转cry1Ac/sck基因抗虫水稻对稻田寄生蜂群落影响的评价

以转cry1Ac/sck双基因抗虫水稻MSA、MSB、MSA4（对照： MH86）及其杂交稻KF6-304（对照： Ⅱ-YM86）为材料,系统地研究了转基因抗虫水稻对稻田寄生蜂群落的影响及生态安全性。结果表明： 在群落水平上,转基因抗虫水稻MSA、MSB、MSA4及杂交稻KF6-304对稻田寄生蜂的物种丰富度、多样性指数、均匀性指数和优势集中性指数的总体情况与时间动态以及个体总数无明显负面影响,但在生长中期转基因稻可降低稻田寄生蜂的个体数量。按寄生蜂功能团分析,MSA与MSA4在水稻生长发育的后期初始,提高卵寄生蜂的数量,MSA、MSB、MSA4及杂交稻KF6-304显著降低以靶标害虫稻纵卷叶螟为寄主的寄生蜂功能团的个体数量,对其他寄生蜂无明显负面影响。     

广西兴安转Bt水稻大田两迁害虫发生动态

以转Bt水稻华恢1号(Cry1Ac/ CryAb融合基因型,简称HH1)及其对照亲本明恢63(简称MH63)稻田两迁害虫稻纵卷叶螟和白背飞虱为研究对象,系统研究转基因抗虫水稻种植下两迁害虫的发生规律及其致害力差异.结果表明:转Bt水稻及其对照亲本上稻纵卷叶螟的落卵量和幼虫发生量无显著差异,但转Bt水稻的卷叶株率和卷叶率都显著低于对照亲本.表明转Bt水稻对靶标害虫稻纵卷叶螟具有较高抗性.转Bt水稻及对照亲本上白背飞虱若虫、成虫及整个种群的发生动态差异不显著,且转Bt水稻对长翅型和短翅型成虫的种群发生影响也不显著.白背飞虱发生高峰期,转Bt水稻上若虫及短翅型成虫发生量均明显高于对照亲本;相反,转Bt水稻上长翅型成虫发生量明显低于对照亲本,且水稻生育后期长翅型成虫雌性比明显低于对照亲本.转Bt水稻大面积商业化种植下其非靶标害虫白背飞虱的发生危害变得更为复杂.     

水稻品种对稻纵卷叶螟抗性的田间鉴定

<正> 稻纵卷叶螟Cnaphalocrocis medinalis(Guenee)原为我国东南部稻区偶发性害虫,六十年代数量上升,七十年代经常大发生,成为水稻生产上的重要害虫。作为综合防治的一个环节,研究水稻品种对稻纵卷叶螟的抗性,筛选鉴定抗虫亲本材料,选育抗虫栽培品种是有现实意义的。Das(1974a)首先报道了稻纵卷叶螟在不同品种稻叶上取食量的差异,其后Das(1974b)、Velusamy(1975)、K.Saivaraj(1979)先后进行了抗稻纵卷叶螟品种的田间筛选。 作者从1977年开始这方面的研究,三年来鉴定了380个水稻品种的田间抗性。现将初步进展小结如下,以供参考。 材料与方法 一、供试水稻品种的来源 供试品种中将近一半是从国际水稻研究所引进的具有各种抗性基因的杂交后代,其余是从亚洲、欧洲、美洲引进的定型品种及湖南省推广的晚稻良种、地方品种。其产地的分布见表1。 二、鉴定方法 鉴定在湖南农学院农场进行。湖南省稻纵卷叶螟一般一年发生五代,以早稻上的第二代     

转cry1Ab/1Ac,cry1C,cry2A基因水稻对田间稻纵卷叶螟及其捕食类天敌的影响

在转Bt基因水稻商业化之前需要评价其对靶标害虫及其天敌的影响.本研究连续2年在3个地点调查了3种转Bt水稻材料和非转基因水稻田间稻纵卷叶螟为害情况,稻纵卷叶螟及其4种捕食类天敌的种群密度、相对丰富度和种群动态.结果表明,水稻材料对卷叶率和稻纵卷叶螟幼虫种群密度有显著影响.非转基因水稻田间卷叶率显著高于3种转Bt基因水稻田.在调查期间,不同地点3种非转基因水稻田间稻纵卷叶螟幼虫的种群密度以及它在植食类功能团内的相对丰富度均显著高于3种转Bt基因水稻田.水稻材料、水稻材料×调查日期、水稻材料×调查年份、水稻材料×调查日期×调查年份对稻纵卷叶螟幼虫的种群动态均有显著影响.但是,3种转Bt基因水稻材料和非转基因水稻田间4种捕食类天敌的相对丰富度、种群密度以及种群动态基本没有显著差异.从上述结果可推论,转Bt基因水稻中的Bt蛋白能明显抑制稻纵卷叶螟的发生,但对4种捕食类天敌没有显著影响.     

