土壤真菌群落和潜在功能对施加外源Bt毒素的响应

【目的】转Bt基因植物和Bt菌生物农药释放的Bt毒素是一类具有生物毒性的潜在环境外源污染物，Bt毒素环境行为和生态效应是转基因植物和植物用转基因微生物安全风险评价的重要内容，但是外源Bt毒素对土壤真菌群落和潜在功能的影响还不清楚。【结果】本研究发现，Bt毒素施加量和培养时间均可以显著影响土壤真菌群落组成，且随着Bt毒素施加量增加和土壤培养时间延长，土壤真菌群落差异性逐渐变大。施加Bt毒素提高了土壤真菌群落香农指数和关联网络的负相关性比例及模块数，因而没有对土壤真菌群落的多样性和稳定性产生负面影响。【结论】上述结果表明，评估Bt毒素的环境行为及微生态效应要关注Bt毒素施加量及其长期影响。随着Bt毒素施加量增加，Phymatotrichopsis、Homalogastra、Geosmithia和Apiotrichum等真菌以及参与蛋白质降解、碳素代谢和磷素代谢的功能基因编码酶相对丰度显著升高，推测上述真菌物种和潜在功能参与了Bt毒素在土壤中的降解和转化过程。研究结果为转Bt基因植物、Bt重组菌生物农药以及Bt毒素的生态安全风险评价提供了科学参考和理论依据。     

转Bt基因作物对丛枝菌根真菌的影响研究进展

在过去的十年里，世界范围内转基因作物尤其是抗虫性转Bt基因作物的品种和种植面积迅速增加。同时，转Bt基因作物的环境安全性评价问题成为人们关注和研究的热点。丛枝菌根真菌（AMF）是生态系统中普遍存在的土壤微生物，能与绝大多数植物种类形成共生关系，在农业生态系统中起重要作用。转Bt基因作物环境释放后，其与AMF问的共生关系是否受所转入Bt基因的影响，以及影响机制需要及时研究。为此，综述了转Bt基因作物与AMF共生特征方面的研究进展，并根据Bt毒素发生的空间和时间规律提出了危害机制以及转Bt基因植物的规模化种植将降低农田系统中的AMF的生物多样性的观点。     

苏云金芽孢杆菌Cry1Aa和Cry3A结构域编码区的融合

苏云金芽孢杆菌（Bacillus thuringiensis,Bt)毒素蛋白由3个不连续的结构域（domains)组成，其中结构域I为膜跨越区，与膜孔道形成有关，结构域Ⅱ，Ⅲ与受体结合有关。利用基因重组技术以鳞翅目昆虫具专一活性的毒素蛋白Cry1Aa与专一性有关的结构域编码区与对鞘翅目昆虫具专一活性的毒素蛋白Cry3A编码基因融合，构建成融合表达载体，为进一步构建Bt工程菌和Bt转基因植物奠定基础。     

转基因棉种植对土壤氧化还原酶活性的影响

转Bt基因棉和转Bt+CpTI基因棉及相应非转基因棉的盆栽试验研究表明,转基因作物生长30天时可向土壤中释放Bt杀虫晶体蛋白,而双抗棉种植时CpTI杀虫晶体蛋白的释放量与作物品种有关;转基因棉种植对土壤过氧化氢酶、脱氢酶和硝酸还原酶的活性影响研究表明,生长30天时,与等价基因系非转基因棉相比,转Bt基因棉和双抗棉A的种植并未使三种酶活性发生显著变化,而双抗棉B的种植使土壤脱氢酶活性显著下降。从杀虫晶体蛋白的释放和对酶活性的影响来看,双抗棉B的种植对土壤的生物活性扰动更大。     

土壤酶活性对大田单季种植转Bt基因及转双价棉花的响应

利用田间试验,以转Bt基因棉花Z30、转双价（Bt+CpTI）棉花SGK321及其相应的等价基因Z16、SY321作为供试对象,研究棉花种植后对土壤水解/氧化还原酶类活性的影响。结果表明,转Bt棉花及转双价棉花的种植对各种土壤酶活性的影响不一致,转Bt棉花及转双价棉花种植显著降低了土壤蛋白酶和脱氢酶活性(P 0.05)。转Bt棉花还显著降低了土壤脲酶活性(P 0.05);转双价棉花种植同时显著降低土壤磷酸二酯酶、β-葡萄糖苷酶、过氧化氢酶和硝酸还原酶活性(P 0.05)。棉花品种及转基因行为（转入基因类型）均未给土壤酸性磷酸单酯酶和FDA活性带来影响;土壤芳基硫酸酯酶所受的影响主要来自于棉花品种自身。     

