重组增效蛋白对Bt和氯氰菊酯防治棉铃虫的增效作用

前期在大肠杆菌中表达得到具有显著生物活性的含粉纹夜蛾颗粒体病毒增效蛋白C端818个氨基酸的融合蛋白P96。以毒饲料法研究P96对苏云金芽孢杆菌和氯氰菊酯杀棉铃虫敏感、抗性品系的增效作用。结果表明：P96对Bt杀棉铃虫敏感、低抗、中抗品系的增效比分别为3．34、4．72和9．82；对氯氰菊酯杀棉铃虫敏感、低抗、中抗品系的增效比分别为2．53、3．38和6．24。随棉铃虫抗性升高，P96的增效作用愈为明显。单对汰选是快速选育棉铃虫抗Bt品系的有效方法。     

Selection and heritability of resistance to Bacillus thuringiensis subsp kurstaki and transgenic cotton in Helicoverpa armigera (Lepidoptera: Noctuidae)

Compared with an unselected susceptible population, a cotton bollworm, Helicoverpa armigera (Hübner), population selected for 22 generations with transgenic cotton leaves (modified Cry1A) in the laboratory developed 11.0-fold resistance to Cry1Ac (one single-protein product MVPII). Resistance to Bacillus thuringiensis Berliner subsp kurstaki (Btk) was selected for 22 generations with a 5.2-fold increase in LC50. The estimated realized heritabilities (h2) of resistance for transgenic-cotton- and Btk-selected populations were 0.1008 and 0.2341, respectively. This reflects the higher phenotypic variation in response to Cry1Ac in the transgenic-cotton-selected population. This variation may have been caused by differences in protein toxin levels expressed in different growth stages of the transgenic cotton. Because of the different slopes of the probit regression lines between Cry1Ac and Btk, the estimated realized h2 cannot be used visually to compare resistance development to Cry1Ac and Btk in H armigera. Thus, the response quotient (Q) of resistance was also estimated. The Q values of resistance for transgenic-cotton- and Btk-selected populations were 0.0763 and 0.0836, respectively. This showed that the rate of resistance development would be similar in both selection populations. This result indicates that the selection of resistance using transgenic cotton is different from that selected using the single toxin. Resistance risk to transgenic cotton and Btk in field populations was assessed assuming different pressures of selection by using the estimated h2. Assuming the h2 of resistance in a field population was half of the estimated h2, and the population received prolonged and uniform exposure to transgenic cotton or Btk causing >70% mortality in each generation, we predicted that resistance would increase 10-fold after <23 generations for Cry1Ac in transgenic cotton-selected-populations and after <21 generations for Btk in Btk-selected populations. Cross-resistance would be expected after <48 generations for Btk in transgenic-cotton-selected populations and after <21 generations for Cry1Ac in Btk-selected population. The results show that the potential to evolve resistance is similar in both transgenic-cotton- and Btk-selected populations, but that cross-resistance development to Btk is slower in transgenic-cotton-selected populations than cross-resistance development to Cry1Ac in Btk-selected populations.     

转Bt基因棉种植前后3个棉铃虫种群对4种杀虫剂的抗性变化

分别于1990年、1992年、2001年和2005年从山东泰安、山东夏津和湖北天门采集3个棉铃虫H elicoverpa arm igera种群,采用点滴法测定了4种杀虫剂对其的毒力,以监测各种群的抗药性变化趋势。结果表明:19901992年3个种群棉铃虫对氰戊菊酯的抗性增长迅速(13.256.1倍);19922001年抗性增长速度有所减缓(3.111.0倍);20012005年抗性水平降低。3个种群棉铃虫对灭多威和辛硫磷的抗性变化趋势与氰戊菊酯的基本相似,但抗性水平较低;对硫丹则均一直处于敏感水平。引起抗药性变化的原因主要与各类药剂的使用频率和转B t基因抗虫棉的大面积推广有关。     

