湖北棉区转Bt基因棉对棉铃虫的控制作用

2000-2001年通过田间系统调查,表明转Bt基因棉（品种为GK19）在湖北江汉平原棉区对棉铃虫抗性稳定。试验设3个处理：转Bt基因棉化防田（使用化学农药控制害虫）、转Bt基因棉自控田（依靠天敌控制害虫）及常规棉对照田（利用综合防治措施控制害虫）。从棉铃虫的第2代到第5代整个发生期内,即使在不进行化学防治的情况下,棉铃虫在Bt棉田的发生量也保持在极低的水平(最高百株虫量为12头)。室内饲养结果表明,转Bt基因棉对棉铃虫的生长发育（幼虫体重、蛹重）有较为明显的影响,使6龄幼虫体重减少25.6%,蛹重减少18.2%。棉铃虫幼虫取食转Bt基因棉组织后,发育迟缓,相对于常规棉喂养的整个发育历期延长17 d,使棉铃虫在田间的危害减少至少一个世代。另外,接虫试验表明,棉铃虫幼虫在常规棉上的取食时间是转Bt基因棉株上的6.1倍,极大地减轻了棉铃虫的危害程度。     

田间条件下不同诱导棉花挥发性物质的鉴定

采用人工机械损伤(处理A)、棉铃虫为害(处理B)、棉铃虫为害+棉铃虫体(处理C)和水杨酸诱导(处理D)4种处理方式,对田间现蕾期棉花植株挥发性化合物进行了定性和定量研究.结果表明：有30多种棉株挥发性化合物被定性检出,对其中10种主要挥发性物质进行定量分析,田间棉株挥发的绿叶气味化合物有3-己酮、2-己酮和3-己醇等;单萜类化合物有α-蒎烯、β-蒎烯、β-月桂烯、3,7-二甲基,1,3,6-辛三烯等;脂肪族化合物有丙酸丁酯、乙酸戊酯、乙酸丁酯和3-甲基丁酸乙酯;芳香族化合物有苯乙酮、苯甲醛等.处理C和处理B的棉花挥发信息化合物种类和释放量均明显多于对照,这两个处理检测到3,7-二甲基,1,3,6-辛三烯;而处理A和处理D未检测到该化合物.与对照相比,处理A挥发物在种类和含量上差异不显著.在田间,处理B、处理C和处理D均能诱导促进棉株挥发物的释放.     

转Bt基因抗虫棉田棉铃虫消长规律及危害特点

通过对转 Bt基因抗虫棉田棉铃虫的系统调查研究表明 :(1)转 Bt基因抗虫棉对棉铃虫蛾、卵无抗性表现 ,田间落卵量与常规棉田无明显差异 ,相反 ,二代棉铃虫百株累计卵量倒比常规棉处理区增加了 2 16～ 2 74粒 ,增长幅度为 30 %～ 38% ;(2 )转 Bt基因抗虫棉对二代棉铃虫幼虫的抗性及控制作用较强 ,基本不用进行药剂防治 ,但对三代棉虫幼虫的抗性控制效果明显下降 ,田间 3龄以上幼虫平均达2 0～ 4 0头 /百株 ,多者达 6 0头以上 ,仍需进行必要的药剂防治 ;(3)棉铃虫在转 Bt基因抗虫棉株上的危害特点与常规棉株有明显的差异 ,表现在幼虫和危害症状均较隐蔽 ,不易被发现。     

转Bt基因棉花及其受体品种主要挥发性物质的测定

应用顶空进样气质联机系统（Headspace-GC-MS)测定了转Bt基因棉花及其对照亲本主要挥发性物质。结果表明棉花营养器官和繁殖器官的主要挥发性物质的差异,棉花植株现蕾期前,叶片中挥发性物质以α-蒎烯为主,而现蕾后,蕾、花和铃中主要以β-月桂烯为主。研究初步发现,外源Bt杀虫蛋白表达对棉花自身主要挥发性物质的合成不会造成不利影响。     

棉铃虫对转Bt基因棉花的行为反应

棉铃虫Helicoverpa armigera(Hübner)是转Bt基因棉花(Bt棉)最重要的靶标害虫之一。害虫对Bt棉的抗性问题是Bt棉产业持续健康发展的最主要威胁。规避行为可以减少害虫与Bt棉的接触而降低生理抗性选择压力。在本研究中,通过比较棉铃虫对Bt棉与常规棉的行为反应评估了棉铃虫对Bt棉的行为规避能力。产卵选择结果显示,棉铃虫成虫在Bt棉上的落卵量显著低于常规棉。幼虫实验结果显示,棉铃虫初孵幼虫在Bt棉植株上的居留时间显著短于在常规棉上的居留时间。无选择条件下,1龄幼虫在对Bt棉叶的取食空洞数以及总取食量均显著低于常规棉叶。综合以上的结果,认为棉铃虫对Bt棉具有一定的行为规避能力,这可能是延缓棉铃虫对Bt棉产生抗性的因素之一。本研究能够帮助有效地预测靶标害虫对Bt作物的抗性风险。     

