转基因抗棉铃虫抗草甘膦棉花对棉铃虫的抗虫性及对二种地老虎幼虫生长发育的影响

为了明确转Bt Cry1A+CP4EPSPS基因抗棉铃虫抗草甘膦棉花对棉铃虫Helicoverpa armigera(Hübner)的抗性水平及对八字地老虎Amathes c-nigrum和黄地老虎Agrotis segetum的影响,作者用转基因棉花和非转基因棉花叶片对棉铃虫、八字地老虎和黄地老虎幼虫进行了室内饲养观察。结果表明,转基因抗棉铃虫抗草甘膦棉花对新疆南部第2代棉铃虫表现为中抗,幼虫校正死亡率为70.4%;棉花叶片在整个生育期均能表达Bt毒蛋白,整个生育期Bt毒蛋白含量呈逐渐降低的趋势。转基因抗棉铃虫抗草甘膦棉花对八字地老虎幼虫有强烈的毒杀作用,可以引起幼虫大量死亡,对存活幼虫的生长发育也有明显的抑制作用,幼虫发育迟缓,不能顺利化蛹;但对黄地老虎幼虫没有明显的影响,幼虫可以正常生长。     

Predation of cotton bollworm by green lacewings in the presence of cotton aphid as alternative prey on transgenic Bt and conventional cotton

Experiments were conducted in small arenas and on whole plants to explore the effect of cotton aphids, Aphis gossypii Glover (Hemiptera: Aphididae), as alternative prey on the predation of Helicoverpa armigera Hübner (Lepidoptera: Noctuidae) larvae by green lacewing larvae, Mallada signatus Schneider (Neuroptera: Chrysopidae). Transgenic Bt (Bollgard II^®) and conventional cotton plants were included to explore potential differences in the predator's performance on these cotton types. In small arenas, the presence of 20 aphids reduced predation on H. armigera larvae by 22% (from 5.5 to 3.3 of 10) by a single lacewing larva over a 24‐h period. The presence of H. armigera reduced predation on aphids by ca. 29% (from 16.8 to 11.0 of 20) over 24 h. On whole plants, the presence of alternative prey had no effect on the number of H. armigera larvae or aphids remaining after 3 days. The presence of H. armigera larvae alone, without the predator, caused a 24% reduction in the numbers of aphids on conventional, but not on Bt cotton plants. The combination of Bt cotton and lacewing larvae caused a 96.6% removal of early‐stage H. armigera larvae, a statistically significant increase over the addition of the proportions (91.6%) removed by each factor measured separately, providing evidence of synergism. These studies suggest that the presence of aphids as alternative prey would not necessarily disrupt the predation by green lacewing on larvae of H. armigera, especially on Bt cotton.     

棉铃虫取食转Bt棉后蛹的抗寒力测定

在棉铃虫HelicoverpaarmigeraH櫣bner人工饲料中分别添加转Bt基因棉叶粉和常规棉叶粉饲喂幼虫,经滞育诱导后,发现2个处理间棉铃虫蛹的滞育率相似,2个处理间滞育蛹的自由水和结合水含量没有差异;但取食含Bt棉叶粉人工饲料的棉铃虫滞育蛹的过冷却点和结冰点显著高于对照组,蛹重及与抗寒性有关的脂肪和糖原含量均显著低于对照组。     

华北地区棉铃虫对转Bt基因抗虫棉抗性适应的模拟模型

通过对华北地区耕作制度和生态系统的了解,在充分考虑种群遗传学、生物学和人为操纵因子等三大因素的基础上,建立了一个预测棉铃虫对转Bt基因抗虫棉抗性适应的模拟模型。在华北地区典型的耕作制度下,如果所有棉田均为Bt棉,则Bt棉的预期寿命为7年;如果只有春播棉为Bt棉(约占棉田总面积的70%),则其寿命为10年。模型的灵敏度分析表明, Bt棉的使用寿命随抗性基因的显性度、初始抗性频率、Bt棉所占比例等因素的增长而迅速缩短。当Bt棉表达的杀虫蛋白量恰好全部杀死敏感基因型(G_SG_S)个体时,Bt棉的预期寿命最短。由于国外采用的“高剂量/庇护所”抗性治理策略不适用于棉铃虫及华北棉区的耕作制度,我国需要加强对其它抗性治理措施(如转双基因抗虫棉)的研究与应用。     