苏云金芽孢杆菌HBF-1芽孢对棉铃虫幼虫生长发育及酶活的影响

苏云金芽孢杆菌HBF-1菌株的芽孢对初孵棉铃虫幼虫无明显杀伤作用,但可以直接影响其正常的生长发育,使幼虫前期体重增加;而后期发育迟缓,历期延长,成虫产卵量明显减少。通过光学显微镜观察发现,用含有HBF-1芽孢的饲料饲喂棉铃虫幼虫,幼虫中肠肠壁细胞遭到破坏;随着虫龄的增大,微绒毛逐渐脱落,有的部位细胞壁破裂。酶活的测定结果表明:初孵棉铃虫幼虫取食含有HBF-1芽孢(100μg/g)的饲料后,幼虫体内的乙酰胆碱酯酶、羧酸酯酶和谷胱甘肽-S-转移酶的活性发生了变化。     

Genetics of Heliothis and Helicoverpa resistance to chemical insecticides and to Bacillus thuringiensis

Genetic linkage maps of Heliothis virescens and Helicoverpa armigera are being used to identify and characterize resistance-conferring genes. The insensitive acetylcholinesterase conferring resistance to organophosphorus insecticides and the insensitive sodium channel conferring resistance to pyrethroids have both been mapped in H. virescens. The linkage mapping approach permits a genetic dissection of resistance, even when the mode of action and lethal target are not precisely known, such as for the insecticidal toxins from the bacterium Bacillus thuringiensis (Bt). We have identified and mapped a major Bt-resistance locus in a strain of H. virescens exhibiting up to 10000-fold resistance to Cry1Ac toxin and are currently developing a linkage map for H. armigera with a set of ‘anchor’ loci to facilitate comparison with H. virescens. Both species are currently experiencing their first significant selective pressure in the field by transgenic cotton expressing Cry1Ac, and timely identification of resistance mechanisms and their underlying genetic basis will be essential in successfully managing the Bt resistance that will eventually appear. ©1997 SCI     

棉铃虫Bt抗性种群的RAPD-PCR初步分析

经室内筛选获得了棉铃虫对Bt杀虫剂、Bt毒蛋白和转Bt基因棉的抗性种群。利用RAPD技术 ,成功地扩增得到 105条多态性条带 ,经过聚类分析发现 ,棉铃虫对Bt产生抗性后在基因水平发生了变异。RAPD技术不仅可以用来鉴定棉铃虫对Bt是否产生抗性 ,而且可以区分不同的Bt抗性种群     

Bt对棉铃虫天蚕素抗菌肽基因(HacD)转录的影响

抗菌肽是昆虫体液免疫的重要组成部分, 对抵御外源致病微生物的侵染具有重要作用。明确苏云金芽胞杆菌Bacillus thuringiensis(Bt)诱导下棉铃虫天蚕素抗菌肽基因(HacD)转录的空间特征和时序特征, 有利于进一步了解昆虫体液免疫与棉铃虫抗Bt发展的关联性。本研究测定了HacD cDNA序列, 对序列进行了蛋白质翻译, 并通过RT-qPCR技术研究了棉铃虫HacD基因在Bt诱导下的转录规律。结果表明, 棉铃虫HacD cDNA全长189 bp, 其中GC含量为48.15%, 肽链由63个氨基酸组成。Bt诱导后棉铃虫幼虫HacD相对转录水平整体呈显著上调趋势, 其中脂肪体中HacD的相对转录水平上调最高, 其次为血淋巴。Bt对棉铃虫幼虫进行短期诱导后, 脂肪体和血淋巴中的HacD基因的相对转录水平均呈持续上调趋势, 分别在诱导24 h和18 h时达到最高, 随后均持续下调。对幼虫进行持续诱导后, F0代初孵幼虫、5龄幼虫、蛹和成虫的HacD基因均可以保持上调的转录特性;传代培养获得的F1代初孵幼虫、5龄幼虫、蛹和成虫的HacD基因转录较F0代下调, 但均高于对照组。综上所述, Bt能够诱导棉铃虫HacD基因转录水平上调。     