转基因抗虫水稻MSA和MSB对非靶标害虫褐飞虱和白背飞虱的影响

本文报道了转SCK+cry1Ac双基因抗虫水稻MSA和MSB对非靶标害虫褐飞虱Nilaparvata lugens 和白背飞虱Sogatella fucifera生物学及田间发生动态的影响。与亲本明恢86相比,MSA对两种飞虱若虫发育历期、初羽化成虫鲜重、羽化率、短翅率、成虫产卵量、卵受精率、单雌子代数及发育进度等生物学指标均无明显影响,而MSB虽然对多数指标影响不明显,但可以引起白背飞虱初羽化雌虫鲜重与短翅率显著下降,还引起褐飞虱初羽化雄虫鲜重明显减轻。在近距离（株间距约 8 cm）寄主选择实验中,褐飞虱对MSA和MSB无明显选择性,而白背飞虱若虫和成虫均明显趋向两种转基因水稻。田间小区实验表明MSB上的稻飞虱（主要为白背飞虱）密度一般低于亲本明恢86,但差异不显著,提示转基因水稻MSB引发非靶标害虫稻飞虱灾变的风险较小。     

转Cry1Ab/Ac基因水稻对稻田浮游动物群落物种多样性的影响

为了解转Bt基因抗虫水稻对稻田浮游动物群落的安全性,本文以转Cry1Ab/Ac基因水稻"华恢1号"(HH1)为试验材料,以其非转基因亲本水稻"明恢63"(MH63)为对照,以浮游动物为指示生物,通过湖南湘潭基地连续3年的大田实验,研究了转Cry1Ab/Ac基因水稻对稻田浮游动物群落物种多样性的影响。结果表明:HH1与MH63两稻田生境浮游动物群落相似性高于0.70,优势种组成相似性高于0.89;浮游动物群落结构各参数的时间动态与总体分析均无显著差异;转Cry1Ab/Ac基因抗虫水稻HH1对稻田浮游动物群落物种多样性无不良影响。     

农杆菌转化获得转B．t．基因水稻及其生物学鉴定

以水稻主栽品种"秀水11"和"春江11"预培养4d未成熟胚为转化受体,经农杆茵LBA4404／pGBI4A2B(含B．t．基因)感染后,筛选出抗性愈伤组织并获得转化植株。其中抗性愈伤组织产生频率达44％～70％,转化植株产生频率达27％～64％。转基因植物总DNA经PCR和Southernblot分子杂交试验表明,T-DNA上的外源基因已整合到水稻的基因组里。对转B．t．基因水稻植株进行了两种水稻主要害虫纵卷叶螟和二化螟的饲虫试验。与对照相比,7d后纵卷叶螟幼虫死亡率达31％～49％,同时纵卷叶螟对转基因水稻叶片的危害程度大大减轻。二化螟在咬食转B．t．基因水稻植株7d后的校正死亡率达15％～71％,30d后多数转基因水稻仍能正常抽穗,表明转基因水稻对上述害虫具有一定抗性。     

水稻三种害虫组合侵害及对籽粒生长的影响

根据江淮稻区水稻害虫发生特点,于1991-1992年在网室和田间采用全组合随机区组设计研究了白背飞虱Sogatella furcifera、稻纵卷叶螟Csapkelocrocis medinalis、褐飞虱 Nilaparvata lugens三种害虫的组合侵害及对籽粒生长的影响。用生长曲线方程拟合籽粒生长过程。由生长曲线方程推导的一些参数分析害虫组合侵害对强、弱势粒生长的影响。分析 结果表明三种害虫组合侵害主效和互作效应均对水稻有显著的影响。随着害虫组合侵害的加重,最大生长速率V_max、旺盛生长期积累的干物重GT等参数变小。田间和网室试验结果表 明害虫组合侵害对弱势粒的影响大于对强势粒的影响。     

