斜纹夜蛾核多角体病毒研究进展

斜纹夜蛾核多角体病毒杀虫剂是一种新型的生物农药,从分子生物学、病毒流行学、对非靶标生物的毒性、工厂化生产及田间药效等方面阐述了斜纹夜蛾核多角体病毒的研究进展.     

春尺蠖NPV生物学特性及室内增殖研究

春尺蠖核型多角体病毒(Apocheima cinerarius nucleopolyhedrovirus,AciNPV)是春尺蠖的主要病原微生物。该文研究了AciNPV的形态结构和室内生物活性,及AciNPV室内生产过程中的适宜感染浓度、感染虫龄和感染温度,从而为该病毒今后的商业化生产和应用提供借鉴。     

家蚕核型多角体病毒ORF 75基因的生物信息学分析

通过对家蚕核型多角体病毒(Bombyx mori,nucleopolyhedrovirus:BmNPV)ORF 75基因核苷酸序列进行生物信息学分析,BmNPVORF 75基因位于病毒基因组70 485 bp和71 264 bp之间,编码259个氨基酸残基的多肽,预测分子相对质量为30.8 kDa,等电点为7.67,分子式为C1409H2157N351O390S19;蛋白在大肠杆菌内的半衰期大于10 h;酸性氨基酸(Asp Glu)占10.5%,碱性氨基酸(Arg Lys)占10.8%;蛋白的不稳定指数为42.67,是不稳定蛋白.利用Prosite数据库扫描,查到4类9个蛋白修饰位点.通过Blast搜索氨基酸同源序列发现,BmNPVORF 75同源蛋白存在于所有测序的鳞翅目昆虫杆状病毒中,不存在于其他非鳞翅目昆虫杆状病毒中.同源序列比对结果,BmNPVORF75蛋白的氨基酸序列与苜蓿银纹夜蛾核型多角体病毒(Autographa californica MNPV)ORF92之间的同源性达到100%,应为相同基因.依照BmNPVORF75同源基因序列相似性,对13种昆虫杆状病毒进行进化树构建分析,其中,BmNPVORF75与薄荷灰夜蛾多粒包埋型核型多角体病毒(Rachiplusia ouMN-PV)ORF89基因进化距离很近,同源性较高.     

家蚕核型多角体病毒ODV囊膜蛋白基因的融合表达及引导外源蛋白进入多角体的研究

为了探索家蚕核型多角体病毒(BmNPV)的包涵体衍生病毒(ODV)囊膜蛋白在病毒粒子包涵入多角体过程中的作用,选择2个ODV囊膜蛋白ODV-E25和ODV-E56作为研究对象,利用能够表达多角体蛋白的BmNPV Polh+Bac-to-Bac表达系统构建了融合表达E25-EGFP和E56-EGFP的重组病毒,在家蚕卵巢培养细胞系(BmN)中进行表达和定位分析。结果表明2种融合蛋白均在BmN细胞中成功表达,并于荧光显微镜下观察到绿色荧光主要分布在细胞核中。从感染的BmN细胞中收集纯化多角体,观察到多角体也能激发出绿色荧光,用Western blot方法进一步证实多角体中含有融合蛋白。这一现象表明当囊膜蛋白基因与外源基因融合表达时,外源融合蛋白能够进入多角体内部,推测这2种ODV囊膜蛋白不仅在病毒粒子包涵入多角体的过程中起信号引导作用,并能引导外源目的蛋白进入多角体。     

Comparative activity of Choristoneura fumiferana nucleopolyhedrovirus propagated in different hosts

The biological activity of the Ireland strain of Choristoneura fumiferana (Clem) nucleopolyhedrovirus (CfMNPV) propagated in different hosts was determined to provide the basis upon which genetically modified CfMNPV, or other naturally occurring isolates, should be compared. Occlusion bodies (OB) derived from CF-203 cells were significantly larger and more pathogenic than those propagated in vivo when tested against the fifth larval instar of C fumiferana (Clem) and C occidentalis Freeman. The dose-responses (LD50 and LD95, expressed as occlusion bodies per larva) of C fumiferana larvae to in vitro-propagated OBs were 274 and 5785, respectively. The values of LD50 and LD95 to C occidentalis larvae were 19 and 118, respectively. There were no significant differences in pathogenicity or size when OBs propagated in C fumiferana larvae were tested against either insect species, nor were there significant differences for OBs propagated in C occidentalis larvae. The LD50 and LD95 of in vivo-produced OBs to C fumiferana were 925 and 61988, respectively. The LD50 and LD95 to C occidentalis were 50 and 453, respectively. OBs propagated in vitro had a mean volume of 13.13 microm3, whereas those propagated in vivo ranged from 0.84 to 1.41 microm3. The median survival time-responses (ST50) of fifth-instar C fumiferana or C occidentalis larvae to OBs propagated in vivo were not significantly different from those propagated in vitro at the dosage levels tested. Values of ST50 of C fumiferana larvae to in vitro- and in vivo-produced OBs at dosages causing less than 50% mortality rangedfrom 9.6 to 9.8 days post-inoculation (dpi), whereas a LD95 dose resulted in ST50 values ranging from 7.3 to 7.7 days. ST50 values of C occidentalis larvae at dosages causing less than 50% mortality ranged from 9.8 to 10.2 dpi, whereas a LD95 dose resulted in ST50 values ranging from 9.5 to 9.8 dpi. The median feeding cessation time-response (FT50) of fifth-instar C fumiferana larvae to OBs propagated in vitro (5.7 days) was not significantly different from the FT50 of those propagated in vivo in either insect species (5.3 and 5.7 days) at the dosage level tested (LD95). No significant differences in FT50 values were observed between OBs propagated in either larval host. The FT50 of C occidentalis larvae to OBs propagated in vitro (7.7 days) was not significantly different from that to those propagated in vivo in C occidentalis larvae (7.6days), but somewhat different (7.2 days) from that to those propagated in C fumiferana larvae. Results indicate that CfMNPV can be propagated in vivo in either C fumiferana or C occidentalis larvae (or sequentially through both) without alteration in infectivity, although the use of the CF-203 cell line yields the most biologically active OBs.     

