灰飞虱体内共生菌Wolbachia的groEL基因克隆及序列分析

利用特异性引物,首次在国内克隆获得灰飞虱(Laodelphax striatellusFalln)体内共生菌Wolbachia的groEL基因全长序列,其开放阅读框为1659bp,可编码552个氨基酸,GenBank登录号为:EF468716,与已发布的韩国分离物核苷酸一致性达99.9%,为进一步研究其与灰飞虱传播水稻条纹病毒的关系奠定了基础.     

灰飞虱Wolbachia群体生物学的遗传特性研究

Ｗｏｌｂａｃｈｉａ是存在于多种昆虫生殖器官的一类共生菌,它可能在宿 主群体中引起细菌质不亲和性而给宿 主带来生殖优势。对灰飞虱体内的Ｗｏｌｂａｃｈｉａｆｊｔｆｄｇｐｗ,ｖ ｇｍ ｐｇｃｆｍｗ ｑｎｎｋｕｄｙｊｒ ｄｏ ｎｕｉ     

植物介导的土耳其斯坦叶螨内共生菌Wolbachia和Cardinium水平传播的比较

为比较研究寄主植物介导的内共生菌Wolbachia和Cardinium的水平传播能力,解释在节肢动物中Wolbachia感染率高于Cardinium感染率的原因。我们运用PCR技术和荧光原位杂交检测Wolbachia和Cardinium的水平传播途径。结果发现Wolbachia在棉花叶片中存在的时间和表达量均高于Cardinium,大多数未感染的土耳其斯坦叶螨能够被2种共生菌感染,且Wolbachia感染率高于Cardinium。证明了Wolbachia的水平传播能力比Cardinium强,这可能是导致自然界中约60%～70%的节肢动物被Wolbachia感染,而只有约20%～30%的节肢动物被Cardinium感染的原因之一。     

Wolbachia 在中国稻田飞虱中的感染和传播

用ＰＣＲ方法检测了采集于不同地域的稻田飞虱共生菌Ｗｏｌｂａｃｈｉａ的感染,发现灰飞虱Ｌａｏｄｅｌｐｈａｘ ｓｔｒｉａｔｅｌｌｕｓ,褐飞虱Ｎｉｌｅｐａｒｖａｔａ ｌｕｇｅｎｓ,白背飞虱Ｓｏｇａｔｅｌｌａ ｆｕｒｃｉｆｅｒａ 为Ｗｏｌｂａｃｈｉａ所感染。克隆了编码Ｗｏｌｂａｃｈｉａ外膜蛋白质的ｗｓｐ基因并进行了序列测定,对ｗｓｐ的ＲＦＬＰ分析证实了这些飞虱为单一Ｗｏｌｂａｃｈｉａ感染,还发现一种能同时     

灰飞虱共生菌Wolbachia引起的细胞质不亲和性

对我国６个地区和日本３个地区灰飞虱的Ｗｏｌｂａｈｉａ感染率，用ＰＣＲ技术进行了检测，结果表明中国的辽宁、北京、上海和云南的灰飞虱的Ｗｏｌｂａｃｈｉａ感染率均接近１００％；四川为５９．６％；而宁夏为０。各地区灰飞虱间的交配实验证明了在Ｗｏｌｂａｃｈｉａ引起的细胞质不亲和现象的存在，Ｗｏｌｂａｃｈｉａ１６ＳｒＤＮＡ的部分测序分析表明，上海、云南和日本出云的灰飞虱发的同为Ｗｏｌｂａｃｈｉａ ｐｉｐｉｅｎ     

生物鉴定和喂养试验证明GNA赋予转基因水稻纯系对褐飞虱的抗性

参照Rao等（1998）的褐飞虱生物鉴定和喂养方法,用水稻褐飞虱生物Ⅰ型的－龄若虫食喂,用基因枪法获得2个转基因水稻纯系。这2个纯系均含有并表达潮霉素抗性基因（hpt),gusA报告基因和雪花莲凝集素基因（gna)。褐飞虱生物鉴定和喂养试验表明,水稻纯系对褐飞虱具有显著的抑制作用。具体表现为降低褐飞虱成活率和繁殖力、延缓褐虱发育以及减少褐飞虱进食是。通过褐飞虱生物鉴定和喂养试验证明,表达GNA的转基因水稻纯系对严重危害水稻生产的褐飞虱具有抗性作用。     

褐飞虱在不同水稻品系上繁殖能力及相关解毒酶系基因表达变化分析

采用水稻敏感品系TN1和抗性品系中浙优、IR56饲养褐飞虱,研究褐飞虱产卵量等繁殖情况,测定解毒酶系中酯酶、细胞色素P450和谷胱甘肽转移酶(GST)及卵黄原蛋白(Vg)等基因在mRNA上的表达变化.结果显示褐飞虱在抗性水稻上产卵量减少,Vg2的表达量极显著下降,表明水稻抗性对褐飞虱的产卵量有抑制作用且Vg2调控产卵.酯酶和P450中的部分基因均在取食抗性水稻的褐飞虱体内表达量较高,存在极显著差异.     

