Cuterebra buccata: immune response in myiasis of domestic rabbits

Naturally infected rabbits (Oryctolagus) were used to define further the nature of the immune response in myiasis due to Cuterebra buccata. Third instar larvae were dissected into four fractions; (1) alimentary tract with attached organs, (2) hemolymph, (3) fat body with tracheae, and (4) cuticle with attached muscles. The antigens provoking immune phenomena in naturally infected rabbits were found to reside in the alimentary tract and hemolymph fractions only. All rabbits which were skin tested were found to exhibit delayed hypersensitivity and to have serum precipitins with specificity against these antigens. Passive cutaneous anaphylaxis activity was demonstrated only in sera from rabbits also exhibiting reaginic and/or Arthus-type skin hypersensitivities. With the use of immunoelectrophoresis four separate antigens were demonstrated in alimentary tract fractions. Larval dissections revealed the alimentary tract to be filled with cellular elements of rabbit whole blood. The immunologic findings are discussed in relation to this newly recognized feeding pattern and it is proposed that sensitization of the host occurs as a consequence of exogenous larval secretions injected at the time of feeding.     

Antisera-mediated in vivo reduction of Cry1Ac toxicity in Helicoverpa armigera

A functional assessment of Bacillus thuringiensis (Bt) toxin receptors in the midgut of lepidopteran insects will facilitate understanding of the toxin mode of action and provide effective strategies to counter the development of resistance. In this study, we produced anti-aminopeptidase (APN) and anti-cadherin sera with purified Cry1Ac toxin-binding APN or cadherin fragments from Heliocoverpa armigera. Antisera were evaluated for their effects on Cry1Ac toxicity through bioassays. Our results indicated that both the anti-APN and anti-cadherin sera reduced Cry1Ac toxicity in vivo, although cadherin antiserum reduced toxicity more than APN antiserum. These results suggest that both APN and cadherin are involved in Cry1Ac intoxication of H. armigera, evidence that the pore formation model may be representative of Cry1Ac toxin mode of action in this insect.     

小麦TaMlo基因的原核表达、抗体制备及细胞化学分析（简报）

白粉病菌（Blumeria graminis）是一类高度专化性的寄生真菌。可侵染650多种单子叶植物和9000多种双子叶植物．能够引起多种麦类作物的白粉病。给农业生产带来巨大的损失。由于白粉病菌生理小种多、变异快。所以利用专化性抗病基因难以解决植物的持久抗病性问题。人们在研究大麦白粉病时。发现大麦Mlo基因的隐性突变可导致大麦对绝大多数白粉病菌生理小种的高效持久的广谱抗病性。Schulze—Lefert等多家实验室合作于1997年成功克隆了野生的Mlo基因。进一步研究表明。该基因编码一种植物特有的具有7个跨膜区和羧基端长尾的膜蛋白（Mlo）,它可能对植物细胞的坏死起负调控作用。但Mlo基因如何表达及其在白粉病菌发育中的作用机制尚不清楚。     

棉铃虫核多角体病毒泛素基因的克隆表达及抗体制备

昆虫杆状病毒和痘病毒是目前已知唯一编码泛素基因的病毒。通过PCR方法,克隆了棉铃虫核多角体病毒(HaSNPV)泛素基因(Ubiquitin,Ubi)。序列分析表明,该基因编码区全长252bp,编码83个氨基酸残基,预计分子量为9.24kDa。将泛素基因克隆到原核表达载体pET28a上,构建重组质粒pETUbi,转化至大肠杆菌BL21(DE3)感受态细胞中,IPTG诱导表达融合蛋白。用Histag抗体检测目的蛋白,Westenblot实验证明所表达的蛋白是带有Histag的重组融合蛋白。通过改变IPTG浓度和诱导时间对表达条件进行了优化。利用NI琼脂糖凝胶亲和层析柱纯化目的蛋白,SDSPAGE鉴定为单一条带,同时用提纯蛋白制备了特异性抗体,为进一步的研究打下基础。     

紫藤脉花叶病毒cp基因在大肠杆菌中的表达及抗血清的制备

采用RT-PCR方法自紫藤脉花叶病毒北京分离物(WVMV-BJ)的基因组中分离出其CP基因,连接到原核表达载体pET22b( )上。获得的重组子pET-WVMVCP转化大肠杆菌BL21(DE3)后,用IPTG进行诱导表达。SDS-PAGE和Western blot分析表明,cp基因在大肠杆菌中获得了高效表达,融合蛋白分子量约为34．4kDa。将融合蛋白纯化后免疫兔子,获得了特异性较高的抗血清。微量免疫沉淀法测定该抗血清的效价为1／1024,酶联法(enzyme-linked immunosorbant assay,ELISA)测定的效价为1／8192。     