取食转Bt基因水稻对稻纵卷叶螟幼虫中肠的组织病理学效应

利用透射电镜观察了稻纵卷叶螟 Cnaphalocrocis medinalis（Guenée）幼虫取食转Bt基因水稻后中肠的组织病理变化。结果表明：稻纵卷叶螟幼虫取食转cry1Ab基因水稻后,中肠上皮细胞的线粒体先发生形态变化,随连续取食时间的延长线粒体出现凝聚、内嵴稀疏、空泡化等,在后期还呈凝聚态随突起脱落或沿杯腔边沿单一排列。内质网的变化也很明显,病变过程中伴随着粗糙内质网的肿胀、核糖体脱落,粗糙内质网增多等现象。细胞核的变化较小,在处理后期出现细胞核拉长、核仁聚集等变化。组织病变程度不一,有的细胞在病变早期就出现了空泡化。     

转cry1 Ac/cpti基因水稻对土壤氨氧化细菌群落组成和丰度的影响

【背景】氨氧化细菌是驱动硝化作用的关键微生物,其群落多样性变化对土壤氮素转化具有重要意义。转基因作物可能通过根系分泌物和植株残体组成的改变对土壤微生物群落产生影响。【方法】本研究通过田间定位试验,利用特异引物进行PCR-DGGE（聚合酶链反应—变性梯度凝胶电泳）和荧光定量PCR,分析了种植转cry1 Ac/cpti双价抗虫基因水稻第3、4年土壤中氨氧化细菌群落组成和丰度的变化。【结果】水稻各生育期（分蘖期、齐穗期和成熟期）内,转cry1 Ac/cpti基因杂交稻Ⅱ优科丰8号（GM）的土壤氨氧化细菌16S rRNA基因群落组成、多样性指数与其对应的非转基因杂交稻Ⅱ优明恢86（CK）间均没有显著差异;以DGGE条带为基础的氨氧化细菌群落组成的冗余分析（RDA）显示,GM和CK的土壤氨氧化细菌群落组成只与水稻生育期存在显著相关性（P=0.002和0.018）;同时,水稻各生育期内土壤氨氧化细菌16S rRNA基因丰度在GM和CK间也没有显著差异,但均随水稻生长而变化且在齐穗期达到最高（P〈0.05）。【结论与意义】稻田土壤氨氧化细菌的群落组成与丰度在水稻不同生育期存在差异,但在转cry1 Ac/cpti基因水稻和非转基因水稻间没有显著差异,即一定时期内种植转cry1 Ac/cpti抗虫基因水稻不会影响土壤氨氧化细菌的群落组成和丰度。     

Baseline susceptibility of Cnaphalocrocis medinalis (Lepidoptera: Pyralidae) to Bacillus thuringiensis toxins in China

Geographic variability in susceptibility of field-collected Cnaphalocrocis medinalis (Guenée) (Lepidoptera: Pyralidae) to Bacillus thuringiensis (Berliner) was studied to establish a geographic baseline for comparison of future population responses to the increased use of B. thuringiensis-based insect control products. Colonies of C. medinalis were established from 10 populations collected from 10 different provinces in the major rice (Oryza savita L.)-growing regions of China. Populations were evaluated for their susceptibility to Cry1Ac and Cry1Ab endotoxin proteins with the leaf-dip bioassay. The LC50 values to Cry1Ac and Cry1Ab were estimated for the 10 populations. Bioassay results indicated that the ranges of LC50 in the second instars of C. medinalis to Cry1Ac and Cry1Ab were from 3.77 to 208.22 mg ([AI])/liter and 0.22-7.05 mg ([AI])/liter, respectively. The relative ratios in susceptibility between the most susceptible and the most tolerant populations were beyond 50-fold for Cry1Ac and 30-fold for Cry1Ab. Moreover, there was a significant positive correlation between susceptibilities to the two toxins tested, suggesting that insect populations that are relatively tolerant to one protein are also relatively tolerant to the other.     

稻田害虫和捕食性节肢动物群落结构和动态

广西南宁市郊稻田害虫和捕食性节肢动物群落,由46科159种组成,其中害虫21科65种;捕食性昆虫12科39种;蜘蛛13科55种。早、晚稻构成群落的物种基本相似,但各物种的数量结构及优势种差异明显。文中分析了晚稻害虫、捕食性昆虫和蜘蛛亚群落的垂直分层、时空格局及其数量变动的原因。     