转Bt基因棉田节肢动物群落营养层及优势功能团的组成与变化

1998年在河北南皮棉区转Bt基因棉棉田和常规棉综防田的生物群落变化的研究表明,与常规棉综防田相比,转Bt基因棉棉田的节肢动物种类增加30.7%,功能团多样性增加66.9%,天敌类和中性节肢动物类的丰富度分别增加97.6%和158.0%,害虫类的丰富度减少45.7%。就功能团的种群数量来看,主要害虫棉铃虫、蚜虫和蓟马种群数量分别减少98.9%、69.0%和72.6%;而次要害虫害蝽、粉虱、叶蝉类昆虫的种群数量分别增加了2.2、3.2和14.8倍,棉花生长中期,转Bt基因棉棉田中的基位物种丰富度大于中位和顶位物种;常规棉田中的差异则更大,出现天敌控制“空缺”。综合分析表明,转Bt基因棉棉田生态系统比常规棉综防田稳定;转Bt基因棉棉田的中性节肢动物和次要害虫在害虫－天敌的营养链中起到了重要的调控作用。     

盐城棉区棉铃虫田间种群对Bt蛋白的抗性基因频率

【背景】转Bt基因抗虫棉已经在我国进行了近20年的大规模商业化种植,产生了显著的经济和环境效益。但是,靶标害虫棉铃虫的抗性是转Bt基因抗虫棉产业健康发展所面临的最大问题,而抗性监测是解决这一问题的必要管理措施。盐城市是江苏省转基因抗虫棉的主产区,但有关该地区棉铃虫对转Bt基因抗虫棉的抗性基因频率未见报道。【方法】于2012年在盐城三龙镇和东台镇棉区采集田间棉铃虫种群,检测了初孵幼虫对花铃期转Bt基因抗虫棉中30幼嫩叶片的敏感性,用区分剂量法检测了2龄幼虫对Bt蛋白的抗性基因频率。【结果】取食转Bt基因抗虫棉叶片后,棉铃虫初孵幼虫在9 d内全部死亡;三龙镇和东台镇棉铃虫2龄幼虫对Bt蛋白的抗性基因频率分别为7.6×10-3和6.9×10-3。【结论与意义】目前,盐城棉区的棉铃虫对转Bt基因抗虫棉仍保持很高的敏感性,棉铃虫种群对Bt蛋白的抗性基因频率没有发生显著变化,但仍需持续监测。     

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。     

新疆棉区转Bt基因棉对棉铃虫生物学的影响

棉铃虫Helicoverpaarmigera高龄幼虫取食转Bt基因棉花组织后 ,化蛹率、羽化率、蛹重、体长均有显著下降 ,在转Bt基因棉花上棉铃虫的取食行为也受到较大的影响 ,表现为取食次数明显减少、吐丝下垂次数明显增加 ;无论是转Bt基因棉花还是常规棉花 ,棉铃虫 3龄幼虫主要分布于繁殖器官上 ,在转Bt基因棉花各繁殖器官上的分布概率为 :花 >棉铃 >棉蕾 ,常规棉花上分布概率为 :棉蕾 >棉铃 >花 ;棉铃虫高龄幼虫取食转Bt基因棉花各组织 ,成虫羽化后产卵量、卵孵化率均有明显下降 ;在较低的棉铃虫虫口密度下 ,转Bt基因棉花对棉铃虫有一定的产卵排趋性。     

转(Bt Cry1Ac+CP_4EPSPS)基因抗虫抗草甘膦棉花对草甘膦的耐受性研究

本研究比较了转Bt Cry1Ac+CP_4EPSPS基因抗棉铃虫抗草甘膦棉花与常规棉花在新疆棉区对草甘膦的耐受性差异。两年的研究结果表明,转Bt Cry1Ac+CP_4EPSPS基因抗棉铃虫抗草甘膦棉花对草甘膦有较好的耐受性,而对照棉花中棉所49对草甘膦耐受性较差。苗期喷施草甘膦后转基因抗棉铃虫抗草甘膦棉花生长发育没有受到影响,而对照棉花中棉所49喷施草甘膦后生长发育受到了明显的影响,个别植株甚至死亡。转Bt Cry1Ac+CP_4EPSPS基因抗棉铃虫抗草甘膦棉花株高、真叶数、蕾数、产量等指标与对照相比差异显著。转Bt Cry1Ac+CP_4EPSPS基因棉花对草甘膦的耐受程度显著高于非转基因棉花。草甘膦对转基因抗草甘膦棉花无负面影响。     