褐飞虱谷胱苷肽转移酶基因cDNA片段的克隆、测序及表达分析

谷胱苷肽转移酶是昆虫体内重要的解毒酶系之一，研究水稻害虫褐飞虱的谷胱苷肽转移酶基因在褐飞虱与水稻互作中的表达变化，可为有效防治褐飞虱提供新的理论依据。利用反转录多聚酶链式反应（RTPCR）技术克隆了褐飞虱谷胱苷肽转移酶基因编码区的eDNA片段，并使用Northern杂交技术检测了该基因对两种不同抗性水稻的分子反应。结果表明，所克隆到的eDNA片段长度为201bp，该片段所编码的氨基酸序列与来自大劣按蚊、细小按蚊、冈比亚按蚊、果蝇和木瓜果实蝇的谷胱苷肽转移酶的片段存在高度同源性。Northern杂交显示，在褐飞虱取食抗性水稻后，谷胱苷肽转移酶基因表达水平明显升高，但褐飞虱取食感虫水稻TN1后，该基因的表达水平没有明显变化。     

植食性昆虫、植物和共生微生物的功能关联

植食性昆虫和植物均能与微生物形成密切关系，它们各自的生态功能及相互关系也常被共生微生物所影响。近年来，随着分子水平研究方法的进展，植食性昆虫和植物中很多可遗传共生微生物（细菌、真菌等）被发现。共生微生物能够在营养、生殖、防御和解毒等方面给宿主带来显著影响，并与宿主形成竞争、互利或寄生等关系。植食性昆虫体内的含菌胞、肠道、血淋巴、唾液腺等常含有重要功能的共生微生物。新的分子生物学手段和高通量测序技术的应用使得我们能够增加对宿主和共生微生物（即使处于低丰度）之间关系的了解。本文尝试总结了植食性昆虫和植物共生微生物的多样性及其关系、昆虫和植物互作机制、昆虫共生菌解毒植物毒素等方面的近期证据，突出强调了应以系统观思维来理解共生微生物、植食性昆虫和植物间的功能关联，并就将来值得研究的问题提出了建议。     

河北省石家庄地区常见飞虱发生概况

近年来,由于小麦和玉米病毒病在河北省的蔓延流行,因而对于传毒媒介昆虫——灰飞虱的研究也逐渐被重视起来。虽然主要传毒昆虫是灰飞虱,但在石家庄地区还有一些其它种类飞虱与灰飞虱混生,因此在研究灰飞虱发生规律的过程中,必然要涉及与灰飞虱混生的其它种类飞虱。为此,今年我们对石家庄地区常见飞虱的发生概况进行了初步的调查。     

Somatic stem cell niche tropism in Wolbachia

Wolbachia are intracellular bacteria found in the reproductive tissue of all major groups of arthropods. They are transmitted vertically from the female hosts to their offspring, in a pattern analogous to mitochondria inheritance. But Wolbachia phylogeny does not parallel that of the host, indicating that horizontal infectious transmission must also occur. Insect parasitoids are considered the most likely vectors, but the mechanism for horizontal transfer is largely unknown. Here we show that newly introduced Wolbachia cross several tissues and infect the germline of the adult Drosophila melanogaster female. Through investigation of bacterial migration patterns during the course of infection, we found that Wolbachia reach the germline through the somatic stem cell niche in the D. melanogaster germarium. In addition, our data suggest that Wolbachia are highly abundant in the somatic stem cell niche of long-term infected hosts, implying that this location may also contribute to efficient vertical transmission. This is, to our knowledge, the first report of an intracellular parasite displaying tropism for a stem cell niche.     

A host parasite interaction rescues Drosophila oogenesis defects

The cytoplasmically inherited bacterium Wolbachia pipientis is a widespread parasite of arthropods that manipulates the reproductive biology of its hosts, often to their detriment, in order to foster its own transmission through egg cytoplasm. Here we report that infection by Wolbachia restores fertility to Drosophila melanogaster mutant females prevented from making eggs by protein-coding lesions in Sex-lethal (Sxl), the master regulator of sex determination. Suppression of sterility by Wolbachia discriminates markedly among similar germline-specific Sxl alleles, and is not observed for mutations in other genes that produce similar 'tumorous ovary' phenotypes, including one that blocks Sxl germline expression. This allele and gene specificity indicates that suppression probably results from a specific interaction with Sxl protein, rather than from a bypass of the normal germline requirement for this developmental regulator or from an effect on Sxl expression. The Sxl-Wolbachia interaction provides a rare opportunity to explore host-parasite relationships at the molecular level in a model insect. Furthermore, demonstration that a parasite infection can counteract the deleterious effects of mutations in host genes illustrates how hosts might become dependent on parasites.     