生境异质度对稻田捕食性天敌及水稻害虫的生态调节有效性

【目的】探索生境高异质度对稻田捕食性天敌及水稻害虫的生态调节有效性,了解这种策略是否会引起其他植食性昆虫成为水稻重要害虫的风险。【方法】2017-2018年连续2年种植单季稻,在稻田边缘种植花生与大豆,构建高异质性边缘生境稻田(rice paddy with high heterogeneous marginal habitats, HHR),调查HHR稻田与简单低异质性边缘生境稻田(rice paddy with low heterogeneous marginal habitats, LHR)中捕食性天敌与水稻害虫功能团的发生规律与相关性,计算益害比。【结果】2017年在HHR稻田中采集到捕食性天敌40种,1 667头;在LHR稻田中采集到捕食性天敌30种,991头。2018年在HHR稻田中采集到捕食性天敌33种,1 384头;在LHR稻田中采集到捕食性天敌34种,1 031头。HHR与LHR两类稻田中获得的捕食性天敌群落重要值P_i≥0.01的物种相似度很高,优势种相似。2017年HHR稻田的捕食性天敌物种丰富度明显高于LHR稻田,这种差异主要由群落重要值P_i<0.01的种类引起。2018年两类稻田的捕食性天敌物种丰富度没有差异。单位样方面积内的捕食性天敌个体数量,HHR中明显高于LHR,这种差异在2017年的分蘖期与成熟期达显著水平(P<0.05),在2018年的开花期极显著(P<0.01)。2017年在HHR中采集到水稻害虫22种,637头;在LHR中采集到水稻害虫19种,743头;物种相似性系数0.88。2018年在HHR中采集到水稻害虫16种,1 011头;在LHR中采集到水稻害虫16种,2 014头;物种相似性系数0.75;主要害虫物种组成结构相同。水稻害虫数量发生的时间动态分析表明,在分蘖期,虽然HHR稻田中的水稻害虫数量明显高于LHR(P<0.05),但此期害虫的数量不多,发生较轻。在孕穗期、开花期与成熟期,HHR稻田中的水稻害虫数量明显低于LHR稻田,这种差异在2017年的成熟期与2018年的孕穗期极显著(P<0.01),在2018年的成熟期差异显著(P<0.05)。2017年HHR和LHR中个体数量益害比N_pi值分别为2.62和1.33;2018年分别为1.37和0.51。【结论】具有高异质性边缘生境的稻田,能提高系统对捕食性天敌物种的涵养潜力,显著提高稻田捕食性天敌个体数量,提高益害比,具有更好的控制害虫的物质基础,促进捕食性天敌对水稻害虫的生态控制效能,不会引起其他植食性昆虫演变为水稻重要害虫风险,可为保护稻田生态系统天敌发挥生态效能提供可借鉴的策略与方法。     

Interaction of gene-cloned and insect cell-expressed aminopeptidase N of Spodoptera litura with insecticidal crystal protein Cry1C

Insecticidal toxins produced by Bacillus thuringiensis interact with specific receptors located in the midguts of susceptible larvae, and the interaction is followed by a series of biochemical events that lead to the death of the insect. In order to elucidate the mechanism of action of B. thuringiensis toxins, receptor protein-encoding genes from many insect species have been cloned and characterized. In this paper we report the cloning, expression, and characterization of Cry toxin-interacting aminopeptidase N (APN) isolated from the midgut of a polyphagous pest, Spodoptera litura. The S. litura APN cDNA was expressed in the Sf21 insect cell line by using a baculovirus expression system. Immunofluorescence staining of the cells revealed that the expressed APN was located at the surface of Sf21 cells. Treatment of Sf21 cells expressing S. litura APN with phosphatidylinositol-specific phospholipase C demonstrated that the APN was anchored in the membrane by a glycosylphosphatidylinositol moiety. Interaction of the expressed receptor with different Cry toxins was examined by immunofluorescence toxin binding studies and ligand blot and immunoprecipitation analyses. By these experiments we showed that the bioactive toxin, Cry1C, binds to the recombinant APN, while the nonbioactive toxin, Cry1Ac, showed no interaction.     