Next‐generation transgenic cotton: pyramiding RNAi and Bt counters insect resistance

Transgenic crops producing insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) are extensively cultivated worldwide. To counter rapidly increasing pest resistance to crops that produce single Bt toxins, transgenic plant ‘pyramids’ producing two or more Bt toxins that kill the same pest have been widely adopted. However, cross‐resistance and antagonism between Bt toxins limit the sustainability of this approach. Here we describe development and testing of the first pyramids of cotton combining protection from a Bt toxin and RNA interference (RNAi). We developed two types of transgenic cotton plants producing double‐stranded RNA (dsRNA) from the global lepidopteran pest Helicoverpa armigera designed to interfere with its metabolism of juvenile hormone (JH). We focused on suppression of JH acid methyltransferase (JHAMT), which is crucial for JH synthesis, and JH‐binding protein (JHBP), which transports JH to organs. In 2015 and 2016, we tested larvae from a Bt‐resistant strain and a related susceptible strain of H. armigera on seven types of cotton: two controls, Bt cotton, two types of RNAi cotton (targeting JHAMT or JHBP) and two pyramids (Bt cotton plus each type of RNAi). Both types of RNAi cotton were effective against Bt‐resistant insects. Bt cotton and RNAi acted independently against the susceptible strain. In computer simulations of conditions in northern China, where millions of farmers grow Bt cotton as well as abundant non‐transgenic host plants of H. armigera, pyramided cotton combining a Bt toxin and RNAi substantially delayed resistance relative to using Bt cotton alone.     

Cotton bollworm resistance to Bt transgenic cotton: A case analysis

Cotton bollworm (Helicoverpa armigera) is one of the most serious insect pests of cotton. Transgenic cotton expressing Cry toxins derived from a soil bacterium, Bacillus thuringiensis (Bt), has been produced to target this pest. Bt cotton has been widely planted around the world, and this has resulted in efficient control of bollworm populations with reduced use of synthetic insecticides. However, evolution of resistance by this pest threatens the continued success of Bt cotton. To date, no field populations of bollworm have evolved significant levels of resistance; however, several laboratory-selected Cry-resistant strains of H. armigera have been obtained, which suggests that bollworm has the capacity to evolve resistance to Bt. The development of resistance to Bt is of great concern, and there is a vast body of research in this area aimed at ensuring the continued success of Bt cotton. Here, we review studies on the evolution of Bt resistance in H. armigera, focusing on the biochemical and molecular basis of Bt resistance. We also discuss resistance management strategies, and monitoring programs implemented in China, Australia, and India.     

转Bt基因棉挥发性气味的化学成分及其对棉铃虫的电生理活性

报道了对转Bt基因棉“GK-12”及其常规棉亲本“泗棉3号”挥发性化学物质的分析结果,以及棉铃虫对Bt棉挥发性化学物质的电生理反应,以期为抗虫棉的生态学和安全性评价提供化学生态学的证据。结果表明,在7～8个真叶期,转Bt基因棉的α-蒎烯和β-蒎烯的相对含量比常规棉高许多,倍半萜烯C和一个含量很低的化合物（该化合物对棉铃虫有电生理活性）是常规棉所没有的。转基因Bt棉的挥发性物质中有9个化合物可引起棉铃虫的触角电位反应。     

Impacts of elevated CO2 on expression of plant defensive compounds in Bt‐transgenic cotton in response to infestation by cotton bollworm

• 1 The allocation of defensive compounds of transgenic Bt (cv. GK‐12) and nontransgenic cotton (cv. Simian‐3) grown in elevated CO₂ in response to infestation by cotton bollworm Helicoverpa armigera (Hübner) was studied in closed‐dynamics CO₂ chambers.
• 2 A significant reduction in foliar nitrogen content and Bt toxin protein occurred when transgenic Bt cotton grew under elevated CO₂. A significantly higher carbon/nitrogen ratio as well as condensed tannin and gossypol contents was observed for transgenic Bt (cv. GK‐12) and nontransgenic cotton in elevated CO₂, in partial support of the carbon nutrient balance hypothesis as a result of limiting nitrogen and excess carbon in cotton plants in response to elevated CO₂.
• 3 The CO₂ level and infestation time significantly affected the foliar nitrogen, condensed tannin, gossypol and Bt toxin protein contents of cotton plants after feeding by H. armigera. The interaction between CO₂ levels × cotton variety had a significant effect on foliar nitrogen content after injury by H. armigera.

