用单克隆抗体结合酶联免疫吸附试验检测棉铃虫幼虫体内的核多角体病毒

在常规ＥＬＩＳＡ间接法的基础上，应用单克隆抗体检测感染棉铃虫核型多角体病毒的棉铃虫幼虫体内病毒粒子。３龄幼虫饲喂表层涂有ＨａＮＰＶ人工饲料后９小时，即可在幼虫抽提液中检出病毒粒子抗原，而典型病虫显症需５～６天后才出现。因此本法是一种灵敏、快速、特异性强的检测昆虫杆状病毒的方法。利用单克隆抗体结合ＥＬＩＳＡ试验分别检测来自江苏和山东不同地区棉田自然死亡的棉铃虫幼虫，表明在江苏和山东不同棉区均可检测到ＨａＮＰＶ病毒粒子，但地区间棉铃虫核型多角体病毒检出频率有显著差异     

棉铃虫核型多角体病毒对宿主昆虫的弱化作用

棉铃虫核型多角体病毒（ＨａＮＰＶ）感染２龄幼虫,感染３ｄ,幼虫体长和体重明显小于对照幼虫。病毒感染４龄幼虫,残留活虫幼虫历期延长,但取食量减少３７．５２％。病毒感染５龄幼虫,感染幼虫蜕皮时间延迟１８ｈ,幼虫历期延长２ｄ。幼虫血淋巴蛋白电泳结果表明,幼虫蜕皮时间延迟主要原因是蜕皮过程延长。病毒感染５龄初幼虫,残留活虫蛹重减轻１４．１９％,成虫产卵量减少２０．５８％。结果表明,ＨａＮ－ＰＶ感染棉铃虫幼     

棉铃虫C型凝集素抑制多角体病毒体内增殖

为探究C型凝集素（C-type lectin）在病毒传播中的可能作用,采用半定量RT-PCR法检测了棉铃虫Helicoverpa armigera不同C型凝集素在核型多角体病毒（nuclearpolyhedrosis virus,NPV）感染后的表达变化,并通过real-time qPCR和RNA干扰技术研究了NPV在棉铃虫体内的增殖状况及对棉铃虫致死率的影响。结果显示,在感染NPV后,棉铃虫血细胞C型凝集素Ha-lectin1和Ha-lectin5基因表达变化明显,其中Ha-lectin1在血细胞和脂肪体中的表达显著上调;注射dsRNA后,Ha-lectin1基因相对于actin的表达量均值由0.19下降到0.04,下降幅度达79%,显示Ha-lectin1基因被成功干扰。随着Ha-lectin1基因的沉默,病毒滴度由0.0015快速升高到1.95,棉铃虫的死亡率也随之增加到97.7%。表明C型凝集素能够抑制NPV在棉铃虫体内的增殖。     

Management of chickpea pod borer, Helicoverpa armigera (Hubner) through Nuclear Polyhedral Virus isolates

Investigations were carried out on Helicoverpa armigera Nuclear Polyhedrosis Virus (HaNPV) isolates collected from different parts of India to study their genetic diversity through molecular characterization and their assessment in the IPM of Helicoverpa armigera on chickpea under field conditions. Variations were found in the DNA profiling of different HaNPV isolates. The similarity matrix relating the different isolates showed similarity co-efficients ranging from 0.38 to 0.82. The highest genetic similarity index of 0.82 was seen between the isolates from Raichur and Guntur followed by 0.77 between the isolates from Coimbatore and Raichur. Field evaluation of different HaNPV isolates against H. armigera in the chickpea ecosystem at the Main Agricultural Research Station, University of Agricultural Sciences, Dharwad, Karnataka for 2 years revealed that the performance of the Gulbarga and Coimbatore isolates was better than that of others in terms of their ability to cause greater larval mortality of H. armigera with a greater yield of chickpea. The Incremental Benefit Cost Ratio was highest with the Gulbarga (2.88) and Coimbatore (2.83) isolates, followed by the Dharwad isolate (2.39). This research paper was presented in the 2nd conference on precision crop protection held at the University of Bonn, Germany during 10–12th October 2007.     

PCR法证实棉铃虫核型多角体病毒对棉铃虫经卵和蛹的垂直传播

根据棉铃虫核型多角体病毒的多角体蛋白基因设计5’引物和3’引物，通过PCR技术成功扩增389bp的目的片段，并对该片段进行了克隆和测序。结果表明，扩增片段为棉铃虫核型多角体病毒多角体蛋白基因序列。该检测方法的灵敏度为10fg，应用此技术从带毒的虫卵和蛹的总DNA中也检测到该病毒的存在。该方法从分子水平上证明了病毒的垂直传递。     

HaNPV对红脉穗螟弱化作用的研究

在实验室条件下测试了棉铃虫核型多角体病毒(Helicoverpa armigera nuclear polyhedrosis virus,HaNPV)对红脉穗螟Tirathaba rufivena Walker生长发育和繁殖力的影响。结果显示:HaNPV感染3龄幼虫6d,幼虫体长和体重分别减少37.53%和62.49%,幼虫发育历期延长11.80d,蛹重和单雌平均产卵量分别降低44.07%和90.23%;感染5龄幼虫3d,幼虫体长和体重分别减少13.02%和22.40%,幼虫发育历期延长6.58d,蛹重和单雌平均产卵量分别降低22.88%和58.44%。表明HaNPV不仅能直接杀死宿主昆虫,而且对残存活虫有很强的弱化作用。     

Effects of Cry1Ac toxin of Bacillus thuringiensis and nuclear polyhedrosis virus of Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) on larval mortality and pupation

In the laboratory, the percentage mortality and pupation of Helicoverpa armigera (Hübner) were investigated when larvae were exposed to Cry1Ac of Bacillus thuringiensis Berliner, nuclear polyhedrosis virus of H. armigera (HaNPV) or Cry1Ac and HaNPV together. The results revealed that interactions between Cry1Ac and HaNPV varied with bioassay method and concentration of the suspension. When larvae were infected using a suspension containing both HaNPV and Cry1Ac, most combinations of Cry1Ac (62.5, 125 and 250 microg mL(-1)) and HaNPV (1.2 x 10(6), 6.0 x 10(6) and 3.0 x 10(7) PIB mL(-1)) showed an antagonistic effect. In the bioassay procedure where larvae were force fed diet containing Cry1Ac 48 h after being infected by HaNPV, interaction between Cry1Ac (0.5, 1, 2, 4 and 8 microg mL(-1)) and HaNPV (6.0 x 10(6) and 3.0 x 10(7) PIB mL(-1)) showed an additive effect, while combinations of Cry1Ac (0.5, 1, 2 and 4 microg mL(-1)) and HaNPV (1.2 x 10(6) PIB mL(-1)) showed an antagonistic effect. In the bioassay procedure where larvae being infected by HaNPV were fed on Cry1Ac diet from neonate to death or pupation, the results suggested that Cry1Ac and HaNPV showed an additive interaction. The percentage mortality was lower in the treatment of larvae infected by transgenic Bt cotton leaf discs containing HaNPV suspension than in the treatment of larvae by conventional cotton leaf discs containing HaNPV, while the pupation rate was higher. The combination of Bt cotton and HaNPV showed antagonism. The present results showed that a combination of Cry1Ac and HaNPV usually resulted in mortality levels greater than in the case of Cry1Ac but not greater than with the virus alone.