非洲狮产气荚膜梭菌病的病原检测

北京动物园一头非洲狮突然死亡,死前曾有神经症状,死后立即解剖,主要病变为肺出血,十二指肠出血。从肺和肾组织分离到Ａ型产气荚膜梭菌,用该菌液０．０５ｍｌ腹腔注射小白鼠致死,用该菌的肉肝胃酶消化汤５ｈ培养的上清液０．１ｍｌ静脉注射小白鼠６ｈ死亡。产气膜梭菌除对草食动物有较强致病作用外,对大型猫科动物也有致病性。     

C型产气荚膜梭菌产α毒素能力分析及免疫原性试验研究

利用16S rRNA分子鉴定技术对病死羊体内分离的19株野毒株产气荚膜梭菌进行了分子生物学鉴定,从中筛选出产毒素稳定的一株C型产气荚膜梭菌(海C1株),并通过优化培养基配方及培养方式提高其产毒素能力,最终利用该菌株研制了羊猝疽灭活疫苗,完成了疫苗对家兔及羊的效力试验.结果显示,该菌株与现用羊瘁狙疫苗生产株分子系统发育树...     

仔猪大肠杆菌和C型产气荚膜梭菌的毒力研究

为了确定不同纤毛抗原的猪大肠杆菌和C型产气荚膜梭菌引起试验动物发病的最小菌(毒)含量,用不同菌数的E.coliC83549(K88)、C83644(K99)和C83710(987P)菌株培养物对出生18～24h的仔猪进行攻毒,用不同毒素含量的C型产气荚膜梭菌毒素分别对出生18～24h的仔猪和家兔进行攻毒。结果表明,引起仔猪发病的大肠杆菌三种菌株的最低含量均为150×108CFU,C型产气荚膜梭菌引起1日龄仔猪和家兔发病的最低毒素含量分别75和80小鼠MLD。     

D型产气荚膜梭菌引起青鹿猝死

对上海动物园 1头猝死的青鹿进行了详细的病原学研究。在死亡动物的肠组织中分离到粗大正直的杆菌 ,有荚膜和芽孢 ,并能使牛乳培养基海绵状发酵 ,可致死小白鼠 ,LD5 0 为 5× 1 0 4CFU。结合肠内容物的毒素中和试验、青鹿的临床症状和病理解剖特征 ,确诊该青鹿的猝死是由于感染了毒力较强的D型产气荚膜梭菌。     

产气荚膜梭菌型特异性多重PCR的研究

建立了一种简单且具有型特异性的产气荚膜梭菌的多重PCR诊断方法.结果表明,该PCR特异性好,只有产气荚膜梭菌呈现阳性,被检验的其他7种梭菌以及大肠杆菌、葡萄球菌和多杀性巴氏杆菌均为阴性.该PCR具有较高的敏感性,最小检测量为4.2×105 CFU/mL,单个菌落稀释10倍后也能被检测出,适用于医院、防疫和兽医诊断等部门,具有重要的公共卫生意义.     

A型产气荚膜梭菌α毒素的提纯试验

应用硫酸铵盐析、SephadexG 2 5和SephadexG 1 0 0层析的方法提纯A型产气荚膜梭菌α毒素 ,并利用卵磷脂水解试验监测α毒素的析出峰。结果表明 ,以 50 % (W/V)硫酸铵盐析A型产气荚膜梭菌α毒素 ,经SephadexG 2 5过柱后能很好地层析脱盐 ,并初步提纯 ,再经SephadexG 1 0 0层析后则能得到相对较纯的α毒素。该方法快速、简便     

Death of a neonatal lamb due to Clostridium perfringens type B in New Zealand

ABSTRACT

Case history and clinical findings: A flock of 20 sheep was kept within three paddocks on a single property. None of the animals in the flock had been vaccinated against any disease for at least three years. Abdominal bloating and haemorrhagic diarrhoea were observed in Lamb 1 at 24 hours-of-age. The lamb subsequently died within an hour of the onset of clinical signs. Lamb 2 was 3-days-old when observed to be recumbent with opisthotonus. The lamb was treated with dextrose, vitamins B1 and B12, and penicillin G, but died 4 hours later.

Pathological findings: Examination of Lamb 1 revealed markedly increased gas within the peritoneum and within dilated loops of intestine. The intestines were dark red and contained large quantities of haemorrhagic fluid. Histology of the intestines revealed peracute mucosal necrosis with minimal accompanying inflammation. The intestinal lumen contained cell debris, haemorrhage, and myriad large Gram-positive bacilli. The intestines of Lamb 2 did not appear bloated or reddened. However, multiple fibrin clots were visible within the pericardial sac. Histopathological examination revealed small foci of necrosis within the mucosa of the distal intestine. The necrotic foci were often associated with large numbers of large Gram-positive bacilli.

Immunohistochemsitry and molecular biology: Intestinal samples from Lamb 1 were processed for Clostridium perfringens immunohistochemistry, which revealed large numbers of intralesional, positively immunostained rods. Fragments corresponding to the expected sizes for genes encoding alpha, beta, and epsilon C. perfringens typing toxins were amplified by PCR from DNA extracted from formalin-fixed sections of intestine.