     

转基因棉花对棉铃虫天敌寄生率的影响

采用分期接虫、分期回收法研究常规棉与转基因棉花品种 (系 )上棉铃虫寄生性天敌在不同棉花品种上的寄生作用。结果表明 ,无论是第 2代棉铃虫发生期间还是第 3 ,4代棉铃虫发生期间 ,无论是棉铃虫卵期还是幼虫期 ,转基因棉花品种 (系 )的棉铃虫卵、幼虫寄生率均显著低于常规棉花品种上的寄生率。棉铃虫卵期寄生蜂主要是拟澳洲赤眼蜂 ,幼虫寄生蜂为棉铃虫齿唇姬蜂等。由此看出转基因棉对棉铃虫寄生性天敌存在非亲和性。     

棉铃虫对转Bt基因棉的抗性筛选及遗传方式的研究

通过室内筛选获得了棉铃虫Helicoverpa armigera (Hübner)对转Bt基因棉的抗性种群,并在此基础上进行了抗性遗传方式的研究。试验结果表明：经16代筛选,棉铃虫对转Bt基因棉的抗性倍数上升到43.3倍。筛选过程中,棉铃虫7日龄幼虫的体重和成虫羽化率变化明显,被选为确定筛选剂量的标准。敏感与抗性棉铃虫杂交,正交、反交的显性度都小于0、杂交过程中雌雄性比基本接近1∶1、假设抗性基因为单基因时的χ²值较低,因此,初步认为棉铃虫对转Bt基因棉的抗性是常染色体单基因控制的不完全隐性遗传。     

转Cry1Ac+Cry2Ab棉对棉铃虫的控制作用及对天敌捕食烟粉虱功能反应的影响

文章以转Cry1Ac基因棉(中棉所41)和常规棉(中棉所49)为对照,研究了转Cry1Ac+Cry2Ab基因棉(639020)在棉花生长的关键时期——蕾期(二代棉铃虫发生期)、花期(三代棉铃虫发生期)和花铃期(四代棉铃虫发生期)对棉铃虫的控制作用,同时研究了639020棉田主要捕食性天敌(中华草蛉幼虫、龟纹瓢虫、小花蝽和草间小黑蛛)对烟粉虱的捕食功能,明确了639020棉花在生长的关键时期对棉铃虫的控制效果及对棉田主要捕食性天敌捕食功能反应的影响。结果表明,639020棉花对二代和三代棉铃虫具有良好的控制作用,抗虫性分别比中棉所41提高了52.85%和16.22%,其中前者差异达显著水平,后者差异不显著。在棉花蕾期、花期和花铃期,639020棉田棉铃虫落卵量都比中棉所41棉田和中棉所49棉田低(除二代棉铃虫发生期);棉铃虫幼虫数量都极显著低于常规棉,且都低于防治指标,但与中棉所41棉田无显著差异。639020棉田中华草蛉、龟纹瓢虫、小花蝽和草间小黑蛛对烟粉虱的捕食功能与中棉所41棉田和常规棉田相比无显著变化。研究结果以期为新型转基因棉花环境安全性研究及其外源基因的抗虫遗传效应和生产应用前景进行安全性评价。     

Yield,development, and quality response of dual‐toxin Bt cotton to Helicoverpa spp. infestations in Australia

