布的重大杂食性农业害虫,长期大量使用化学药剂防治棉铃虫导致其对不同种类杀虫剂产生了抗性。抗性分子机制的阐明有利于棉铃虫的科学防控和抗性治理。该文主要综述棉铃虫对化学杀虫剂的抗性发展现状,以及近年来棉铃虫抗药性分子机制的研究进展,包括解毒酶代谢能力增强、靶标敏感性降低和表皮穿透能力下降等方面,并就未来研究工作和棉铃虫抗性治理新技术进行了展望。     

棉蚜对新烟碱类杀虫剂的抗性现状及其治理策略

棉蚜Aphis gossypii Glover是棉花生产中最严重的害虫之一。长期以来防治棉蚜主要依赖于化学杀虫剂,其中新烟碱类杀虫剂扮演着十分重要的角色,但由于其长期、大量的使用,棉蚜已对该类杀虫剂产生了较高水平的抗性,严重影响了对棉蚜的防治效果。抗性机制研究表明,棉蚜对新烟碱类杀虫剂产生抗性的机制主要涉及解毒代谢能力增强和靶标敏感性下降。细胞色素P450、羧酸酯酶、谷胱甘肽S-转移酶、UDP-葡糖基转移酶等解毒酶基因过量表达介导的解毒代谢增强和烟碱型乙酰胆碱受体β1亚基突变引起的靶标敏感性下降是棉蚜对新烟碱类杀虫剂产生抗性的关键。针对我国棉蚜对新烟碱类杀虫剂抗性问题突出的现状,为更好地利用该类药剂防治棉蚜,亟需根据棉蚜抗性机制制定合理的抗性治理策略。本文将从棉蚜对新烟碱类杀虫剂的抗性现状、抗性机制以及抗性治理策略等方面进行综述,以期为抗性棉蚜的科学治理提供参考。     

灰飞虱对杀虫剂的抗性分子机制研究进展

灰飞虱是中国长江流域和黄淮地区重要的农业害虫,由于杀虫剂的广泛与大量使用,已导致其对多种杀虫剂产生了抗性。深入研究其抗药性分子机制,可为灰飞虱抗性的快速检测和治理提供重要理论基础。文章总结了灰飞虱对毒死蜱、吡虫啉、溴氰菊酯、噻嗪酮、氟虫腈和乙虫腈等杀虫剂的抗性分子机制研究进展,主要包括抗性相关解毒酶和转运蛋白基因的筛选与功能验证,以及靶标位点突变等重要研究成果,指出该研究领域当前存在的问题主要有抗性基因的功能验证及调控路径、抗性新基因的鉴定及交互抗性和多重抗性机制不明确等,并展望了其未来发展方向,认为:可利用CRISPR/Cas9基因编辑技术验证抗性基因功能;可将转录组测序结合生物信息学手段用于鉴定新抗性基因及抗性调控基因,以探明交互抗性和多重抗性机制;应深入至蛋白组学水平探讨抗性机制;需开发配套的高效田间施药技术,以达到杀虫剂减施增效的目的。     

主要农业害虫对茚虫威的抗性现状及其治理策略

茚虫威属于噁二嗪类杀虫剂,与大多数杀虫剂不同的是其进入害虫体内需要经活化代谢转变成N-去甲氧羰基代谢物(decarbomethoxylated metabolite,DCJW)后不可逆地阻断钠通道,进而发挥杀虫活性。茚虫威由于其作用机制不同于常见的使钠离子通道延迟关闭的菊酯类药剂而被广泛用于鳞翅目和一些同翅目、鞘翅目害虫的防治。抗药性是任何杀虫药剂使用后面临的问题,茚虫威也不例外,许多害虫对其产生了不同程度的抗性。昆虫对茚虫威产生抗性的机制包括酯酶活性、谷胱甘肽S-转移酶(glutathione S-transferase,GST)和P450活性的增加以及分子靶标F1845Y、V1848I、L1014P的突变,这些对茚虫威抗性机制的研究基本都是基于抗性种群和敏感种群开展的,需要进一步验证其对抗性研究的贡献度。针对我国田间害虫种群对茚虫威的抗性现状,及时实施对茚虫威有效的抗性治理是迫切的。对于茚虫威的抗性治理除了传统的杀虫药剂轮用、混用外,需要利用其作用机制特点开展抗性治理策略研究。一是充分利用其活化代谢的特点,开展组合药剂的研究应用;二是菊酯类药剂和茚虫威的作用机制均与钠离子通道有关,...     