Antennal response of cotton bollworm (Heliocoverpa armigera) to volatiles in transgenic Bt cotton

Abstract:  Volatile compositions in transgenic Bt cotton variety 'GK-97' and its parental variety 'Simian No. 3' were analysed, and antennal response of cotton bollworm ( Heliocoverpa armigera ) to the volatiles were investigated. Volatile components in Bt cotton and regular cotton were almost same, except for absence of a sesquiterpene and a minor unknown compound in regular cotton. The results of gas chromatography-electroantennographic detection (GC-EAD) experiments showed that cotton bollworm antennae responded to seven compounds and two minor unknown compounds in Bt volatiles, among which concentrations of α -pinene and β -pinene in Bt cotton were relatively higher than those in regular cotton, and one minor unknown compound was absent in regular cotton.     

转Bt基因抗虫棉与常规棉中几种同工酶的比较——转基因植物安全性评价生理指标初探

经基因改造的转基因抗虫棉在我国已开始进入大规模商业化应用,对此进行安全性评价是一个十分重要的生态学问题。本文比较了抗虫棉和常规棉中过氧化物酶、多酚氧化酶、酯酶等几种重要同工酶的酶活和酶谱的差异,发现在一定生长期中,抗虫棉的可溶性过氧化物酶活性显著高于常规棉,酯酶的酶活和酶谱在两种棉花中也存在着差异。认为这些差异可能影响抗虫棉的次生代谢,进而导致一些综合的生态学效应,提出植物生理上的改变也是转基因植物安全性评价研究的重要内容。     

Leaf surface factors of transgenic Bt cotton associated with the feeding behaviors of cotton aphids: A case study on non-target effects

The present paper reports case study results of the risk assessment of transgenic Bt cotton on a non-target pest, cotton aphid, Aphis gossypii. Several types of techniques, i.e., electrical penetration graph (EPG), light and electron microscopy, bioassays and chemical analysis, were applied to investigate physical and chemical leaf factors of 2 transgenic Bt cotton lines (GK12 and GK19) and their pa-rental non-Bt cotton line (Simian3) associated with searching and feeding behaviors of cotton aphids on leaves or leaf extracts of cotton plants. EPG results showed that there were some differences among behaviors of cotton aphids on 2 Bt cotton and 1 non-Bt cotton lines. Cotton aphids performed similarly to leaf surface extracts from 3 cotton lines; and leaf surface chemicals, mainly volatiles and waxes, were almost identical in the components and concentrations among the cotton lines. However, three cotton lines were quite different from each other in the densities of certain kinds of covering trichomes. Therefore, the relationships between the physical characteristics and the searching behaviors of cotton aphids on the three cotton lines were constructed as the regression equations. Glandular trichomes and covering trichomes with 5 branches influenced the cotton aphids' searching behaviors effectively; and other trichomes with other branches affected aphids in varying ways. These results demonstrated that leaf surface physical factors of transgenic Bt cotton lines different from their parental non-Bt line could affect the penetration behaviors of non-target cotton aphids. Cotton aphids penetrate and feed more easily on two Bt cotton lines than on the non-Bt cotton line.     

Influences of elevated CO2 and pest damage on the allocation of plant defense compounds in Bt‐transgenic cotton and enzymatic activity of cotton aphid

Abstract Plant allocation to defensive compounds by elevated CO₂‐grown non‐transgenic and transgenic Bt cotton in response to infestation by cotton aphid, Aphis gossypii (Glover) in open‐top chambers under elevated CO₂ were studied. The results showed that significantly lower foliar nitrogen concentration and Bt toxin protein occurred in transgenic Bt cotton with and without cotton aphid infestation under elevated CO_2. However, significantly higher carbon/nitrogen ratio, condensed tannin and gossypol were observed in transgenic Bt cotton “GK‐12” and non‐transgenic Bt cotton ‘Simian‐3’ under elevated CO_2. The CO₂ level and cotton variety significantly influenced the foliar nitrogen, condensed tannin and gossypol concentrations in the plant leaves after feeding by A. gossypii. The interaction between CO₂ level × infestation time (24 h, 48 h and 72 h) showed a significant increase in cotton condensed tannin concentrations, while the interaction between CO₂ level × cotton variety significantly decreased the true choline esterase (TChE) concentration in the body of A. gossypi. This study exemplified the complexities of predicting how transgenic and non‐transgenic plants will allocate defensive compounds in response to herbivorous insects under differing climatic conditions. Plant defensive compound allocation patterns and aphid enzyme changes observed in this study appear to be broadly applicable across a range of plant and herbivorous insect interactions as CO₂ atmosphere rises.     