To verify current thresholds for Bollgard II^® cotton in Australia, the impact of Helicoverpa spp. (Lepidoptera: Noctuidae) larvae on yield, development, and quality under various infestation intensities and durations, and stages of growth, was tested using small plot field experiments over two seasons. Infestation with up to 80 eggs m^?1 of Helicoverpa armigera (Hübner) and Helicoverpa punctigera Wallengren showed that species, infestation level, and stage of growth had no significant effect on yields of seed‐cotton or lint and on maturity and fibre quality. The duration of infestation of white flowers with H. punctigera neonates (maximum of every day for up to 4 weeks) had no impact on the yield of seed‐cotton or lint, maturity, and fibre quality, but when 100% of flowers were infested (compared with 0 or 50%), seed‐cotton and lint yields were significantly reduced and maturity was delayed. Infestation with up to 18 medium H. armigera larvae m^?1 at several plant stages did not significantly affect yields of seed‐cotton and lint, maturity, and fibre quality. A heliocide spray applied on a commercial farm at the current threshold resulted in a significantly higher lint yield, compared with a farm where no spray was applied. In conclusion, Bollgard II^® cotton is highly resistant to Helicoverpa spp. infestation.     

转Bt基因抗虫棉棉盲蝽防治指标研究

2005-2006年,采用田间接虫试验的方法,分别研究了江苏沿海棉区棉盲蝽优势种绿盲蝽和中黑盲蝽对Bt棉的为害与产量损失。结果表明,Bt棉田的被害株率和产量损失均随着接虫量的加大而增加,其对数相关均达极显著水平,直线相关也达显著至极显著水平;Bt棉田棉盲蝽的防治可以若虫数量与被害株率两项指标来把握,2种盲蝽混合种群第2、3、4代虫量推广防治指标分别为百株有2、3龄若虫5头、10头和20头上下;或被害株率依次为3%、8%和15%左右。     

转Bt基因抗虫棉对棉铃虫拒食作用及其机理研究

转Bt基因抗虫棉对 3 ,4,5龄棉铃虫幼虫的抗性表现形式为拒食作用 ,且随幼虫龄期的增加拒食作用明显降低 ,其中对 5龄幼虫的拒食作用很低。取食Bt棉后 ,3 ,4,5龄棉铃虫幼虫中肠消化酶比活力均较对照有所减退 ,且随幼虫龄期的增加减退率明显降低 ,其中对 5龄幼虫减退率最低。由此 ,解释了Bt棉对 3龄及 3龄以上棉铃虫幼虫抗性表现形式、抗性随不同幼虫龄期的差异性 ,及其抗性差异性的消化机理。     

转CpTI-Bt基因棉和转Bt基因棉对棉铃虫幼虫存活、生长及营养利用的影响

2000年7月中旬和8月中旬, 分别测定了采自田间的转CpTI-Bt基因双价抗虫棉（SGK321, 以下简称CpTI-Bt棉）和转Bt基因抗虫棉（中30,以下简称Bt棉）对棉铃虫Helicoverpa armigera幼虫存活、生长的影响。结果表明：7月中旬两种转基因抗虫棉抗虫效果均较好,尤其是CpTI-Bt棉棉叶和花瓣对4龄幼虫3天内致死率为92%以上;8月中旬两种转基因棉的抗虫活性均明显降低,且Bt棉的杀虫活性显著低于CpTI-Bt棉,其幼虫死亡率与对照受体棉中16的死亡率之间无显著差异,仅显著抑制了幼虫的生长;石远321(SGK321受体品系)的花瓣具有一定的抗虫活性,可显著降低取食幼虫的体重,甚至造成部分幼虫死亡; CpTI-Bt棉中,花瓣和棉叶的抗虫性明显高于蕾和铃心。对5龄幼虫取食棉铃1日后的营养指标测定结果显示： 两种转基因抗虫棉处理的幼虫相对生长率和相对取食量均显著低于石远321,但两者之间无显著差异; CpTI-Bt棉处理的幼虫近似消化率显著低于石远321和Bt棉,但其食物利用率显著高于石远321和Bt棉。     

Comparing the predatory performance of green lacewing on cotton bollworm on conventional and Bt cotton