基于全长转录组测序的稻秆潜蝇解毒代谢相关基因筛选

为筛选水稻害虫稻秆潜蝇Chlorops oryzae潜在的解毒代谢酶基因，利用PacBio Sequel Ⅱ测序平台对稻秆潜蝇幼虫进行全长转录组测序，基于测序结果筛选稻秆潜蝇的解毒代谢相关基因谷胱甘肽S-转移酶（glutathione S-transferase，GST）基因、羧酸酯酶（carboxylesterase，CarE）基因和细胞色素P450（cytochrome P450，CYP450）基因，并检测其在稻秆潜蝇不同发育阶段的相对表达量。结果表明，对稻秆潜蝇进行测序得到18 100条去冗余转录本序列，共有16 283条序列得到注释。通过比对分析共筛选出8条GST、12条CarE和28条CYP450基因序列，这些基因在稻秆潜蝇不同发育阶段的表达量具有显著差异，表明不同虫态的稻秆潜蝇其解毒代谢能力可能不同。此外，还筛选到1 452个lncRNA序列，以及176个潜在的lncRNA靶基因序列，其中4个与解毒代谢基因相关。表明筛选获得的稻秆潜蝇解毒代谢酶基因及lncRNA靶基因可用于后续该虫的潜在抗药性研究及防治药剂筛选。     

Cytochrome P450 monooxygenase-mediated permethrin resistance confers limited and larval specific cross-resistance in the southern house mosquito, Culex pipiens quinquefasciatus

The cytochrome P450-dependent monooxygenases (P450s) are an important enzymatic system that metabolizes xenobiotics (e.g., pesticides), as well as endogenous compounds (e.g., hormones). P450-mediated metabolism can result in detoxification of insecticides such as pyrethroids, or can be involved in the bioactivation and detoxification of insecticides such as organophosphates. We isolated (from the JPAL strain) a permethrin resistant strain (ISOP450) of Culex pipiens quinquefasciatus, having 1300-fold permethrin resistance using standard backcrossing procedures. ISOP450 is highly related to the susceptible lab strain (SLAB) and the high resistance to permethrin is due solely to P450-mediated detoxification. This is the first time in mosquitoes that P450 monooxygenase involvement in pyrethroid resistance has been isolated and studied without the confounding effects of kdr. Resistance in ISOP450 is incompletely dominant (D = +0.3), autosomally linked, and monofactorally inherited. It is expressed in the larvae, but not in adults. Cross-resistance to pyrethroids lacking a 3-phenoxybenzyl moiety (tetramethrin, fenfluthrin, bioallethrin, and bifenthrin) ranged from 1.5- to 12-fold. ISOP450 had only limited (6.6- and 11-fold) cross-resistance to 3-phenoxybenzyl pyrethroids with an α-cyano group (cypermethrin and deltamethrin, respectively). Examination of cross-resistance patterns to organophosphate insecticides in ISOP450 showed an 8-fold resistance to fenitrothion, while low, but significant, levels of negative cross-resistance were found for malathion (RR = 0.84), temephos (RR = 0.73), and methyl-parathion (RR = 0.55). The importance and uniqueness of this P450 mechanism in insecticide resistance is discussed.     