Diagnosis: Lamb dysentery due to C. perfringens type B.

Clinical relevance: C. perfringens bacteria have a worldwide distribution, but disease due to C. perfringens type B has only been diagnosed in a small number of countries and has never been reported in New Zealand or Australia. C. perfringens type B produce both beta toxin and epsilon toxins, therefore both haemorrhagic enteritis and systemic vascular damage can develop. As many animals are exposed to C. perfringens without developing disease, there must be additional unknown factors that resulted in disease in these particular sheep. Vaccines that specifically protect against C. perfringens type B are available and may be recommended for use in smaller non-commercial flocks, as in the present case.     

仔猪产气荚膜梭菌肠毒血症病原诊断及疫苗研究Ⅲ.7株A型产气荚膜梭菌菌株的产毒素试验

本试验对7株A型产气荚膜梭菌菌株在不同培养基、不同培养温度和不同培养时间的产毒力进行了试验。结果表明，培养基的种类和培养时间对菌株的产毒素能力影响较大，而培养温度则影响较小。所试菌株中C57-1菌株的产毒素能力最强，是制备疫苗的最佳菌株。     

CTB-CPA融合基因构建、表达及其产物的免疫原性

构建成含有编码霍乱毒素B亚单位 (CTB)和产气荚膜梭菌α毒素 (CPA)第 70～ 370位氨基酸的融合基因CTB CPA ,并在大肠杆菌中获得表达。经限制性内切酶鉴定和核苷酸序列分析表明 ,CPA基因在重组质粒中与CTB基因的连接向位是正确的 ,位于CTB基因的下游并与CTB基因处于同一阅读框架。重组菌株BL2 1(DE3) (pECTB CPA)经IPTG诱导后 ,其表达产物经SDS PAGE和Western blot检测表明 ,重组菌株可以表达CTB CPA融合蛋白。表达的产物以包涵体的形式存在 ,可分别被抗CT和A型产气荚膜梭菌抗毒素识别 ,表达量占菌体总蛋白的 2 2 7%。用表达产物免疫小鼠后 ,所产生的对A型产气荚膜梭菌毒素攻击的免疫保护力高于用单独的A型产气荚膜梭菌类毒素免疫的小鼠所产生的免疫力 ,证明所表达的融合蛋白中CTB起到免疫佐剂的作用 ,增强CPA的免疫原性     

Effects of supplementation with digestible undegradable protein in late pregnancy on ewe colostrums production and lamb output to weaning

The aim of this study was to investigate the effects of supplementation with digestible undegradable protein (DUP) in diet during late pregnancy on colostrum production, lamb survival and growth. For 3 weeks prior to lambing, 41 Afshari ewes were assigned at random to three dietary treatments, namely a control (CDUP), moderate (MDUP) and high (HDUP) DUP group. The ewes (n = 14) in CDUP group were fed a protein maintenance level for pregnant ewes [83 g metabolizable protein (MP)/kg dry matter (DM)], while ewes in the MDUP (n = 13) and HDUP (n = 14) groups were fed diets provided 14% (94.7 g MP/kg DM) and 24% (104.3 g MP/kg DM) MP in excess of these requirements respectively, which this increment was coming from DUP (DUP/MP ratio 43.8; 51.0% and 57.6% respectively). All diets were isoenergetic (12 MJ ME/kg DM). All ewes received the same lactation diet after lambing. Ewes in HDUP tend to produce more colostrums (p < 0.1) and had higher protein, fat, and solids non-fat content (p < 0.05) in colostrums during the first 24 h after lambing. MDUP group had higher colostrum yield compared to CDUP (p < 0.1). The changes in body weight and body condition score of ewes were similar for all diets but placenta weight was higher in MDUP group (p < 0.05) than for HDUP and CDUP ewes. Lamb body weight at weaning was higher for males compared to females (p < 0.05), but was not different between treatments. HDUP ewes had higher blood glucose concentration in the pre-lambing period than other treatments (p < 0.05). In this period, increasing the level of DUP in diet increased the content of blood urea nitrogen (p < 0.05). It was concluded that increasing the DUP content of diet for 3 weeks prior to lambing above the standard requirements resulted in an increase in colostrums production but had no effect on lamb outcome to weaning.     