转Bt基因棉秸秆还田利用对土壤肥力的影响

随着转基因的快速发展,大量转Bt棉秸秆的合理利用和处理是不可忽视的重要课题之一。为明确Bt棉秸秆还田利用的可行性和安全性,本研究以不同抗虫转Bt基因棉和常规棉花‘泗棉3号’为研究材料,在分别种植1、2年后将秸秆机械粉碎后原位还田,测试土壤中Bt蛋白残留量、土壤酶活性及养分含量的变化,分析Bt棉秸秆原位还田对土壤肥力特性的影响。研究结果表明,秸秆还田40 d后, Bt棉样地土壤中Bt残留蛋白检测值较低,均与非转基因棉样地无显著性差异。棉秸秆还田后,土壤脲酶、蔗糖酶、蛋白酶、多酚氧化酶、过氧化氢酶、碱性磷酸酶活性皆较秸秆还田前增加,但土壤纤维素酶活性较之前降低。棉秸秆还田使土壤中有机质、有效磷、碱解氮、速效钾和全氮等养分含量及pH明显增加,而Bt抗虫棉与常规棉秸秆还田后对土壤肥力的影响不存在显著差异。对土壤综合肥力指数评价结果表明,秸秆还田对土壤肥力提升与Bt棉抗虫水平无关,土壤肥力指数在两年间由Ⅲ级水平上升至Ⅱ级水平。综上, Bt棉花秸秆还田不会造成土壤综合肥力降低,相反能有效提升土壤肥力;同时还田利用措施可对转基因植株有效灭活,与转基因植物秸秆利用和无害化处理要求相契合。生产中用于Bt转基因棉花秸秆利用和处理在一定程度上是安全可行的。     

转Bt基因棉叶对土壤微生物多样性的影响

应用Biolog方法研究了转Bt基因棉粉碎叶腐解对土壤微生物群落结构功能多样性的影响。取腐解10d、25d、40d、55d、70d土样分析土壤微生物群落多样性指数及土壤微生物对聚合物、胺类、氨基酸、糖、羧酸和其他类碳源利用情况。结果表明:在腐解过程中,转Bt基因棉粉碎叶土壤微生物群落丰富度下降,群落多样性显著降低,而群落优势集中性明显提高;转Bt基因棉粉碎叶影响了土壤微生物群落对碳源的利用程度,表现为可显著增加对糖类、胺类和氨基酸类碳源的利用,初期显著降低对羧酸类碳源的利用,对聚合物类和其他类碳源的利用率无显著影响;主成分分析表明转Bt基因棉粉碎叶对土壤微生物群落原有结构功能影响具有持续性。     

转Bt基因棉粉碎叶对土壤微生物群落功能多样性的影响

采用Biolog GN2微平板法,研究转Bt基因棉粉碎叶添加到土壤后,对土壤微生物群落功能多样性的影响。结果表明,在转Bt基因棉粉碎叶添加到土壤后,第10天和第40天的平均颜色变化率(AWCD)显著提高,表明微生物群落的代谢加快,活动强度加大;土壤微生物群落对糖类、胺类、氨基酸类和羧酸类4类碳源的优势利用顺序发生明显变化,表明土壤微生物群落结构及其功能多样性发生了一定改变;土壤微生物群落对聚合物类和其他类碳源的利用率无显著影响,但显著提高对糖类、胺类和氨基酸类碳源的利用率,并仅在第10天显著降低对羧酸类碳源的利用率,表明降解糖类、胺类和氨基酸类碳源的微生物可能是转Bt基因棉粉碎叶影响的主要土壤微生物类群。     

Bt蛋白在不同矿物上的吸附动力学及其影响因素研究

<正>苏云金芽胞杆菌(Bacillus thuringiensis,简称Bt)近缘于蜡状芽胞杆菌,革兰氏染色阳性,是一种产生伴孢晶体,能寄生于昆虫体内引起昆虫发病的芽胞杆菌。自Vaeck等[1]首次报道转Bt基因抗虫植物以来,从苏云金芽孢杆菌中提取的Bt毒素蛋白被广泛运用于育种实践,这些作物在大田种植过程     

