二芳酰肼类昆虫生长调节剂商品化品种的特点简介

蜕皮激素(molting hormore，MH)是由昆虫前胸腺分泌的一种甾类激素，对昆虫蜕皮行为及蜕皮后的发育起调控作用。蜕皮激素及其类似物曾被考虑用作杀虫剂，但是由于其结构复杂，提取、合成成本太高；而分子体积大、亲水性强等特点又使其不易渗透昆虫表皮；而且昆虫已具有很强的分解代谢甾类结构的能力，使其迅速失去杀虫活性，上述原因使其不能实现田间应用。1988年Rohm and Hass公司的科学家们发现二芳酰肼类化合物RH-5849(图1)能引起受试昆虫早熟、不完全、致死性蜕皮，诞生了一类新的昆虫生长调节剂——蜕皮激素拮抗剂，使蜕皮激素类似物走向了实用化。     

麦红吸浆虫成虫和幼虫滞育过程中蜕皮激素的定量分析

为明确麦红吸浆虫成虫和幼虫滞育过程中体内蜕皮激素的变化规律,采用放射免疫分析法测定了麦红吸浆虫成虫和幼虫滞育过程中的蜕皮激素滴度.结果表明,同一天羽化的成虫,雌虫蜕皮激素滴度为7.43pg/头,显著高于雄虫;滞育前后及滞育期幼虫蜕皮激素滴度存在显著差异,滞育前为5.22pg/头,进入滞育后显著降低;滞育期间,冬季蜕皮激素滴度为6.64～7.10pg/头,显著高于其它季节;滞育解除后蜕皮激素滴度显著降低,之后随着幼虫的生长发育逐渐升高,至化蛹前约1周达到最高,为15.22 pg/头.滞育期间同期裸露幼虫与结茧幼虫,以及不同滞育年限幼虫的蜕皮激素滴度差异均不显著.     

Β-蜕皮激素对异色瓢虫羧酸酯酶基因表达及酶活性影响的初步研究

为了探明β-蜕皮激素对异色瓢虫的作用及其安全剂量,本研究利用实时定量PCR(RT-PCR)和比色法测定了异色瓢虫3龄幼虫施用不同浓度的β-蜕皮激素体内羧酸酯酶活性与基因表达情况.结果表明:用0.1、0.5、1 mg/L β-蜕皮激素处理异色瓢虫3龄幼虫,在处理后4～48 h内CarE mRNA水平和CarE比活力呈现先升高、保持正常水平或略下降、再升高、然后很快恢复正常的规律.到处理后的第48 h,处理组的CarE mRNA表达量和CarE比活力均恢复到正常水平.所以,认为在低于1 mg/L浓度下应用β-蜕皮激素控制害虫对异色瓢虫的3龄幼虫安全.而利用5 mg/L或10 mg/L的β-蜕皮激素处理异色瓢虫3龄幼虫,CarE mRNA表达量和CarE比活力均表现出长期的被抑制效应,特别是到处理24 h后,CarE mRNA表达量和CarE活性不仅未能恢复到正常水平,反而一直处于极低的水平.因此,认为这两个浓度对异色瓢虫3龄幼虫不安全.     

喜树碱对小菜蛾体内保幼激素及蜕皮激素滴度的影响

为明确喜树碱对小菜蛾Plutella xylostella生长发育的抑制作用及其机制,设置0.5、1.0、2.0mg/m L剂量处理其3龄幼虫,采用浸叶饲喂法和高效液相色谱法测定了喜树碱对小菜蛾体内蜕皮激素和保幼激素滴度的影响。结果显示,喜树碱各剂量处理均显著抑制了小菜蛾幼虫的生长发育,抑制率最高达到28.42%;各处理小菜蛾化蛹时间均延迟,异常蛹增多,羽化率降低;其中2.0 mg/m L处理后4 d其化蛹率为6.67%,对照组化蛹率为86.60%。0.5、1.0 mg/m L处理下,小菜蛾体内蜕皮激素滴度显著低于对照组,仅为对照组的0.06倍;而保幼激素滴度增加,分别可达到28.74mg/m L和36.29 mg/m L,是对照组的9倍左右。表明喜树碱可影响小菜蛾体内蜕皮激素和保幼激素的平衡,延迟和影响了小菜蛾的正常发育进程,可实现对小菜蛾种群的控制作用。     