Regulation of DNA synthesis by guanosine-5′-diphosphate, cyclic guanosine-3′,5′-monophosphate, and cyclic adenosine-3′,5′-monophosphate in mouse lymphoid cells

The effect of various adenine and guanine nucleotides and nucleosides on DNA synthesis was studied in various types of mouse lymphoid cells. Two out of the ten compounds tested, namely guanosine-5′-diphosphate (GDP) and cyclic guanosine-3′,5′-monophosphate (cGMP) increased the thymidine incorporation into the DNA of the spleen cells and counteracted completely or partially the inhibitory action of cyclic adenosine-3′,5′-monophosphate (cAMP) on spleen cells stimulated by various B or T cell mitogens. GDP seems to act preferentially on thymus cells while cGMP acts better on bone marrow cells. The possible significance of the results for the mechanism of the mitogenic signal is discussed.     

Echinococcus granulosus: evaluation of purified antigens immunoreactivity

A new method is presented for the isolation of purified Echinococcus granulosus antigens from sheep hydatid fluid. Echinococcus granulosus antigens were separated from the host's serum contaminants by absorbing sheep serum proteins with specific immunoabsorbents.No net sensitivity gain was obtained by using these purified antigens rather than crude sheep hydatid fluid in hemagglutination and immunoelectrophoretic tests.The presence of two major antigens (≥ 400,000 and 150,000 MW) was confirmed, the larger component being clearly the most immunoreactive.Evidence of a third slightly antigenic fraction of low molecular weight is presented.The effectiveness of labeled major Echinococcus granulosus antigens in radioimmunoprecipitation test is reported.     

水稻瘤矮病毒P8蛋白的结构分析及其表达

为揭示水稻瘤矮病毒(RGDV)外层衣壳蛋白P8的结构特点及其在大肠杆菌中的表达特性。利用生物学软件和在线分析工具,对RGDV P8蛋白结构特征进行了生物信息学分析;同时将通过RT-PCR扩增的P8基因克隆到pGEX-4T-1上,构建pGP8转化大肠杆菌E.coli BL21(DE3),在IPTG诱导下表达。利用表达的融合蛋白免疫家兔,制备了抗血清,并以RGDV、水稻矮缩病毒(RDV)、水稻条纹病毒（RSV）、水稻锯齿叶矮缩病毒(RRSV)4种病毒作为供试材料进行特异性检测。分析显示P8蛋白在C端位置有一个典型的跨膜螺旋区,整个蛋白也具有Phytoreo P8家族共有的典型结构域Phytoreo P8 domain。SDSPAGE和蛋白印记分析表明,RGDV P8融合蛋白分子量约为73kDa,以包涵体的形式获得了高效表达。获得的效价高、特异性强的抗血清,为水稻瘤矮病毒的快速检测及P8蛋白的功能研究奠定了基础。     

单核增生李斯特菌Internalin A的克隆表达与抗体制备

目的克隆表达单核增生李斯特菌Internalin A（InlA）,并以之为抗原制备检测单核增生李斯特菌的抗体。方法通过PCR技术从单核细胞增生李斯特菌4b中扩增出inlA基因,克隆筛选和测序鉴定后,最终构建该基因的原核表达质粒pGEX-4T-InlA,谷胱甘肽树脂亲和层析纯化表达产物后,免疫小鼠分别制备相应的多抗和单抗。结果在大肠埃希菌中成功表达了InlA,并对其进行了纯化,融合表达产物分子量约为110 kD;免疫小鼠获得的抗血清效价达到1∶1600;得到了3株抗InlA的单克隆抗体杂交瘤细胞株,腹水单抗效价为1∶1×10^5-1∶3×10^5。2种抗体与其他病原菌均无交叉反应。结论通过表达单核增生李斯特菌的特异性蛋白制备的抗体,能有效地消除交叉反应,提高检测的特异性。     

Ichthyophthirius multifiliis Has Membrane-Associated Immobilization Antigens

Sera from fish that survive infections with the ciliated protozoon, Ichthyophthirius multifiliis , immobilize the parasite in vitro. In order to identify ceil surface antigens involved in the immobilization response, integral membrane proteins were extracted from tomites with Triton X-l14 and used to immunize rabbits. The rabbit antisera immobilized the parasite in vitro and antigens were localized to cell and ciliary plasma membranes by indirect immunofluorescent microscopy. The membrane protein fractions from both whole cells and tomite cilia were characterized by 1 - and 2-dimensiona! SDS-PAGE. A 43,000-dalton (D) glycoprotein with an isoelectric point of 7.0 is the predominant protein in these fractions, comprising 12% and 60% of the total protein of whole cell and ciliary membranes, respectively. Western blot analysis of ciliary proteins with immune rabbit sera indicated that the 43,000-D glycoprotein is the principal antigen.