转Bt基因棉种植对根际土壤生物学特性和养分含量的影响

通过盆栽试验,比较了转Bt基因棉Bt新彩1和Bt基因的受体棉新彩1根际土壤可培养微生物种群数量、酶活性及养分含量的变化。结果表明,Bt新彩1根际土壤可检测到Bt蛋白,且花期达到峰值56.14 ng/g;与对照新彩1相比,Bt#61472;新彩1根际土壤更有利于细菌和真菌的生长和繁殖,放线菌数量没有显著变化。Bt新彩1的根际土壤碱性磷酸酶活性受到抑制,在生长旺盛期脱氢酶活性受到激活,而根际土壤蛋白酶、脲酶和蔗糖酶活性无显著变化;苗期、花期Bt新彩1根际土壤有机质、全氮、速效氮和钾含量没有显著变化,且苗期速效磷含量也没有显著改变,花期其含量显著降低。     

Environmental fate and effects of Bacillus thuringiensis (Bt) proteins from transgenic crops: a review

This paper reviews the scientific literature addressing the environmental fate and nontarget effects of the Cry protein toxins from Bacillus thuringiensis (Bt), specifically resulting from their expression in transgenic crops. Published literature on analytical methodologies for the detection and quantification of the Cry proteins in environmental matrices is also reviewed, with discussion of the adequacy of the techniques for determining the persistence and mobility of the Bt proteins. In general, assessment of the nontarget effects of Bt protein toxins indicates that there is a low level of hazard to most groups of nontarget organisms, although some investigations are of limited ecological relevance. Some published reports on the persistence of the proteins in soil show short half-lives, whereas others show low-level residues lasting for many months. Improvements in analytical methods will allow a more complete understanding of the fate and significance of Bt proteins in the environment.     

离子强度和三氧化物对土壤吸附Bt毒素的影响

Chemical reactions and fate of the toxins of Bacillus thuringiensis(Bt) in the soil environment are causing increasing concerns due to the large-scale cultivation of transgenic Bt plants.In this study,the effect of ionic strength(0-1 000 mmol kg 1) adjusted by NaCl or CaCl 2 on adsorption of Bt toxin by a lateritic red soil,a paddy soil and these soils after chemical removal of organic-bound or free Fe and Al oxides,as well as by pure minerals(goethite,hematite and gibbsite) which are widespread in these soils,were studied.The results indicated that when the supporting electrolyte was NaCl,the adsorption of Bt toxin by the lateritic red soil and paddy soil increased rapidly until the ionic strength reached 250 mmol kg 1 and then gradually slowed down with the increase of ionic strength;while in case the supporting electrolyte was CaCl 2,the adsorption of Bt toxin enhanced significantly at low ionic strength(< 10 mmol kg 1) and then decreased as the ionic strength increased.The adsorption of Bt toxin by the tested minerals and soils after the removal of organic-bound or free Fe and Al oxides also increased with increasing ionic strength controlled by NaCl.Removing organic-bound Fe and Al oxides obviously increased the adsorption of Bt toxin in the tested soils.Differently,removing free Fe and Al oxides increased the Bt adsorption by the paddy soil,but decreased the adsorption by the lateritic red soil.The study indicated that the varieties of ionic strength and the presence of Fe and Al oxides affected the adsorption of Bt toxin by the soils,which would contribute to the further understanding of the fate of Bt toxin in the soil environment and provide references for the ecological risk assessment of transgenic Bt plants.     

Transgenic cotton could safely be grown since CpTI toxin rapidly degrades in the rhizosphere soil

Abstract

Cultivation of transgenic plants is debated worldwide. Potential environmental risks have to be considered, before acceptance of expanding cultivation, despite the advantages of the use of fewer pesticides. Here, the potential effects on soil ecosystems of transgenic plants have been studied. As a model, genetically engineered cotton producing cowpea trypsin inhibitor (CpTI) has been used. The degradation of CpTI in the rhizosphere of the transgenic CpTI+Bt (Bacillus thuringiensis) cotton cultivar SGK321 was assessed. During plant development, concentrations of CpTI toxin in the rhizosphere were measured using an enzyme-linked immunosorbent assay (ELISA). As the plants developed, the residue of CpTI in the rhizosphere increased, and reached a peak at topping stage (100 days after planting). After this stage, the residue began to decrease, and was nil the following year (258 days after planting). The conclusion is that genetically engineered cotton can safely be cultivated since no accumulation of substances released from the transgenic plants was persistent in the soil.     