Insecticide Resistance in the Western Flower Thrips, Frankliniella occidentalis

The western flower thrips, Frankliniella occidentalis Pergande (Thysanoptera: Thripidae) is a serious pest on a wide range of crops throughout the world. F. occidentalis is difficult to control with insecticides because of its thigmokinetic behaviour and resistance to insecticides. Pesticide resistance can have a negative impact on integrated pest management programmes with chemical control as one of the components. Resistance to a number of different insecticides has been shown in many populations of F. occidentalis. This flower thrips has the potential of fast development of resistance owing to the short generation time, high fecundity, and a haplodiploid breeding system. The mechanisms conferring insecticide resistance in insects can be divided into four levels. First, an altered behaviour can aid the insect to avoid coming into contact with the insecticide. Second, a delayed penetration through the integument will reduce the effect of the insecticide at the target site. Third, inside the insect, detoxification enzymes may metabolise and thereby inactivate the insecticide. Fourth, the last level of resistance mechanisms is alterations at the target site for the insecticide. Knowledge of resistance mechanisms can give information and tools to be used in management of the resistance problem. Recently, studies have been carried out to investigate the underlying mechanisms conferring resistance in F. occidentalis. It appears that resistance in F. occidentalis is polyfactorial; different mechanisms can confer resistance in different populations and different mechanisms may coexist in the same population. Possible resistance mechanisms in F. occidentalis include: reduced penetration, detoxification by P450-monooxygenases, esterases and glutathione S-transferases, and alterations of acetylcholinesterase, the target site for organophosphate and carbamate insecticides. Target site resistance to pyrethroids (knockdown resistance) may also be a resistance mechanism in F. occidentalis.     

Metabolic mechanisms of insecticide resistance in the western flower thrips, Frankliniella occidentalis (Pergande)

The interactions between six insecticides (methiocarb, formetanate, acrinathrin, deltamethrin, methamidophos and endosulfan) and three potential synergists (piperonyl butoxide (PBO), S,S,S-tributyl phosphorotrithioate (DEF) and diethyl maleate (DEM)) were studied by topical exposure in strains selected for resistance to each insecticide, and in a susceptible strain of Frankliniella occidentalis (Pergande). In the susceptible strain PBO produced appreciable synergism only of formetanate, methiocarb and methamidophos. Except for endosulfan, PBO synergized all the insecticides to varying degrees in the resistant strains. A very high level of synergism by PBO was found with acrinathrin, which reduced the resistance level from 3344- to 36-fold. PBO slightly synergized the carbamates formetanate (4.6-fold) and methiocarb (3.3-fold). PBO also produced a high synergism of deltamethrin (12.5-fold) and methamidophos (14.3-fold) and completely restored susceptibility to both insecticides. DEF did not produce synergism with any insecticide in the resistant strains and DEM was slightly synergistic to endosulfan (3-fold). These studies indicate that an enhanced detoxification, mediated by cytochrome P-450 monooxygenases, is the major mechanism imparting resistance to different insecticides in F occidentalis. Implications of different mechanisms in insecticide resistance in F occidentalis are discussed.     

Resistance to insecticides in winter- and summer-forms of pear psylla,Psylla pyricola

Summer-form pear psylla, Psylla pyricola Foerster, from sprayed pear were resistant to endosulfan (2·4-fold), methiocarb (2·5-fold), ethylan (5·8-fold), azinphos-methyl (7·7-fold), and fenvalerate (40·1-fold). Esterase (3·8-fold), glutathione transferase (1·8-fold), and cytochrome P-450 monooxygenase (1·6-fold) detoxification enzyme activity was higher in resistant than in susceptible summer forms. Synergism by piperonyl butoxide and S,S,S-tributylphosphorotrithioate (DEF) was added evidence for cytochrome P-450 monooxygenases and esterases as resistance mechanisms. Reduced penetration may also have contributed to resistance, as indicated by a 1·6-fold slower penetration of azinphos-methyl in resistant than susceptible summer-forms. Similar differences in insecticide toxicity and esterase and glutathione transferase activities were observed between winter-forms of resistant and susceptible pear psylla. Winter-forms of P. pyricola were up to three times more tolerant to insecticides than summer-forms. Higher cytochrome P-450 monooxygenase activity (1·7-fold) and slower azinphosmethyl penetration (2·1-fold) in winter-forms may have contributed to their greater insecticide tolerance; however, sequestration may also have been involved.     