Effect of Bt genetic engineering on indirect defense in cotton via a tritrophic interaction

We present a tritrophic analysis of the potential non-intended pleiotropic effects of cry1Ac gene derived from Bacillus thurigiensis (Bt) insertion in cotton (DeltaPine 404 Bt Bollgard® variety) on the emission of herbivore induced volatile compounds and on the attraction of the egg parasitoid Trichogramma pretisoum (Hymenoptera: Trichogrammatidae). Both the herbivore damaged Bt variety and its non-Bt isoline (DeltaPine DP4049 variety) produced volatiles in higher quantity when compared to undamaged plants and significantly attracted the egg parasitoids (T. pretiosum) when compared to undamaged plants. However, Trichogramma pretiosum did not differentiate between the transgenic and nontransgenic varieties, suggesting that the ratios between the compounds released by herbivory damaged -Bt cotton and herbivory damaged-nonBt cotton did not change significantly. Finally, no detrimental effect of the Bt genetic engineering was detected related to the volatile compounds released by Bollgard cotton on the behavior of the natural enemy studied.     

Stylet penetration behaviors of the cotton aphid Aphis gossypii on transgenic Bt cotton

Stylet penetration behaviors of cotton aphids Aphis gossypii Glover on a transgenic cotton line "GK-12" expressing Bt toxic protein of Cry1A (Bt cotton) and a non-Bt conventional cotton line "Simian-3" (CK cotton) were recorded with the direct current electrical penetration graph (DC-EPG) technique. Cotton aphids reared on Bt cotton (abbreviated as Bt-aphids) and its parental non-Bt control line (CK-aphids) for more than 20 generations each, were used for recordings on two cotton lines. Among 47 selected parameters reflecting the activities of aphid stylets within plant tissues, there were eight parameters of CK-aphids showing significant differences between the performances of CK-aphids on Bt cotton and CK cotton, while for Bt-aphids, all the parameters were statistically equal between the performances on the two cotton lines. All parameters with significant differences indicated that CK-aphids could penetrate into Bt cotton more easily, but the phloem saps of Bt cotton were not as good as those of regular cotton for CK-aphids. Based on the present results, we concluded that there were some factors in Bt cotton affecting penetration behaviors of CK-aphids, but it just took several generations for CK-aphids to completely adapt Bt cotton, and Bt-aphids could feed on two cotton lines without difficulty.     

Feeding behaviour of Helicoverpa armigera larvae on insect-resistant transgenic cotton and non-transgenic cotton

Abstract: Feeding behaviour of Helicoverpa armigera Hübner (Lep.; Noctuidae) larvae on non‐transgenic Bacillus thuringiensis (Bt) cotton (Gossypium hirsutum L.), Zhong 30, and transgenic cowpea trypsin inhibitor (CpTI)‐Bt cotton, SGK 321, and non‐transgenic cotton, Shiyuan 321, was investigated in both choice tests and no‐choice tests. The results of choice tests suggested that neonates have the ability to detect and avoid transgenic cotton. In the choice tests of neonates with both transgenic and non‐transgenic cotton leaves, a significantly greater proportion of larvae and higher consumption were observed on non‐transgenic cotton than on the transgenic Bt or CpTI‐Bt cotton. In the choice tests with leaves of two transgenic cotton lines, the proportion of neonates on leaf discs of the two lines was not significantly different, but there was significantly higher consumption on CpTI‐Bt transgenic cotton than that on Bt transgenic cotton. In addition, significantly more neonates were found away from the leaf discs, lower consumption and higher mortality were achieved in the choice test with two transgenic cotton leaves than in the choice tests containing non‐transgenic cotton leaves. Leaves and buds were examined in choice tests of fourth instars. It appeared that fourth instars were found in equal numbers on transgenic and non‐transgenic cotton, except when larvae were exposed to leaves for 3 h. However, the total consumption on transgenic cotton was lower than that of the non‐transgenic cotton, so fourth instars may still have the capacity to detect transgenic cotton and reduce feeding on it, although they showed no preference on either transgenic or non‐transgenic cotton. More larvae were found off diet in the treatments with leaves than that of buds, and the number of injured leaf discs by per fourth instar was significantly higher than that of buds in choice tests, suggesting that leaf is a less preferred organ for H. armigera larvae, elicited more larval movements. Similarly, in no‐choice tests of fifth instars, significantly fewer feeding time and more moving time occurred on leaf than that of bud, boll and petal. When cotton line was considered, compared with non‐transgenic cotton, significantly lower feeding time and higher resting time occurred on the two transgenic cottons. Overall, H. armigera larvae have the ability to detect the transgenic Bt and CpTI‐Bt cottons or the less preferred organs and selectively feed more on the non‐transgenic cotton or the preferred organs, especially the neonates, which have a high capacity for avoiding transgenic cotton.     

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。