Optimization of production of nucleopolyhedrovirus ofHelicoverpa armigera throughout larval stages

Studies were carried out to optimize production of nucleopolyhedrovirus of American bollworm,Helicoverpa armigera (Hübner), by treating larvae individually with an inoculum dose that allowed maximal larval growth and also gave the highest occlusion bodies (OB) yield/larva. The maximum virus yield of 12.2x 10⁹ OB/larva was obtained when 6-day-old larvae were fed individually with a dose of 1 x 10³ OB. Topical spiracular treatment of larvae as old as 8 days with 10 μ of 2x 10⁷ OB ml^-1 gave the highest yield, of 15.2x 10⁹ OB from 13-day-old larvae, of 12.8x 10⁸ OB from prepupae and of 1.49x 10⁸ OB from pupae at the time of their death. These studies showed that dietary inoculum is the best route for 6-day-old larvae and topical spiracular treatment is the best for 8-day-old larvae.     

中红侧沟茧蜂对棉铃虫核型多角体病毒的传播作用

中红侧沟茧蜂Microplitis mediator与核型多角体病毒(nucleopolyhedrovirus, NPV)是棉铃虫Helicoverpa armigera的两种重要生物防治因子。中红侧沟茧蜂传播NPV对于利用二者协同防治棉铃虫具有重要意义。本研究探讨了给中红侧沟茧蜂饲喂含病毒蜂蜜水、体表喷洒病毒液、中红侧沟茧蜂在染毒宿主体内产卵、从染毒宿主体内发育、蜂茧浸泡病毒5种带毒方式的传播病毒效率,以及饲喂带病毒蜂蜜水方式下的传播机制。结果表明,饲喂带病毒蜂蜜水和体表喷洒病毒时中红侧沟茧蜂传毒率较高,在连续传毒的3 d内传毒效率分别为15.1%、9.1%～9.3%、2.4%～4%,其他3种方式传毒效率较低。在田间防治时可以利用中红侧沟茧蜂的传毒作用采用病毒与寄生峰的协同防控策略。     

美国白蛾核型多角体病毒室内大量增殖的研究

研究了饲毒时美国白蛾幼虫虫龄和饲毒剂量对美国白蛾核型多角体病毒(HcNPV)青岛分离株产量的影响,从而探寻HcNPV室内大量增殖的最佳条件。结果表明,虫龄、剂量以及虫龄和剂量的组合对单头幼虫的HcNPV产量、幼虫死亡率以及病毒总产量均具有极显著的影响,在室内大量增殖HcNPV时,以1.48×106 OBs/头的饲毒剂量饲喂5龄和6龄幼虫均可以获得最高的病毒产量,百头幼虫的病毒总产量分别达到1.55×1011 OBs和1.50×1011 OBs。     

辽宁地区樗蚕的生物学特性及人工饲育技术研究

樗蚕(Samia cynthia Walker et Felde)是一种珍贵的泌丝及食用昆虫资源。通过室内及野外人工饲育的方法,观察分布在辽宁地区的樗蚕的形态特征和生活习性,发现与分布于其它地区的樗蚕存在不同程度的差异。辽宁地区的樗蚕年发生1-2代,喜食臭椿(Ailanthus altissima)、山花椒(Zanthoxylum schinifollum)、枣树(Zizyphus jujube)、黄檗(Phellodendron amu-rense)等的树叶,幼虫4眠5龄,龄期经过25-37d,饲料种类不同可造成龄期差异;以蛹滞育。人工饲育樗蚕可采用小蚕室内育,大蚕野外育的饲养形式;种茧保护可采用柞蚕(Antheraea pernyi)的种茧越冬保护技术与方法,春季种茧出库后在温度20～23℃、相对湿度75%～85%的条件下暖茧,成虫交配时间超过4h的卵孵化率较高;樗蚕的主要病害有细菌性软化病、白僵病、微粒子病,春、秋季野外饲养的樗蚕易被蚕饰腹寄蝇(Blepharip azebina)寄生危害。樗蚕茧丝的纤度为1.80～3.60dtex,茧层丝胶练减率平均为25%,茧丝中的丝素蛋白约78%。