Insecticide resistance management strategies against the western flower thrips, Frankliniella occidentalis

Western flower thrips (WFT), Frankliniella occidentalis (Pergande), is an economically important pest of a wide range of crops grown throughout the world. Insecticide resistance has been documented in many populations of WFT. Biological and behavioural characteristics and pest management practices that promote insecticide resistance are discussed. In addition, an overview is provided of the development of insecticide resistance in F. occidentalis populations and the resistance mechanisms involved. Owing to widespread resistance to most conventional insecticides, a new approach to insecticide resistance management (IRM) of F. occidentalis is needed. The IRM strategy proposed consists of two parts. Firstly, a general strategy to minimise the use of insecticides in order to reduce selection pressure. Secondly, a strategy designed to avoid selection of resistance mechanisms, considering cross-resistance patterns and resistance mechanisms.     

Gene expression in pyriproxyfen-resistant Bemisia tabaci Q biotype

Pyriproxyfen is a biorational insecticide that acts as a juvenile hormone (JH) analogue and disrupts insect development with an unknown molecular mode of action. Pyriproxyfen is one of the major insecticides used to control the whitefly Bemisia tabaci (Gennadius) and comply with integrated pest management (IPM) programmes, resulting in minimal effects on the environment, humans and beneficial organisms. During the last few years, resistance to pyriproxyfen has been observed in several locations in Israel, sometimes reaching a thousandfold or more. No information exists about the molecular basis underlying this resistance that may lead to understanding the mode of action of pyriproxyfen and developing molecular markers for rapid monitoring of resistance outbreaks. In this communication, a cDNA microarray from B. tabaci was used to monitor changes in gene expression in a resistant B. tabaci population. Based on statistical analysis, 111 expressed sequence tags (ESTs) were identified that were differentially upregulated in the resistant strain after pyriproxyfen treatment. Many of the upregulated ESTs observed in the present study belong to families usually associated with resistance and xenobiotic detoxification such as mitochondrial genes, P450s and oxidative stress, genes associated with protein, lipid and carbohydrate metabolism and others related to JH-associated processes in insects such as oocyte and egg development.     

Field-simulator study of insecticide resistance conferred by esterase-, MACE- and kdr-based mechanisms in the peach-potato aphid,Myzus persicae (Sulzer)

Insecticide sprays were applied to Myzus persicae (Sulzer) populations carrying various combinations of three insecticide resistance mechanisms (esterase-based metabolic resistance and two target site mechanisms, known as MACE and kdr), supported on host plants growing in field simulator cages. The study showed that MACE confers extreme resistance to pirimicarb and triazamate (carbamate insecticides) but not to deltamethrin + heptenophos (16 + 1) (Decisquick) or dimethoate (an organophosphorus insecticide). Resistance to dimethoate depends solely on levels of esterase-based resistance, while resistance to Decisquick depends on kdr and esterase. None of the four insecticides is effective against aphids carrying MACE combined with extreme esterase-based resistance. This knowledge, in association with current monitoring of the mechanisms, will play an important role in making decisions on insecticide use against M persicae in the UK. © 1999 Society of Chemical Industry     

丽蝇蛹集金小蜂P450基因家族的发现和分子进化分析

丽蝇蛹集金小蜂是一类重要的天敌昆虫,大量地应用于生物防治中。虽然寄生蜂不是化学农药的直接作用靶标,但其在自然环境中可能接触到化学农药。P450基因家族是昆虫产生抗药性的重要原因之一,同时也参与了昆虫激素的合成与降解。因此,研究P450基因家族对充分利用丽蝇蛹集金小蜂,以及保护其免受农药毒杀具有重要的意义。本文利用生物学信息方法从丽蝇蛹集金小蜂基因组筛选到91个P450基因。家族分析表明,丽蝇蛹集金小蜂具有15个P450基因超家族、27个亚家族。对P450基因的正选择和基因转换开展了进一步分析,研究发现5个基因族存在正选择位点,2个基因族存在基因转换,正选择和基因转换之间存在明显的关联性。各个亚家族的选择压力分析表明,CYP9P亚家族受到选择压力最大（ω=118．9355）。通过MEME程序在丽蝇蛹集金小蜂P450基因上识别到5个频率为90％以上的motif,其中“LAE[NH]P”为新发现的motif.