昆虫生长调节剂的开发现状与发展趋势

本文简要地介绍了与环境友好，相容的昆虫生长调节剂包括几丁质抑制剂，保幼激素类似物和蜕皮激素似物的开发现状和发展趋势，昆虫生长调节剂的发展虽缓慢，但前景广阔。     

温度和外源激素对白蜡吉丁肿腿蜂翅型分化的影响

为探讨影响寄生蜂翅型分化的环境因素,促进天敌昆虫的利用,在室内比较了白蜡吉丁肿腿蜂Sclerodermus pupariae在21、24、27、30℃和33℃5个温度条件下翅型分化情况,并在24℃下通过对该蜂寄主麻竖毛天牛Thyestilla gebleri幼虫注射早熟素Ⅱ和蜕皮激素,比较不同激素处理对寄生蜂子代雌蜂有翅率的影响。研究结果表明,白蜡吉丁肿腿蜂在27℃和30℃下子代雌蜂的有翅率显著高于其他温度,分别高达62.02%和68.04%。24℃下该蜂雌蜂有翅率最低,仅为0.97%,确定24℃为该蜂的无翅型温度T_(wl)。在T_(wl)下早熟素Ⅱ处理寄主幼虫显著提高了寄生蜂子代雌蜂的有翅率(15.62%)。而用3种浓度的蜕皮激素溶液处理寄主幼虫,其育出的寄生蜂子代翅型均无显著分化,蜕皮激素对白蜡吉丁肿腿蜂的翅型分化无显著影响。在对白蜡吉丁肿腿蜂人工繁育过程中,建议将饲养温度设置为30℃来提高该蜂种群的雌蜂有翅率。结合早熟素Ⅱ具有翅化效应的结果,可以对寄主幼虫进行早熟素Ⅱ处理或者直接在幼期处理子代蜂,从而实现对该天敌资源的高效利用。     

荻草谷网蚜ABCG1基因的克隆及表达模式分析

荻草谷网蚜Sitobion miscanthi是严重威胁我国小麦生产安全的迁飞性害虫。蜕皮激素是参与蚜虫翅型分化调控的内激素, 在有翅成蚜体内保持高滴度, 且诱导后代产生更高比例的无翅蚜, 其进出靶细胞需要经过细胞膜上特定蛋白的转运。ATP结合盒转运蛋白家族G亚家族(ATP-binding cassette transporter G, ABCG)中的 ABCG1是通过跨膜转运昆虫类固醇、对蜕皮激素信号进行负调控的功能蛋白之一, 在蚜虫中尚未见报道。本文克隆了荻草谷网蚜ABCG1(SmisABCG1)基因, 并进行了序列比对、系统进化分析以及不同组织部位和发育时期表达模式分析。结果显示, SmisABCG1基因的开放阅读框全长为1 851 bp, 编码616个氨基酸, 含7个跨膜结构域, 符合ABCG蛋白家族典型结构特性, 基因登录ID:OP626323。昆虫间ABCG1较保守, 该蛋白系统进化关系与各自物种间亲缘关系的远近保持一致。其中, SmisABCG1与来自豌豆蚜、禾谷缢管蚜、棉蚜、花生蚜和雪松长足大蚜等的ABCG1氨基酸序列高度一致(>87%), 以上蚜虫聚为一支。与SmisABCG1亲缘关系最近的是豌豆蚜的ABCG1, 其次是半翅目的褐飞虱、白背飞虱和灰飞虱, 与膜翅目的新疆菜叶蜂、阿里山潜蝇茧蜂以及鞘翅目的赤拟谷盗、蜂箱小甲虫亲缘关系较远。该基因在伪胚胎和成蚜阶段高表达。包含伪胚胎的有翅、无翅成蚜整蚜SmisABCG1的转录水平无显著差异, 但其在来自有翅成蚜的伪胚胎中的转录水平高于无翅成蚜伪胚胎, 证实无翅成蚜自身的转录水平较高, 而有翅成蚜较低。进一步分析显示这一差异主要是无翅蚜胸部显著高表达所导致。基于该蛋白对蜕皮激素负调控, 与有翅成蚜转录水平低, 但蜕皮激素水平更高相符合。     

