多重PCR检测病死鸡中沙门氏菌方法的研究

为了建立能够同时检测病死鸡样品中的沙门氏菌、鸡白痢沙门氏菌和肠炎沙门氏菌的多重PCR 方法.采用煮沸法提取 DNA 做为模板,选择分别针对沙门氏菌属、鸡白痢沙门氏菌和肠炎沙门氏菌的特异性基因(invA、fliC、sdfⅠ)设计引物,优化反应体系和反应参数,建立多重PCR检测方法.应用该方法对国内17家鸡场369份病死鸡样品进行沙门氏菌的检测,检测结果与传统细菌培养法的结果进行比较.结果表明,优化过的多重 PCR 可同时对沙门氏菌种、属进行鉴定,灵敏度为4.6×102 CFU/mL;多重 PCR 方法共检测出沙门氏菌34株,其中鸡白痢沙门氏菌21株、肠炎沙门氏菌5株,与传统细菌培养法检测结果的符合率分别为91.2%、90.5%和80.0%.     

鸡白痢沙门氏菌O12特异性噬菌体的分离及生物学特性研究

采用双层平板培养法从成都某养鸡场的污水样本中分离得到一株鸡白痢沙门氏菌O12 (宿主菌)的噬菌体(Ph-1), 并对菌斑形态及理化特性进行了初步研究. 试验结果显示, 纯化后该噬菌体的噬菌斑直径约3mm; 经高于60℃温度处理1h 后噬菌体失活; PH5-7时该噬菌体可保持较高效价和较强的紫外线耐受性; 该噬菌体的最佳感染复数(MOI)为 0.01-0.001; 一步生长曲线显示该噬菌体的潜伏期为40min, 裂解期持续60min.     

鸡白痢沙门氏菌O12特异性噬菌体的分离及生物学特性研究

采用双层平板培养法从成都某养鸡场的污水样本中分离得到一株鸡白痢沙门氏菌O₁₂(宿主菌)的噬菌体(Ph-1), 并对菌斑形态及理化特性进行了初步研究. 试验结果显示, 纯化后该噬菌体的噬菌斑直径约3mm; 经高于60℃温度处理1h 后噬菌体失活; PH5-7时该噬菌体可保持较高效价和较强的紫外线耐受性; 该噬菌体的最佳感染复数(MOI)为 0.01-0.001; 一步生长曲线显示该噬菌体的潜伏期为40min, 裂解期持续60min.     

SpiC/SsaM蛋白质复合物的原核表达及纯化

沙门氏菌毒力岛2中spiC所编码的效应蛋白在沙门氏菌得以于宿主吞噬性细胞内的存活中起到重要作用．SpiC与位于相同操纵元的SsaM特异结合，暗示了SpiC／SsaM蛋白质复合物参与转运元的形成并调控其他效应蛋白的分泌．为了从蛋白质结构角度研究SpiC／SsaM复合物的生物学功能，原核体系中共表达了SpiC和SsaM．与SpiC形成稳定的复合物，改善了SsaM单独表达不可溶的情况．胰蛋白酶限制性酶切实验显示：相对单独存在于溶液中的SpiC，形成复合物的SpiC更加稳定．得到大量高纯度的胰蛋白酶处理后的SpiC／SsaM复合物，并进行了结晶试验，为后续结构学研究奠定了基础．     

快速检验食品中沙门氏菌和O157:H7大肠杆菌

利用沙门氏菌快速检验装置(Oxoid公司出口）和可视免疫沉淀物（VIP）试剂盒（BioControlSystems公司出口）对广西桂林市的七星区某农贸市场的20种食品进行了沙门氏菌和O157：H7大肠杆菌的快速抽样检验，结果发现，其中的11种食品的沙门氏菌呈阳性反应，4种食品的O157：H7大肠杆菌也呈阳性反应，与目前国内普遍使用的传统检验方法相比，该方法具有快速、简便、高效、准确的特点。     

鼠伤寒沙门氏菌集体食物中毒临床报告

目的：对一起集体食物中毒进行检验分析报告。方法：对55例食物中毒者的食用食物、患者及厨师的分泌物进行病原学分析。通过细菌培养,血清学检查鉴定细菌类型。结果：中毒者的食用食物,分泌物均培养出鼠伤寒沙门氏菌。结论：食物污染经消化道传播仍是沙门氏菌属感染的主要途径,注意食品卫生是防止食物中毒的重要环节。     

Specific anti-tumor effect induced by attenuated Salmonella typhimurium vaccine expressing extracellular region of vascular endothelial growth factor receptor 2

