抗虫转基因植物对非靶标节肢动物生态影响的研究进展

重组DNA技术的发展为培育高效的抗虫作物提供了前所未有的便利条件。通过转基因技术,全世界已培育出众多转基因抗虫植物品系。其中,表达苏云金芽孢杆菌(Bt)基因的作物品系如Bt棉花和Bt玉米已在很多国家大规模种植,在害虫控制方面发挥了重要的作用。转基因抗虫作物可能带来的生态风险问题,如对农田非靶标节肢动物的潜在影响,一直受到相关研究者及民众的广泛关注。至今,已有大量研究论文发表。本文在总结、归纳前人研究的基础上,阐述了从实验室到田间多层次评价转基因抗虫作物对非靶标生物影响的一般研究程序和方法,并简要综述了Bt玉米和Bt棉花2种已商业化种植的转基因抗虫作物对农田非靶标节肢动物生态影响的研究进展。现有研究表明:当前种植的Bt作物所表达的Cry蛋白杀虫专一性非常强,对农田非靶标节肢动物没有毒性;且Bt作物的利用降低了广谱化学杀虫剂的施用量,从而提高了非专一性害虫天敌的种群密度,加强了对害虫的控制,并有效地保护了生态环境和农民健康。因此,Bt作物可以作为害虫综合防治(IPM)的一个策略,结合其他防治措施可加强对害虫的有效控制。     

转基因植物对土壤污染相关研究

随着转苏云金芽孢杆菌基因（Bacillus thuringiensis, Bt）作物（后面全用转Bt基因作物代替）的推广应用，有关转Bt基因作物对土壤中微生物的生态风险尚无统一的结论。丛枝菌根(Arbusular Mycorrhiza, AM）真菌在土壤中分布广泛，是监测转Bt基因作物生态风险评估的重要微生物群落。Bt杀虫蛋白的表达及释放，可能会使AM真菌共生过程中一些重要阶段的生态共生效应受到影响。转Bt基因作物对AM真菌的影响可作为转Bt基因作物生态风险评价的重要指标。文章以紫云英为宿主植物，通过盆栽实验，人为添加提取的Bt蛋白，检测Bt蛋白对AM真菌侵染率、酶的活性以及生物量的影响，以探索Bt蛋白对AM真菌侵染性及共生效应的影响，从而为转Bt基因作物的生态风险评估提供参考。     

转Bt基因玉米的生态安全性研究进展

随着转基因作物的应用和推广 ,转 Bt基因作物释放后对生态环境及其它方面产生的潜在影响越来越受到重视。分别从生物活性杀虫晶体蛋白在土壤中的残留特性、杀虫晶体蛋白对土壤中非目标生物的影响、转 Bt基因玉米植株体成分的变化、转Bt基因玉米花粉中杀虫晶体蛋白的表达特性及其在田间和马力筋叶片上的散积状况、花粉中表达的杀虫晶体蛋白对君主斑蝶的毒性、君主斑蝶幼虫暴露在 Bt花粉中的概率及综合风险评价估算等方面对转 Bt基因玉米产生的杀虫晶体蛋白与土壤生态环境的相互作用、花粉对非目标生物影响的研究现状进行了综述。通过对转 Bt基因作物生态安全性的科学评价和广泛宣传 ,以确保生物技术的健康发展。     

转Bt抗虫稻对地上非靶标节肢动物的生态风险性

通过转入Bt基因使作物获得抗虫性,减少杀虫剂施用量,为害虫管理开辟了一条重要途径。但公众关注与担忧其环境释放可能存在生物安全与生态风险。本文在系统评述转基因抗虫水稻的生态风险、安全性评价研究内容与方法基础上,重点分析了转基因抗虫水稻对非靶标害虫、捕食性天敌、寄生蜂、中性昆虫及其在食物链上传递的生态风险与安全性,提出了未来研究发展的方向。     

转Bt基因水稻对二化螟绒茧蜂生物学特性的影响

用转Bt基因 (cry1Ab) 水稻KMD1饲喂3龄、4龄和5龄二化螟Chilo suppressalis一定时间后, 作为二化螟绒茧蜂Apanteles chilonis的寄主,研究了转基因水稻经寄主对绒茧蜂生物学特性的影响。结果发现：KMD1处理后,三个龄期幼虫的寄生率都显著下降,其中4龄和5龄达极显著水平;3龄和4龄上的结茧率显著低于对照;蜂蛹历期均短于对照,但仅3龄差异显著;从5龄幼虫所羽化的雄蜂寿命显著短于对照;蜂茧长显著短于对照;而对卵＋幼虫期、茧块茧数、蜂羽化率及性比均无显著影响。     