We compared the survival of Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) eggs and larvae on Bt and conventional cotton, in the presence or absence of the generalist predator, green lacewing larvae, Mallada signatus, (Schneider) (Neuroptera: Chrysopidae). In small arenas, green lacewings consumed a similar number of H. armigera eggs (ave. 15.8 ± 1.3 on conventional, 12.6 ± 1.4 on Bt cotton per predator over 24 h) and larvae (ave. 6.8 ± 0.7 conventional, 6.5 ± 0.8 Bt per predator over 24 h) whether on Bt or conventional cotton leaves. Likewise, similar numbers of eggs were consumed by each lacewing larva searching whole plants of either Bt (ave. 15.5 ± 0.6 of 49 over 24 h) or conventional (ave. 13.6 ± 1.1 of 49 over 24 h). On conventional plants over 72 h, survival of H. armigera larvae was 72.8% and decreased to 37.7% when lacewings were present, giving a net consumption rate of 35.1% (8.6 prey per predator over 72 h). On Bt cotton plants, 13.6% of the H. armigera larvae survived after 72 h and this decreased to 1.7% when lacewings were present. This combination of mortality factors operated synergistically. Helicoverpa armigera larvae moved to fruiting structures on conventional or Bt cotton but failed to survive in the squares (young flower buds) when the impacts of Bt and lacewings were combined. The removal of first to second instar H. armigera larvae from squares of Bt cotton by predators has the potential to reduce immediate pest damage and, perhaps more importantly, remove potentially Bt‐resistant genotypes.     

Determining the major Bt refuge crops for cotton bollworm in North China

Evaluation of the effectiveness of refuge strategies involved in cotton bollworm Bt resistance management would be aided by technologies that allow monitoring and quantification of key factors that affect the process under field conditions. We hypothesized that characterization of stable carbon and nitrogen isotopes in adult bollworm, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) moths may aid in determining the larval host that they developed upon. We found moths reared from larvae fed on peanut, soybean or cotton, respectively, could be differentiated using isotopic analyses that also corresponded to their respective larval host origins. These techniques were then used to classify feral second‐generation bollworm moths caught in Bt cotton (Gossypium hirsutum) fields into different populations based on their isotopic signatures. In 2006–2007 feral moths captured in Bt cotton fields predominantly correlated with the peanut (Arachis hypogea) having served as their larval host, indicating this is the most important refuge crop for Bt‐susceptible bollworm individuals (providing 58%?64% individuals) during independent moth peaks for the second generation in North China. The remaining feral moths correlated with soybean (Glycine max) (0?10%); other C₃ plant (20%?22%) and non‐C₃ plant (12%?14%) host types also provided some Bt‐sensitive moths. Field observations showed that peanut constitutes the primary refuge crop contributing to sustaining Bt‐susceptible moths dispersing into cotton in North China. These results suggest that peanut may be a more effective refuge to sustain Bt‐susceptible bollworm individuals and reduce the risk of development of a Bt‐resistant biotype.     

Responses of Thrips tabaci to odours of herbivore‐induced cotton seedlings

Herbivore‐induced changes in plants have been widely viewed as defensive responses against further insect attack. However, changes in plants as a consequence of herbivore feeding can elicit various responses in herbivores; these are variable, context dependent, and often unpredictable. In this laboratory study, the responses of Thrips tabaci Lindeman (Thysanoptera: Thripidae) to volatiles emitted by intact and herbivore‐damaged or mechanically damaged cotton seedlings [Gossypium hirsutum L. (Malvaceae)] were investigated in dual‐choice olfactometer assays. Thrips tabaci showed increased attraction to seedlings subject to foliar mechanical damage and those with foliar damage inflicted by conspecifics or Tetranychus urticae Koch (Acari: Tetranychidae), upon which it preys. However, T. tabaci did not discriminate between intact seedlings and those with foliar damage inflicted by Helicoverpa armigera Hübner (Lepidoptera: Noctuidae), two other species of thrips, Frankliniella schultzei Trybom and Frankliniella occidentalis Pergrande (Thysanoptera: Thripidae), or those with root damage inflicted by Tenebrio molitor L. (Coleoptera: Tenebrionidae). Attraction of T. tabaci was also affected by herbivore density on damaged plants. That is, seedlings damaged by higher densities of T. urticae or T. tabaci were more attractive than seedlings damaged by lower densities of the corresponding arthropod. Although attracted to plants damaged by conspecifics or T. urticae, T. tabaci showed greater attraction to seedlings damaged by T. urticae than to seedlings damaged by conspecifics. Results are discussed in the context of the responses of F. schultzei and F. occidentalis to herbivore‐induced cotton seedlings, highlighting the complexity, variability, and unpredictability of the responses of even closely related species of insects to plants under herbivore attack.