The purposes of this research were to study the stable expression of exogenous gene encoding therapeutic protein in attenuated Salmonella typhimurium, observe the metabolism of oral gene vaccine carried by attenuated Salmonella typhimurium in BALB/c mouse, and investigate the feasibility of prevention and treatment of tumors by the recombinant bacteria. Recombinant plasmid pcDNA3.1 VEGFR2(n1-7) was transformed into competent attenuated Salmonella typhimurium SL3261 to develop oral DNA vaccine SL3261-pcDNA3.1 VEGFR2(n1-7). To observe whether the exogenous gene can be expressed in the recombinant bacteria, PCR was performed to amplify the CMV promoter of the eukaryotic expression vector as the proof of stable expression of exogenous protein; transmission elec- tron microscopy (TEM) was applied to observe the morphology of the recombinant bacteria to confirm that the exogenous gene has no impact on the growth of the bacteria, and then BALB/c mice were immunized with the gene vaccine. After inoculation of the gene vaccine, the recombinant bacteria SL3261 could be detected in the tissues such as small intestine, colon, liver and spleen. And then, mice in each group were challenged with tumor cells. The results of animal experiment showed that tumor growth of the mice in experimental group was inhibited and survival time of immunized mice was prolonged compared with control groups. A higher lymphocyte infiltration in tumors from animals treated with DNA vaccine was observed. Immunohistochemical analysis of tumor samples revealed an en- hanced accumulation of CD8 cytotoxic T lymphocytes, as well as an increase in CD4 cells in the tumors of animals treated with the oral gene vaccine compared to tumors from control group mice. Ultrastructure of the tumor tissue showed that tumor cells in the samples of the immunized mice were well-differentiated. Our research confirmed that the exogenous gene can be stably expressed in the attenuated Salmonella typhimurium and has no impact on the growth of the recombinant bacteria; the exogenous gene can de delivered to the host by attenuated Salmonella typhimurium to produce anti-tumor effect with no obvious cytotoxity to the host. In this study, it is established that attenuated Salmonella typhimurium could be used as a vector for oral gene vaccine, and our study provided a theoretical basis for the body distribution and the metabolism of the recombinant bacteria. This strategy may provide a simple, safe and effective way for the prevention and treatment of tumors.     

活菌制剂对雏鸡白痢沙门氏菌的体外拮抗及预防效果试验

从健康鸡的新鲜粪便中分离到乳酸杆菌和粪链球菌,佐以地衣芽孢杆菌,按不同比例制成活菌制剂。比较其对雏鸡白痢沙门氏菌的体外拮抗作用。结果表明,复合菌的组合适宜,对雏鸡白痢沙门氏菌的生长有很强的抑制作用。用此复合菌培养物预防人工感染的雏鸡白痢病,其预防保护率为８５％。     

In vitro mutagenicity testing of a potent,new, benzoyl urea insect growth regulator

Summary Bacterial mutagenicity assay to detect potential chronic toxicity of a potent, new, benzoyl urea insect growth regulator (CGA-112913 or IKI-7899, formerly UC-62644) was conducted using 5 histidine auxotrophs ofSalmonella typhimurium. Tests within the concentration range of 0.9–500 g (saturating)/plate of the compound with and without the 5–9 mammalian metabolic activation system showed no mutagenic effects clearing the way for long-term chronic toxicology studies.The authors thank Professor Bruce N. Ames for supplying the tester strains together with all the relevant information for conducting the bioassays and Ms. Norma Charlebois for her meticulous technical assistance.     

以减毒猪霍乱沙门菌为载体的抗O型口蹄疫病毒重组活菌苗在家兔体内免疫应答的初步研究

将含有口蹄疫病毒疫苗基因的真核表达质粒pBO1经过中间宿主鼠伤寒沙门菌LB5010修饰后,电转化到减毒的猪霍乱沙门菌C500中,构建成抗O型口蹄疫病毒的重组活菌苗pBO1／S．cho．血清凝集实验验证,筛选到的转化菌仍然具有猪霍乱沙门菌的血清学特性,即具有免疫原性．采用口服方式免疫家兔,检测重组菌在家兔体内诱导的免疫应答．家兔免疫后8周,分离脾脏T细胞,检测T细胞增殖情况．结果显示,口服DBO1／S．fho的家兔T细胞在FMDV特异性抗原的刺激下有明显的增殖反应,刺激指数SI〉11;免疫后2周和8周分别取家兔静脉血,用液相阻断ELISA法检测血清中针对FMDV的特异性抗体,pBO1／S．cho组家兔产生的抗体效价为1／32。