二芳酰肼类昆虫生长调节剂的构效关系研究进展

二芳酰肼类化合物是近年来开发的具有蜕皮激素活性的非甾醇类昆虫生长调节剂,能引起昆虫,特别是鳞翅目幼虫早熟、不完全、致死性蜕皮。文章总结近期文献,结合课题组的研究工作,对该类化合物中各个结构片断对其杀虫活性的影响进行了探讨,对结构与活性的关系进行了综述。     

昆虫神经内分泌研究的生物技术应用展望

昆虫的激素主要有三种,蜕皮激素、保幼激素与神经激素。另外,前列腺素、生物胺、雌性激素有时也作为昆虫激素。迄今,昆虫内分泌研究主要集中在蜕皮激素类与保幼激素类,原因是这些激素的分子相对简单,能制约蜕皮与变态这两个关键过程。打破这两个生理过程就能有效地防治害虫。不少保幼激素类似物能干扰变态与生殖,已商品化地应用于蝇、蚊、跳虱、(虫非)蠊等害虫的防治。神经激素一般是肽,因此结构变化比蜕皮激素及保幼激素大。尽管有关神经激素的资料不多,但从长远来看,其应用于害虫防治的潜力要出蜕皮激素和保幼激素大。它们是昆虫主要的调控因子,影响很多的生理过程,如保幼激素与蜕皮激素的分泌,以及生长、变态、生殖,自发生理平衡等,大约有24例研究报道,指称神经激素能调节昆虫的生长与发育、生殖、代谢与生理的自发平衡等生理过程。但仅有极少数昆虫神经激素被分离,提取,完全或部分地搞清它的化学结构。本文论述以神经内分泌为基础的害虫防治有关事项。     

Comparison of the activity of non‐steroidal ecdysone agonists between dipteran and lepidopteran insects,using cell‐based EcR reporter assays

BACKGROUND: Diacylhydrazine (DAH) analogues have been developed successfully as a new group of insect growth regulators, called ecdysone agonists or moulting accelerating compounds. These DAHs have been shown to manifest their toxicity via interaction with the ecdysone receptor (EcR) in susceptible insects, as does the natural insect moulting hormone 20‐hydroxyecdysone (20E). A notable feature is their high activity and specificity, particularly against lepidopteran insects, raising the question as to whether non‐lepidopteran‐specific analogues can be isolated. However, for the discovery of ecdysone agonists that target other important insect groups such as Diptera, efficient screening systems that are based on the activation of the EcR are needed. RESULTS: In this study, a dipteran‐specific reporter‐based screening system with transfected S2 cells of Drosophila melanogaster Meig. was developed in order to discover and evaluate compounds that have ecdysone agonistic or antagonistic activity. A library of non‐steroidal ecdysone agonists containing different mother structures with DAH and other related analogues such as acylaminoketone (AAK) and tetrahydroquinoline (THQ) was tested. None of the compounds tested was as active as 20E. This is in contrast to the very high activity of several DAH and AAK congeners in lepidopteran cells (Bombyx mori L.‐derived Bm5 cells). The latter agrees with a successful docking of a DAH, tebufenozide, in the binding pocket of the lepidopteran EcR (B. mori), while this was not the case with the dipteran EcR (D. melanogaster). Of note was the identification of two THQ compounds with activity in S2 but not in Bm5 cells. Although marked differences in activity exist with respect to the activation of EcR between dipterans and lepidopterans, there exists a positive correlation (R = 0.724) between the pLC₅₀ values in S2 and Bm5 cells. In addition, it was found through protein modelling that a second lobe was present in the ligand‐binding pocket of lepidopteran BmEcR but was lacking in the dipteran DmEcR protein, suggesting that this difference in structure of the binding pocket is a major factor for preferential activation of the lepidopteran over the dipteran receptors by DAH ligands. CONCLUSIONS: The present study confirmed the marked specificity of DAH and AAK analogues towards EcRs from lepidopteran insects. THQ compounds did not show this specificity, indicating that dipteran‐specific ecdysone‐agonist‐based insecticides based on the THQ mother structure can be developed. The differences in activity of ecdysone agonists in dipteran and lepidopteran ecdysone‐reporter‐based screening systems are discussed. Copyright © 2010 Society of Chemical Industry     