高效苏云金杆菌KN11杀虫剂产品开发及应用

苏云金杆菌（Bacillus thuringiensis,Bt）商品化制剂是目前应用范围最为广泛,占据市场容量最大的微生物农药,在对害虫的生物防治中发挥了巨大的作用。本文综述了高效苏云金杆菌KN11产品开发背景、菌株信息、产品特性、登记情况、应用技术等,并对未来KN11产品改进及应用进行了展望。     

Seasonal Dynamics of Metabolic Mechanisms Mediating Pyrethroid Resistance in Helicoverpa armigera in Central India

Very high cypermethrin and fenvalerate resistance frequencies were recorded in Helicoverpa armigera (Hübner) populations in central India during the 1993–94, 1994–95 and 1995–96 cropping seasons. Synergism assays and biochemical analyses of detoxification enzyme levels indicated that mono-oxygenases and esterases were important metabolic mechanisms mediating pyrethroid resistance. Piperonyl butoxide- (PBO) and profenofos-suppressible pyrethroid resistance were correlated with enhanced levels of cytochrome P450 and general esterases respectively. Enzyme assay data indicated that high cytochrome P450 levels generally coincided with low esterase activity and vice versa. Similarly, synergist bioassays showed that PBO-insensitive resistance was frequently associated with profenofos-sensitive resistance and vice versa. Oxidase- and esterase-mediated mechanisms evidently alternated in a reciprocal manner, with perceptible shifts in relative importance occurring during mid-October in all three seasons and in late January in 1995. Apart from metabolic mechanisms, a synergist-insensitive resistance mechanism (believed to be nerve insensitivity), accounted for an average of 51, 30 and 28% of cypermethrin resistance during the 1993–94, 1994–95 and 1995–96 seasons respectively. © 1997 SCI.     

A perspective on management of Helicoverpa armigera: transgenic Bt cotton,IPM, and landscapes

Helicoverpa armigera is a major pest of agriculture, horticulture and floriculture throughout the Old World and recently invaded parts of the New World. We overview of the evolution in thinking about the application of area‐wide approaches to assist with its control by the Australian Cotton Industry to highlight important lessons and future challenges to achieving the same in the New World. An over‐reliance of broad‐spectrum insecticides led to Helicoverpa spp. in Australian cotton rapidly became resistant to DDT, synthetic pyrethroids, organophosphates, carbamates and endosulfan. Voluntary strategies were developed to slow the development of insecticide resistance, which included rotating chemistries and basing spray decisions on thresholds. Despite adoption of these practices, insecticide resistance continued to develop until the introduction of genetically modified cotton provided a platform for augmenting Integrated Pest Management in the Australian cotton industry. Compliance with mandatory resistance management plans for Bt cotton necessitated a shift from pest control at the level of individual fields or farms towards a coordinated area‐wide landscape approach. Our take‐home message for control of H. armigera is that resistance management is essential in genetically modified crops and must be season long and area‐wide to be effective. © 2016 Society of Chemical Industry     

Regional reversion of insecticide resistance in Helicoverpa armigera (Lepidoptera: Noctuidae) is associated with the use of Bt cotton in northern China

A continuous programme for monitoring resistance of Helicoverpa armigera (Hubner) to commonly used insecticides was undertaken for assessing the impact of Bt cotton planting on the evolution of the pest resistance from 1994 to 2002 in China. The bioassay results showed that the resistance levels per year in field populations of H armigera to lambda-cyhalothrin, phoxim and endosulfan decreased, respectively, to 9-15-fold, 11-14-fold and 6-8-fold in 2001-2002 from 197-262-fold, 52-74-fold and 18-38-fold before the cultivation of Bt cotton in 1997. This significant increase in susceptibility to insecticides is expected to result in a reduction in insecticide application for H armigera control in Bt cotton. It is concluded that Bt cotton is playing an important role in the long-term management of H armigera by increasing the potential for natural and chemical control of the pest.     