Call for Papers--Bt Research （ISSN 1925-1939）

Bt Research （ISSN 1925-1939） is a new launched, open access and peer-reviewed journal that disseminates significant creative reviews and opinions or innovative research work in the area of Bacillus thuringiensis, including the isolation and identification of novel Bt strains, identification of novel Bt toxic genes and their functions, the insecticidal mechanism Bt toxics, Bt genetic engineering, transgenic Bt plants, the resistance mechanism of target-insect to Bt toxins, and the development of novel experimental methods and techniques for Bt Research.     

导入外源Bt基因与受体作物次生代谢物质的交互关系研究

转Bt基因作物是目前商品化进程最快的转基因作物之一,外源Bt基因的导入为培育有效控制有害生物的作物品种提供了新的手段,但外源基因的导入一定程度上改变了作物自身的基因序列,可能引起作物的代谢过程发生改变。本文对转Bt基因作物中导入的Bt基因所产生的杀虫蛋白与作物自身次生代谢物质含量及其效应的关系进行简要综述,结果表明基因的改造和重组已经影响到转Bt基因作物的一些代谢过程。     

Effect of Transgenic Bt Cotton on Bioactivities and Nutrients in Rhizosphere Soil

The influence of transgenic Bacillus thuringiensis (Bt) cotton (BtXincai1) and its corresponding nontransgenic isoline (Xincai1) on the microorganisms, enzyme activity, and nutrient content of rhizosphere soil was studied through experiments in potted plants. The calcareous drab soil samples were collected (0–15 cm deep) from an experimental field in Shanxi Agricultural University (China) in 2005. The pots were categorized in different groups with replicates for each variety (transgenic BtXincai1 and general Xincai1). The rhizosphere soil samples were collected at different growth periods (seedling, bud, flowering, peak boll, boll opening, and harvest). The Bt protein and other microbial properties in the soil samples were determined by using selected methods (material and methods session). The results demonstrated that the concentration of the Bt protein in the rhizosphere soil of BtXincai1 reached a peak at 56.14 ng g^?1 during the flowering period. However, the Bt protein would not continuously accumulate in the soil. The rhizosphere soil of BtXincai1 was more suitable for the growth and proliferation of bacteria and fungi but it had no significant impact on the number of actinomycetes. BtXincai1 had some inhibitory effects on alkaline phosphatase activity in the rhizosphere soil, and it might promote dehydrogenase activity during the blooming period. However, it had no significant influence on protease, urease, or sucrase activities. Further, it had no significant impact on the contents of organic matter, total nitrogen, available nitrogen, or potassium in rhizosphere soil. It could significantly decrease the content of available phosphorus during the flowering period. Based on this study, the sensitive reactions of microorganisms and the activities of alkaline phosphatase and dehydrogenase might be considered as three potential indexes for assessing the risk posed by transgenic Bt cotton to soil ecology.     

Soil persistence of <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> (Bt) toxin from transgenic Bt cotton tissues and its effect on soil enzyme activities

A silty loam soil was incubated with the leaves and stems of two transgenic Bacillus thuringiensis (Bt) cotton varieties and nontransgenic Bt cotton to study the soil persistence of the Bt toxin from the decomposing transgenic Bt cotton tissues and its effect on soil enzyme activities. The results showed that after Bt cotton tissue amendment, Bt toxin was introduced into soil upon decomposition; about 50% of the introduced Bt toxin persisted in soil for at least 56 days. No Bt toxin was detected in the nontransgenic Bt cotton-amended soil; the amount of Bt toxin was the highest in the soil treated with the residue with the higher Bt toxin content. Activities of soil urease, acid phosphomonoesterase, invertase, and cellulase were stimulated by the addition of Bt cotton tissues, whereas activity of soil arylsulfatase was inhibited. Probably cotton tissue stimulated microbial activity in soil, and as a consequence, enzyme activities of soil were generally increased. This effect can mask any negative effect of the Bt toxin on microbial activity and thus on enzyme activities.