转Bt基因抗虫作物对非靶标害虫的影响

转基因作物的长期大面积种植,在为农业生产带来惠益的同时,对农业生态系统的健康和稳定可能会产生潜在的影响。转基因作物表达的Bt蛋白对靶标害虫起到较好的控制效果,而对Bt蛋白不敏感的非靶标害虫种群可能会迅速发展起来,对作物造成为害。随着抗虫转基因作物的连续多年种植,科学家们对于田间杀虫剂施用量的增减看法不尽一致。通过总结已有的研究报道,本文以Bt玉米和Bt棉花为例,分析了大田中非靶标害虫暴发的现状,以及暴发的主要原因(如杀虫剂的使用、害虫天敌减少和物种替代)。在生产实践中,抗虫作物的长期大面积释放导致广谱杀虫剂施用量减少,田间非靶标害虫数量上升。因此今后需要继续开展更多的研究来综合评估种植转Bt基因作物产生的长期潜在影响,优化害虫防治措施,避免非靶标害虫暴发。     

转Bt基因棉花生态风险评价的研究进展

转Bt基因抗虫棉(Gossypium spp.)是目前国内释放面积最大的转基因作物,其生态风险问题从一开始就受到密切的关注.从生态风险评价的角度,分转基因棉花中Bt杀虫蛋白的时空表达及其对害虫的控制效果、Bt基因通过花粉传播而扩散的风险、害虫对Bt棉花抗性的进化风险、Bt棉花对非目标生物体影响的风险等几个方面,综述了Bt棉安全性评价的最新研究进展,为生物安全管理提供咨询意见,并提出了目前针对Bt棉亟待研究的内容.期望本文能够为推动生物安全的研究和生物技术的发展做出一定的贡献.     

从作物驯化谈转基因

正1994年,美国Calgene公司研制的延熟保鲜转基因番茄在美国批准上市,成为世界上第一个批准商业化生产的转基因作物。之后,改良营养成分的转基因油菜、抗虫转Bt基因玉米、耐除草剂草甘膦转基因大豆等多种性状的转基因作物相继研发成功,并被批准大面积商业化种植。根据国际农业生物技术应用服务组织(ISAAA)发布的最新统计资料显示,截至2018年年底,     

转Bt基因水稻对土壤微生态系统的潜在影响

随着转基因作物商品化应用的增多,对其进行生态风险性评价尤为重要.国内外对转基因作物中外源基因向野生亲缘物种漂移的可能性、昆虫对抗虫转基因作物的耐受性以及转基因作物对生物多样性的潜在影响等问题进行了广泛的研究.文中从Bt杀虫结晶蛋白在土壤中的残留特性、Bt杀虫晶体蛋白对土壤微生物可培养类群和土壤酶活性的影响等方面对转Bt基因抗虫水稻的潜在生态风险性进行了简要综述,以期为同类研究提供有益的信息.     

转基因植株毒素可渗漏

纽约大学的研究人员,通过室内种植转基因Bt玉米植株试验表明：该玉米可免受钻心虫危害,其时效可达２５天之久。然而,在此种玉米根系的周围土壤中,可以提取Bt毒素物。他们认为此现象是使用转基因植株对土壤生态学冲击的例证。美国目前栽种有６００万公顷的Bt玉米,据推测在杀死危害玉米害虫的同时,对土壤中其它有机物的影响也是非常显著的。由于该毒素在土壤中存留时间较长,从而可使目标害虫增加抗药性。如果目标害虫增加了抗性,则表明以该种技术制成的转基因作物宣告失败。环境专家提示大家,现在是再次提出评价转基因玉米对环境风…     

棉田棉铃虫寄生峰对常规棉及转Bt棉品种的趋性反应

连续4年采用人工定期释放棉铃虫卵、幼虫和定期回收法,研究了不同棉花品种与棉铃虫卵期、幼虫期寄生蜂种群数量变化的关系．通过对卵、幼虫寄生率反正弦转换后进行方差分析表明,无论是在棉铃虫卵期还是在其幼虫期,转基因棉田中棉铃虫卵和幼虫的被寄生率始终显著低于其亲本对照棉．运用“Y”型嗅觉仪测定转基因棉及其亲本对照棉对棉铃虫幼虫寄生蜂——中红侧沟茧蜂的选择性行为反应,用成对数据进行方差分析,并进行了“T”测验．结果表明,转基因棉对棉铃虫寄生蜂有较强的忌避反应,且取食转基因棉的棉铃虫幼虫与被害转基因棉两者的共同组合与单独被害棉之间对寄生蜂的忌避效应基本一致．     