Wolbachia variability and host effects on crossing type in Culex mosquitoes

Wolbachia is a common maternally inherited bacterial symbiont able to induce crossing sterilities known as cytoplasmic incompatibility (CI) in insects. Wolbachia-modified sperm are unable to complete fertilization of uninfected ova, but a rescue function allows infected eggs to develop normally. By providing a reproductive advantage to infected females, Wolbachia can rapidly invade uninfected populations, and this could provide a mechanism for driving transgenes through pest populations. CI can also occur between Wolbachia-infected populations and is usually associated with the presence of different Wolbachia strains. In the Culex pipiens mosquito group (including the filariasis vector C. quinquefasciatus) a very unusual degree of complexity of Wolbachia-induced crossing-types has been reported, with partial or complete CI that can be unidirectional or bidirectional, yet no Wolbachia strain variation was found. Here we show variation between incompatible Culex strains in two Wolbachia ankyrin repeat-encoding genes associated with a prophage region, one of which is sex-specifically expressed in some strains, and also a direct effect of the host nuclear genome on CI rescue.     

Cloning and characterization of a gene encoding phage-related tail protein (PrTP) of endosymbiont Wolbachia

Wolbachia is an obligatory, maternally inherited intracellular bacterium, known to infect a wide range of arthropods. It has been implicated in causing cytoplasmic incompatibility (CI), parthenogenesis, the feminization of genetic males and male-killing in different hosts. However, the molecular mechanisms by which this fastidious bacterium causes these reproductive abnormalities have not yet been determined. In this study, we report on the cloning and characterization of the gene encoding phage-related tail protein (PrTP) from Wolbachia in Drosophila melanogaster CantonS (wMelCS) and from Wolbachia in Drosophila melanogaster yw67c23 (wMel) by representational difference analysis (RDA) and ligation-mediated PCR (LM-PCR). The functionality of a bipartite nuclear localization signal sequence (NLS) of the gene was also successfully tested in Drosophila S2 cells. PrTP expression in various strains of Wolbachia was investigated. Our results suggest that PrTP may not induce CI directly. However, the existence of prtp provided direct evidence of phage-mediated horizontal gene transfer (HGT) that might play an important role in a variety of reproductive abnormalities of Wolbachia.     

Isolation and characterization of a potential transposable element from Wolbachia

Wolbachia are a group of Rickettsia-like bacteria which parasitize the cells of a wide range of anthropoid. These microorganisms are associated with the reproductive and developmental abnormalities io their hosts. To study the molecular mechanism underlying such phenomena, we analyzed the genomic difference between Wolbachia with different cytoplasmic incompatibility (CI) phenotype using representational difference analysis method. A potential transposable element, which exists in the strong CI-inducing strain wRi, was isolated. This element was designated as Wolbachia insertion sequence element (WISE).     

Infectious parthenogenesis

Parthenogenesis-inducing Wolbachia bacteria are reproductive parasites that cause infected female wasps to produce daughters without mating. This manipulation of the host's reproduction enhances the transmission of Wolbachia to future generations because the bacteria are passed on vertically only from mothers to daughters. Males are dead ends for cytoplasmically inherited bacteria: they do not pass them on to their offspring. Vertical transmission of Wolbachia has been previously considered to be the main mode of transmission. Here we report frequent horizontal transmission from infected to uninfected wasp larvae sharing a common food source. The transferred Wolbachia are then vertically transmitted to the new host's offspring. This natural and unexpectedly frequent horizontal transfer of parthenogensis-inducing Wolbachia intraspecifically has important implications for the co-evolution of Wolbachia and their host.     

Molecular Identification of a Wolbachia endosymbiont in Trichogramma dendrolimi

Wolbachia is a common and widespread group of bacteria found in arthropods. These bacteria have evolved various mechanisms for manipulating reproduction of their host. The presence of Wolbachia in a lab strain of the arrhenotokous species Trichogramma dendrolimi was observed by the amplification and sequencing of part of the wsp gene. Aligning the resulting sequences with already published ones, the phylogenetic relationships between Wolbachia found in Trichogramma dendrolimi and in other Trichogramma wasps was established, and the phylogenetic relationships of Wolbachia in Trichogramma were not congruent with their hosts Trichogramma. Some factors contributing to this uncongruence are discussed here.