Receptor-binding affinity and larvicidal activity of tetrahydroquinoline-type ecdysone agonists against Aedes albopictus

Tetrahydroquinolines (THQs), a class of nonsteroidal ecdysone agonists, are good candidates for novel mosquito control agents because they specifically bind to mosquito ecdysone receptors (EcRs). We have recently performed quantitative structure–activity relationship (QSAR) analyses of THQs to elucidate the physicochemical properties important for the ligand–receptor interaction. Based on previous QSAR results, here, we newly synthesized 15 THQ analogs with a heteroaryl group at the acyl moiety and evaluated their binding affinity against Aedes albopictus EcRs. We also measured the larvicidal activity of the combined set of previously and newly synthesized compounds against A. albopictus to examine the contribution of receptor-binding to larvicidal activity. Multiple regression analyses showed that the binding affinity and the molecular hydrophobicity of THQs are the key determinants of their larvicidal activity.     

沉默蜕皮激素受体对雌性中黑盲蝽生殖力的影响

中黑盲蝽Adelphocoris suturalis是一种重要的农业害虫,主要为害棉花、果树和牧草等作物。昆虫保幼激素(juvenile hormone, JH)与蜕皮激素(molting hormone, 20E)是调控昆虫生殖的主要因素。本研究运用基因克隆、基因沉默(RNAi)、实时荧光定量PCR(qPCR)等技术研究蜕皮激素受体(ecdysone receptor, EcR)、超气门蛋白(ultraspiracle protein, USP)在中黑盲蝽生殖调控中的作用。结果表明,中黑盲蝽EcR基因的ORF全长1 413 bp,编码470个氨基酸。预测蛋白质分子量为53.65 kD,pI值为7.79。中黑盲蝽USP基因ORF全长1 209 bp,编码402个氨基酸。预测蛋白质分子量为44.99 kD, pI值为7.94。与对照相比,注射dsEcR、dsUSP后6～18 d的中黑盲蝽体内EcR基因、USP基因在转录水平分别被显著抑制;卵巢的挂卵量分别仅有对照组的31.5%、86.6%;单雌产卵量减少36.0%～80.4%,显著低于对照。EcR和USP基因沉默后,产卵前期与对照相比无显...     

Isoforms of the heteropteran Nezara viridula ecdysone receptor: protein characterisation, RH5992 insecticide binding and homology modelling

BACKGROUND: Certain bisacylhydrazine compounds such as tebufenozide (RH5992) have been shown to act as order‐specific insecticides. Their compatibility with predatory Heteroptera, which are used as biological control agents, has also been demonstrated. However, the molecular mode of action of these ecdysone agonists has not been explored in a heteropteran, much less one that is a significant agricultural pest, such as Nezara viridula. RESULTS: Alternatively spliced ligand‐binding regions of the N. viridula ecdysone receptor were expressed, purified and characterised by 2D gel analysis, mass spectrometry, homology modelling and competitive binding of a bisacylhydrazine insecticidal compound (RH5992) and various ecdysteroids. Ligand binding by the two splice isoforms was indistinguishable, and relative affinities were found to occur in the order muristerone A > ponasterone A > 20‐hydroxyecdysone > inokosterone > RH5992 > α‐ecdysone. CONCLUSION: The predicted difference in amino acid sequence between the ligand‐binding domains of the N. viridula ecdysone receptor splice variants was verified by mass spectrometry. Both splice variant isoforms exhibit a greater affinity for the bisacylhydrazine insecticide RH5992 than do the other hemipteran ecdysone receptors characterised to date. Their affinities for a range of ecdysteroids also distinguish them from the ecdysone receptors of other Hemiptera characterised thus far. Homology models of both N. viridula receptor isoforms provide further insight into the bisacylhydrazine‐ and ecdysteroid‐binding properties of these receptors, including their similar affinity for 20‐hydroxyecdysone and the postulated pentatomomorphan moulting hormone makisterone A. Copyright © 2011 Society of Chemical Industry     