棉田棉铃虫寄生峰对常规棉及转Bt棉品种的趋性反应

连续4年采用人工定期释放棉铃虫卵、幼虫和定期回收法,研究了不同棉花品种与棉铃虫卵期、幼虫期寄生蜂种群数量变化的关系.通过对卵、幼虫寄生率反正弦转换后进行方差分析表明,无论是在棉铃虫卵期还是在其幼虫期,转基因棉田中棉铃虫卵和幼虫的被寄生率始终显著低于其亲本对照棉.运用“Y”型嗅觉仪测定转基因棉及其亲本对照棉对棉铃虫幼虫寄生蜂———中红侧沟茧蜂的选择性行为反应,用成对数据进行方差分析,并进行了“T”测验.结果表明,转基因棉对棉铃虫寄生蜂有较强的忌避反应,且取食转基因棉的棉铃虫幼虫与被害转基因棉两者的共同组合与单独被害棉之间对寄生蜂的忌避效应基本一致.     

Heritable transgenesis of Parastrongyloides trichosuri: a nematode parasite of mammals

Germline transformation of a parasitic nematode of mammals has proven to be an elusive goal. We report here the heritable germline transformation of Parastrongyloides trichosuri, a nematode parasite whose natural hosts are Australian possums of the genus Trichosurus. This parasite can undergo multiple free-living life cycles and these replicative cycles can be maintained indefinitely in the laboratory. Transformation was achieved by microinjection of DNA into the ovary syncytium of either free-living or parasitic adult females. By selecting for the transgenic progeny of successive free-living life cycles, it was possible to establish and maintain transgenic lines. All three transgenic lines tested were shown capable of establishing patent infections in possums and to transmit the functional transgene to their progeny. The transgene, driven by the Pt hsp-1 promoter, was constitutively expressed in intestinal cells at all stages of both parasitic and free-living life cycles, although gene silencing appears to occur in some transgenic progeny. This is the first report of heritable transgenesis in a parasitic nematode of a mammal and we discuss a variety of previously inaccessible experimental avenues that will now be possible with this powerful model system.     

5S rRNA sequences of 12 species of flatworms: implications for the phylogeny of the Platyhelminthes

5S rRNAs from 12 species of free living and parasitic platyhelminthes were sequenced. In the phylogenetic analysis, attention was focused on the statistical estimates of the trees corresponding to existing phylogenetic hypotheses. The available 5S rRNA data agree well with widely accepted views on the relationships between the Acoela, Polycladida, Tricladida, and Neorhabdocoela; our analysis of the published 18S rRNA sequences also demonstrated good correspondence between these views and molecular data. With available 5S rRNA data the hypothesis that the dalyellioid turbellarians is the sister group of the Neodermata is less convincing than the hypotheses proposing the Neodermata as the sister group of the Neorhabdocoela, or of the Seriata, or of the branch uniting them. A relatively low rate of base replacement in parasitic flatworms, probably, accounts for the uncertain position of the Neodermata, while a relatively high rate in planarians may explain a relatively too early divergence of the Tricladida in several published phylogenetic trees constructed from various rRNA data.     

Ripening in papaya fruit is altered by ACC oxidase cosuppression

Papaya (Carica papaya) is a very important crop in many tropical countries but it is highly susceptible to parasitic diseases, physiological disorders, mechanical damage and fruit overripening. Here we report a study on ACC oxidase cosuppression and its effects on papaya fruit ripening. Papaya ACC oxidase was isolated using PCR and embriogenic cells transformed by biolistic using the CaMV 35S promoter to drive the expression of the PCR fragment in sense orientation. Fifty transgenic lines were recovered and 20 of those were grown under field conditions. Southern analysis showed incorporation of the transgene in different copy numbers in the papaya genome. Fruits were evaluated in terms of texture (firmness), colour development, respiration and ethylene production. A sharp reduction in ethylene and CO2 production was detected, whereas softening and colour development of the peel were also altered. Overall, transgenic fruits showed a delay in ripening rate. A reduction in mRNA level for ACC oxidase in transgenic fruit was clearly detectable by northern blot. More studies are necessary before this technology can be used to extend the shelf life of papaya fruit.     