荧光增白剂对苏云金芽孢杆菌毒力的增效作用及其缈外防护功效

测试了3种荧光增白剂Tinopal LPW,VBL和JD－3对苏云金杆菌杀虫剂杀棉铃虫初孵幼虫毒力的影响，并检测了荧光增白剂VBL作为佐剂在Bt受紫外线照射的兴保护功效。结果表明，这3种二苯乙烯类光增白剂以10g/kg与Bt复配后的LC50值与不加荧光增白剂的Bt的LC50值相比，其增效比值分别为：1．23、1．49和2．00。说明这3种荧光增白剂对Bt杀铃虫的活性有不同程度的增效作用。而且复混剂（10g/kg荧光增白剂VBL＋Bt）在紫外线处理8h后，其杀虫活性仍保持70％以上，说明了荧光增白剂VBL是Bt一种好的光保护剂，能提高Bt抗紫外线的能力。     

拮抗细菌对Bt制剂杀虫效果的影响

从土壤、棉花根际和棉铃虫肠道内分离、筛选出细菌８株,初步鉴定为芽孢杆菌属、假单胞杆菌属和肠杆菌属,对水稻白叶枯、棉铃疫霉、棉花黄萎病原菌进行抑菌试验,拮抗细菌以菌悬液和细胞菌体分别与Ｂｔ制剂混配感染饲料,以２龄棉铃虫生测的结果表明,培养液无杀虫效果,拮抗细菌发酵液、菌体对Ｂｔ制剂杀虫效果有不同的影响。拮抗细菌中５株芽孢杆菌对Ｂｔ制剂有较好的增效作用,其杀虫效果均达７５％以上,防效接近ＨＤ┐７３无晶体突变株     

Patterns of resistance to organophosphate insecticides in field populations of Helicoverpa armigera in Pakistan

The pattern of organophosphate resistance in field populations of Helicoverpa armigera (Hübner) was monitored in Pakistan from 1994 through 1997 using an IRAC leaf-dip method. Generally, moderate to high resistance was found to an orthophosphorate, monocrotophos and to a dithiophosphorate, ethion. Resistance to the commonly used thiophosphorates, chlorpyrifos and profenofos, was very low during 1994 and 1995 but showed an upward trend during 1996 and 1997. Resistance factors to the other thiophosphorates such as quinalphos, parathion-methyl, methamidophos and triazophos remained very low. Baseline lethal concentrations for ethion, methamidophos and triazophos were at least four to five times higher than those for the other organophosphates tested, suggesting their low intrinsic efficacy against H armigera. © 1999 Society of Chemical Industry     

Inheritance of resistance to a Bacillus thuringiensis toxin in a field population of diamondback moth (Plutella xylostella)

Inheritance of resistance to the Bacillus thuringiensis Berl. CryIA(b) crystal protein was studied in Plutella xylostella L. (diamondback moth). A field population 50-fold more resistant to CryIA(b) than a control susceptible strain was used. Dose-mortality curves of the resistant population, the susceptible strain and the F₁ from the two reciprocal crosses were compared. Resistance transmission to the F₁ was dependent on the sex of the resistant progenitor. Sex ratio of the survivors to high doses of CryIA(b) in the F₁ of the two reciprocal crosses did not corroborate the preliminary hypothesis of resistance being due to a recessive sex-linked allele. Since, in a previous work, the loss of CryIA(b) binding capacity of resistant insects had been demonstrated, binding to midgut tissue sections from F₁ individuals was also analysed. The presence of binding in all of the F₁ preparations showed that, at least, a recessive autosomal allele was responsible for the loss of binding capacity in the resistant population.