Assessment of species specificity of moulting accelerating compounds in Lepidoptera: comparison of activity between Bombyx mori and Spodoptera littoralis by in vitro reporter and in vivo toxicity assays

BACKGROUND: Dibenzoylhydrazine analogues have been developed successfully as a new group of insect growth regulators, called ecdysone agonists or moulting accelerating compounds. A notable feature is their high activity against lepidopteran insects, raising the question as to whether species‐specific analogues can be isolated. In this study, the specificity of ecdysone agonists was addressed through a comparative analysis in two important lepidopterans, the silkworm Bombyx mori L. and the cotton leafworm Spodoptera littoralis (Boisd.). RESULTS: When collections of non‐steroidal ecdysone agonists containing different mother structures (dibenzoylhydrazine, acylaminoketone, tetrahydroquinoline) were tested, in vitro reporter assays showed minor differences using cell lines derived from both species. However, when compounds with high ecdysone agonist activity were examined in toxicity assays, larvicidal activity differed considerably. Of note was the identification of three dibenzoylhydrazine analogues with > 100‐fold higher activity against Bombyx than against Spodoptera larvae. CONCLUSION: The present study demonstrated that species‐specific ecdysone‐agonist‐based insecticides can be developed, but their species specificity is not based on differences in the activation of the ecdysone receptor but rather on unidentified in vivo parameters such as permeability of the cuticle, uptake/excretion by the gut or metabolic detoxification. Copyright © 2010 Society of Chemical Industry     

Nonsteroidal ecdysone receptor agonists use a water channel for binding to the ecdysone receptor complex EcR/USP

The ecdysone receptor (EcR) possesses the remarkable capacity to adapt structurally to different types of ligands. EcR binds ecdysteroids, including 20-hydroxyecdysone (20E), as well as nonsteroidal synthetic agonists such as insecticidal dibenzoylhydrazines (DBHs). Here, we report the crystal structures of the ligand-binding domains of Heliothis virescens EcR/USP bound to the DBH agonist BYI09181 and to the imidazole-type compound BYI08346. The region delineated by helices H7 and H10 opens up to tightly fit a phenyl ring of the ligands to an extent that depends on the bulkiness of ring substituent. In the structure of 20E-bound EcR, this part of the ligand-binding pocket (LBP) contains a channel filled by water molecules that form an intricate hydrogen bond network between 20E and LBP. The water channel present in the nuclear receptor bound to its natural hormone acts as a critical molecular adaptation spring used to accommodate synthetic agonists inside its binding cavity.     

烟粉虱MED隐种蜕皮激素受体基因cDNA克隆、转录与组织表达

蜕皮激素受体(ecdysone receptor,Ec R)是调控昆虫蜕皮、变态和生殖等过程中基因表达的重要调控因子。为研究Ec R在烟粉虱Bemisia tabaci生长发育中的作用,利用RT-PCR和RACE等技术扩增了烟粉虱MED隐种Ec R基因的c DNA全长序列,并利用实时荧光定量PCR技术检测其在烟粉虱不同发育时期相对表达量的变化。Bt Ec R c DNA序列全长2 844 bp,含有一个1 518 bp的开放阅读框,共编码505个氨基酸,预测蛋白分子量为56 k D,等电点为6.43,含有特殊结构功能域:DNA结合域(DBD_Ec R)和配体结合域(LBD_Ec R),该序列编码的蛋白质序列与其它已报道的昆虫Ec R蛋白序列具有很高的相似性,命名为Bt Ec R(Gen Bank登录号:KR534774)。Bt Ec R在MED隐种若虫、雌成虫各发育时期均有表达,若虫期在伪蛹前期达到最高值,成虫期呈现先升高后降低的趋势,且在羽化后第7天达到最高值,约为羽化第1天的23倍。研究结果为揭示Bt Ec R在烟粉虱整个生长发育过程中的作用提供了重要依据。