The effect of hFVIII transgene on the chromosomal aneuploidy rate in rabbits

The aim of this study was to compare the chromosomal aneuploidy rate between transgenic and non-transgenic rabbits derived from the F4 generation. Chromosomal analysis was carried out on bone marrow samples of New Zealand White transgenic (carrying human factor VIII gene) and non-transgenic rabbits (F4 generation) each having a different genetic background (female no. 1-3-5 line I and female no. 1-9-7 line II). C-metaphase plates were obtained from the bone marrow lymphocytes synchronized by the addition of 0.25 microg/ml colcemide. No significant difference in chromosomal aneuploidy between transgenic (61%) and non-transgenic (51.27%) rabbits of line I was observed. A higher but non-significant aneuploidy rate between transgenic and non-transgenic rabbits was found in line II, on the other hand a significant difference (P<0.05) was observed in diploidy rate. In conclusion, chromosomal aneuploidy rates in this experiment were higher than published previously in other reports.     

Parasitic nematodes, proteinases and transgenic plants.

Parasite proteinases have important functions in host-parasite interactions. Consequently, they have been investigated as targets for the control of both plant and animal parasites. Plant parasitic nematodes cause estimated annual losses to world agriculture of US$100 billion and, currently, their control often relies on highly toxic nematicides, with associated environmental risks. The potential of disrupting digestive proteinases for plant parasitic nematode control, via expression of proteinase inhibitors in transgenic plants, is summarized here by Catherine Lilley, Pauline Devlin, Peter Urwin and Howard Atkinson. They then consider whether the approach of expressing antinematode proteins in plants can be adapted for control of certain animal parasitic nematodes.     

Antibody expressing pea seeds as fodder for prevention of gastrointestinal parasitic infections in chickens

Background

Coccidiosis caused by protozoans of genus Eimeria is a chicken parasitic disease of great economical importance. Conventional disease control strategies depend on vaccination and prophylactic use of anticoccidial drugs. Alternative solution to prevent and treat coccidiosis could be provided by passive immunization using orally delivered neutralizing antibodies. We investigated the possibility to mitigate the parasitic infection by feeding poultry with antibody expressing transgenic crop seeds.

Results

Using the phage display antibody library, we generated a panel of anti-Eimeria scFv antibody fragments with high sporozoite-neutralizing activity. These antibodies were expressed either transiently in agrobacteria-infiltrated tobacco leaves or stably in seeds of transgenic pea plants. Comparison of the scFv antibodies purified either from tobacco leaves or from the pea seeds demonstrated no difference in their antigen-binding activity and molecular form compositions. Force-feeding experiments demonstrated that oral delivery of flour prepared from the transgenic pea seeds had higher parasite neutralizing activity in vivo than the purified antibody fragments isolated from tobacco. The pea seed content was found to protect antibodies against degradation by gastrointestinal proteases (>100-fold gain in stability). Ad libitum feeding of chickens demonstrated that the transgenic seeds were well consumed and not shunned. Furthermore, feeding poultry with shred prepared from the antibody expressing pea seeds led to significant mitigation of infection caused both by high and low challenge doses of Eimeria oocysts.

Conclusion

The results suggest that our strategy offers a general approach to control parasitic infections in production animals using cost-effective antibody expression in crop seeds affordable for the animal health market.     

Effect of pea lectin expressed transgenically in oilseed rape on pollen beetle life-history parameters

When investigating resistance to herbivorous insects in transgenic plants, it is primarily the damaging stage of the insect that is targeted. In our study, we showed that small effects of the transgenic plant on the pest performance might lead to a clear effect on the pest population level. Pollen beetle, Meligethes aeneus (Fabricius) (Coleoptera: Nitidulidae), larvae were reared on three transgenic lines of Brassica napus L. (Brassicaceae) expressing pea (Pisum sativum L.) lectin at 0.2, 0.6, and 0.7% of total soluble protein in anthers, and then monitored until the adult stage. Two lines without the gene construct and consequently containing no pea lectin were used as controls. The egg viability, larval weight, development time, and survival rate were recorded. In a separate experiment, the development time from egg to adult, adult weight, and winter survival of pollen beetles developed on intact plants were recorded. Survival of larvae to adult stage was significantly lower on the transgenic plant lines, independent of lectin level. Survival during hibernation was only 2.4% and was not affected by plant line. Combining previously published results from adult feeding and oviposition assays with the new data on larval development we summarize the effect of the transgenic oilseed rape on the pollen beetle, from oviposition in spring to overwintered adults of the new generation. Net reproductive rate was then shown to be reduced by half on the transgenic oilseed rape compared to the control plants.