A型产气荚膜梭菌灭活疫苗类毒素含量与疫苗攻毒保护率之间的平行关系研究

测定了制苗菌液中外毒素对小白鼠的最小致死量,并将该制苗菌液按《兽用生物制品规程》（以下简称《规程》）[1]方法制成A型产气荚膜梭菌灭活疫苗,按《规程》方法进行安全和效力检验。结果表明：6批疫苗安全检验全部合格;疫苗保护率与疫苗中类毒素含量呈正相关。     

体外培养中硒对黄曲霉毒素B_1损伤凡纳滨对虾肝胰腺细胞的影响

本试验旨在研究体外培养中硒(Se)对凡纳滨对虾(Litopenaeus vannamei)肝胰腺细胞抗氧化能力的影响以及对黄曲霉毒素B1(AFB1)毒性的缓解作用。在体外培养肝胰腺细胞的基础上,设定含不同浓度(0、0.15、0.30、0.45、0.60、0.75μg/m L)Se的培养液,研究Se对肝胰腺细胞抗氧化能力的影响;另用浓度为1 600μg/L的AFB1侵染细胞,探究Se对AFB1毒性的缓解作用。结果表明:当Se浓度处于0~0.45μg/m L时,随着Se浓度的增加,丙二醛(MDA)含量呈现降低趋势,但各个浓度之间并没有显著差异(P0.05),过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GSH-Px)活力随着Se浓度的增加而增强,Se浓度为0.45μg/m L时GSH-Px活力显著高于其他浓度(除0.30μg/m L)时(P0.05);当Se浓度处于0.45~0.75μg/m L时,随着MDA含量升高的同时CAT、GSH-Px活力下降。当细胞受到浓度为1 600μg/L的AFB1侵染后,随着培养时间的增加,MDA含量表现一直升高的趋势,同时CAT、GSH-Px活力都有一定程度的下降;当细胞受到浓度为1 600μg/L的AFB1和浓度为0.45μg/m L的Se共同作用后,随着培养时间的增加,MDA含量有一定程度的改善,CAT、GSH-Px活力得到一定程度的恢复。由此可见,在体外培养条件下,浓度为0.45μg/m L的Se有利于提高凡纳滨对虾肝胰腺细胞的抗氧化能力,且在一定程度上能缓解AFB1对肝胰腺细胞的毒性作用。     

硒对黄曲霉毒素B1中毒雏鸭肾组织抗氧化功能的影响

为观察黄曲霉毒素B1(aflatoxin B1,AFTBl)对试验雏鸭肾组织抗氧化功能的影响及亚硒酸钠对AFTBl的颉颃效应.本试验以雏鸭为试验动物,7日龄雏鸭90只,共分为3组,每组各30只.第Ⅰ组设为空白对照组,灌胃同等量二甲基哑砜;第Ⅱ、Ⅲ组为试验组.每天分别按0.1 mg/kg体重剂量给第Ⅱ、Ⅲ组雏鸭灌胃AFTB11次,连续投药21 d,试验期间给第Ⅲ组雏鸭每天按1 mg/kg体重剂量灌胃亚硒酸钠(Na2 SeO3)1次.分别在给雏鸭投药后7、14、21d,检测雏鸭肾组织总抗氧化能力(T-AOC)、超氧化物歧化酶(SOD)、过氧化氧酶(CAT)、谷胱甘肽过氧化物酶(GSH-Px)、谷胱甘肽还原酶(GR)及丙二醛( MDA)等抗氧化指标.试验结果显示,第Ⅱ、Ⅲ组雏鸭肾组织SOD、CAT、GSH-Px及GR活性与T-AOC均显著低于第Ⅰ组(P＜0.05),而MDA显著高于第Ⅰ组(P＜0.05);与第Ⅱ组相比,经灌胃投用亚硒酸钠的第Ⅲ组雏鸭肾组织各项抗氧化指标均得到明显的改善(P＜0.05).结果表明,黄曲霉毒素B1导致雏鸭肾组织抗氧化功能发生显著的变化,而亚硒酸钠能明显改善其变化.     

Displaying epitope B and epitope 7 of porcine reproductive and respiratory syndrome virus on virus like particles of porcine circovirus type 2 provides partial protection to pigs

The Cap of porcine circovirus type 2 (PCV2) can be assembled into virus like particles (VLPs) in vitro that have multiple loops located on the particle surface. This would make it a good vehicle for displaying exogenous proteins or epitopes. We derived two epitopes, epitope B (EpB, S³⁷HIQLIYNL⁴⁵) and epitope 7 (Ep7, Q¹⁹⁶WGRL²⁰⁰) from Gp5 of the highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV). We replaced the core region of Loop CD (L⁷⁵PPGGGSN⁸²) and the carboxyl terminus (K²²²DPPL²²⁶) of PCV2 Cap, respectively, to construct a bi-epitope chimeric PCV2 Cap. Its immunogenicity and protective effects were evaluated as one PRRSV subunit vaccine. The chimeric PCV2 Cap was soluble, efficiently expressed in an Escherichia coli expression system, and could be self-assembled into chimeric virus like particles (cVLPs) with a diameter of 12–15 nm. Western blotting confirmed that the cVLPs could be specifically recognized by anti-PCV2, anti-EpB and anti-Ep7 antibodies. The cVLPs vaccine could alleviate the clinical symptoms and reduce the viral loads after HP-PRRSV challenge in 100–120 days old pigs. These data suggest that the cVLPs vaccine could provide pigs with partial protection against homologous PRRSV strains, and it provides a new design for additional PRRSV subunit vaccines.