Interaction of Gene-Cloned and Insect Cell-Expressed Aminopeptidase N of Spodoptera litura with Insecticidal Crystal Protein Cry1C

Insecticidal toxins produced by Bacillus thuringiensis interact with specific receptors located in the midguts of susceptible larvae, and the interaction is followed by a series of biochemical events that lead to the death of the insect. In order to elucidate the mechanism of action of B. thuringiensis toxins, receptor protein-encoding genes from many insect species have been cloned and characterized. In this paper we report the cloning, expression, and characterization of Cry toxin-interacting aminopeptidase N (APN) isolated from the midgut of a polyphagous pest, Spodoptera litura. The S. litura APN cDNA was expressed in the Sf21 insect cell line by using a baculovirus expression system. Immunofluorescence staining of the cells revealed that the expressed APN was located at the surface of Sf21 cells. Treatment of Sf21 cells expressing S. litura APN with phosphatidylinositol-specific phospholipase C demonstrated that the APN was anchored in the membrane by a glycosylphosphatidylinositol moiety. Interaction of the expressed receptor with different Cry toxins was examined by immunofluorescence toxin binding studies and ligand blot and immunoprecipitation analyses. By these experiments we showed that the bioactive toxin, Cry1C, binds to the recombinant APN, while the nonbioactive toxin, Cry1Ac, showed no interaction.     

转Cry1Ac+Cry2Ab棉花生长势及其对棉田节肢动物物种丰富度的影响

【背景】转基因棉花在商业化种植之前,必须评价其环境安全性。其中新型棉花材料的生存竞争能力和对物种丰富度的影响是评价的重要内容。【方法】以转Cry1Ac＋Cry2Ab基因棉为试验材料,转Cry1Ac棉花中棉所41和非转基因棉花中棉所49为对照品种,分别于2014年5~9月对棉花株高、主茎叶片数、叶绿素含量、比叶面积、果枝数、蕾铃数等生长参数进行比较,同时对二代、三代和四代棉铃虫发生期棉田物种丰富度进行系统调查。【结果】转Cry1Ac＋Cry2Ab棉花的生长势与转Cry1Ac棉花和非转基因棉花基本相当,没有表现出明显的竞争优势;产量构成参数在成铃和脱落等方面比非转基因棉表现出良好的优势。对棉田节肢动物物种丰富度的影响表明,转Cry1Ac＋Cry2Ab棉花对靶标害虫棉铃虫具有良好的控制效果,对主要刺吸性害虫棉蚜、棉蓟马、烟粉虱、绿盲蝽与天敌龟纹瓢虫、草间小黑蛛、草蛉和小花蝽等的种群丰富度在个别时期有所影响,但总体上与转Cry1Ac棉田和非转基因棉田没有显著性差异。【结论与意义】转Cry1Ac＋Cry2Ab棉花无竞争优势,但目标性状优势较好;对棉田节肢动物物种丰富度无明显影响。研究结果为新型转Cry1Ac＋Cry2Ab棉花对棉田环境安全方面的研究进一步补充了内容,为转基因棉花的环境安全评价提供科学数据。     

Impacts of nitrogen fertilizer on major insect pests and their predators in transgenic Bt rice lines T2A‐1 and T1C‐19

Nitrogen is a critical factor for plant development and nitrogen input is one of the important tactics to enhance the development and yield of crops. Nevertheless, nitrogen input could influence the occurrence of insects positively or negatively. Nitrogen is also one of the main elements composing the insecticidal crystal (Cry) protein. Cry protein production could affect nitrogen partitioning in Bt plants and as such nitrogen input may influence insect pest management in transgenic Bt rice, Oryza sativa L. (Poaceae). To test this possibility, we evaluated the impacts of nitrogen regimes on the main insect pests and their predators on two Bt rice lines, T2A‐1 and T1C‐19, expressing Cry2A and Cry1C, respectively, and their non‐transgenic parental counterpart MH63. The results showed that Cry proteins with different nitrogen regimes have enough insecticidal activity on rice leaffolder, Cnaphalocrocis medinalis Guenée (Lepidoptera: Crambidae), in both laboratory and field experiments. Laboratory studies indicated that relevant parameters of ecological fitness in brown planthopper, Nilaparvata lugens (Stål) (Hemiptera: Delphacidae), a non‐target insect pest, were significantly affected by nitrogen input both on Bt and MH63 rice lines. Nymphal survival, female adult longevity, and egg hatchability in N. lugens differed significantly among rice varieties. The experiments conducted in rice fields also demonstrated that nitrogen was positively correlated with the abundance of N. lugens on Bt rice, similar to that on MH63 rice. The abundances of two predators – the wolf spider Pirata subpiraticus (Boesenberg & Strand) (Araneae: Lycosidae) and the bug Cyrthorhinus lividipennis Reuter (Hemiptera: Miridae) – were significantly affected by rice growth stages but not by nitrogen input and rice varieties. In conclusion, the above results indicate that high nitrogen regimes for Bt rice (T2A‐1 and T1C‐19) and non‐Bt rice (MH63) cannot facilitate the management of insect pests.