鸽禽流感免疫防制程序的探讨

对不同龄期鸽进行疫苗免疫和采用不同免疫间隔进行疫苗免疫后抗体消长动态及母源抗体的检测结果:在7～35日龄初免,7天后产生抗体,28～35天抗体滴度达到较高点,效果较好;首免后间隔20～90天二免,效果较优,当母鸽为高抗体水平(8.1log2)时产下的子代母源抗体至15日龄时已衰减至3.7log2。据此,提出了鸽场禽流感的免疫程序。     

鸡抗大肠杆菌血清抗体和卵黄抗体变化规律研究

用猪源大肠杆菌四价(K88、K99、987P、F41)灭活疫苗免疫蛋鸡,微量凝集法定期检测蛋鸡血清抗体和卵黄抗体的效价。结果表明,血清抗体比卵黄抗体出现早,免疫蛋鸡血清中和卵黄中4种抗体的消长规律基本相似,但不同抗体水平之间存在明显的差异,整个试验期间血清抗体水平均比卵黄抗体水平高。     

Comparative Pathogenesis of Experimental Infections with Salmonella gallinarum in Local and Commercial Chickens

The pathogenicity of a virulent strain of Salmonella gallinarum was studied in local chickens and commercial layers. Sixty 4-month-old chickens were used. Of these, 50 chickens were orally infected with S. gallinarum, comprising 25 commercial layers and 25 local chickens. Five chickens in each group were used as uninfected negative controls. The clinical signs and pathological features of acute, subacute and chronic fowl typhoid were observed in both groups. Chickens in both groups seroconverted, but the antibody titre was significantly higher (p<0.001) in the commercial layers. The antibody titre remained high to the end of the experiment in all the surviving chickens. Only one commercial layer chicken died during the course of the experiment. The PCV decreased significantly (p<0.001) in the infected chickens of both groups as compared to the controls. The viable cell count of S. gallinarum in the liver and spleen reached a maximum on day 9 after infection in both groups. However, there was a significantly higher cell count (p<0.05) in the commercial layers. The severity of the disease appeared to be slightly greater in the commercial layers, although the susceptibility was similar in both groups. It was concluded that, under experimental conditions, local chickens are just as susceptible to S. gallinarum infection as are commercial layers.     

A Single-chain Fragment Variable Recombinant Antibody against F5 Fimbria of Enterotoxigenic <Emphasis Type="Italic">Escherichia coli</Emphasis> Inhibits Agglutination of Horse Red Blood Cells Induced by F5 Protein

Bovine colibacillosis caused by enterotoxigenic Escherichia coli (ETEC) is a worldwide problem. Adhesion of ETEC to intestinal cell receptors mediated by the surface protein F5 fimbriae is the initial step in the establishment of colibacillosis. Prevention of ETEC F5⁺ adhesion to enterocytes protects newborn calves against collibacillosis. On the enterocytes, the F5 fimbriae bind to a ganglioside that is also found on horse red blood cells. Thus, the presence of F5 fimbriae induces haemagglutination, which is useful as an indicator in a functional assay system. In this study, recombinant anti-F5 scFv antibody fragment produced in E. coli HB2151 reacted with F5 fimbriae in ELISA and Western immunoblot, and prevented haemagglutination induced by the binding of the F5 fimbriae to its natural host receptors on horse red blood cells. Given the ease with which recombinant antibodies can be mass-produced, the presently described scFv may hold promise as a prophylactic agent for colibacillosis.     

Improved immunological detection of S<Emphasis Type="Italic">pongospora subterranea</Emphasis>

The genus Spongospora has two members which are important pathogens of vegetables, S. subterranea f.sp. subterranea (Sss) and S. subterranea f.sp. nasturtii (Ssn). The close taxonomic relationship of these formae speciales is based on similar cystosori morphology. The potato disease powdery scab, caused by Sss, is difficult to control. The key control measure is avoidance, aimed at planting clean seed in clean soil. For the development of routine tests for the presence of the pathogen on tubers and in soil, a monoclonal antibody (MAb) was developed using Sss cystosori as immunogen. It detected less than one Sss cystosorus and recognised Sss material from many parts of the world. No cross-reactions with other Plasmodiophoromycetes including Plasmodiophora brassicae, Polymyxa betae, Polymyxa graminis and different Streptomyces species causing common and netted scab of potatoes were observed. A novel tuber sample test method was developed using a kitchen peeling machine. This detected two tubers with one powdery scab lesion each in a sample including eighteen uninfected tubers. When soil samples spiked with cystosori were tested with the MAb, different Sss infestation levels could be discriminated. Ssn cystosori gave absorbance values in ELISA as high as Sss cystosori, whereas fresh crook roots of watercress containing Ssnzoosporangia and plasmodia or mud from an Ssninfected watercress bed gave low absorbance values or no reaction. The potential of these findings for the development of a disease control management are discussed.     

河南省部分地区奶牛肉孢子虫感染的血清学调查

为了解河南省奶牛肉孢子虫病流行情况,2018年3月至2019年9月用间接酶联免疫吸附试验(ELISA)对采自河南省13个地区58个奶牛场的1819份血样进行了肉孢子虫抗体检测分析。结果肉孢子虫抗体阳性牛场31个,场阳性率53.4%,阳性样品424份,样品平均阳性率为23.3%;不同生长阶段牛群阳性率差异极显著(P<0.01),产奶牛(24月龄以上)阳性率最高(27.9%),犊牛(6月龄以下)阳性率最低(13%);不同季节阳性率差异极显著(P<0.01),春季最高(35.1%),冬季最低(13.2%)。此调查结果为河南省奶牛肉孢子虫病防控提供了科学依据。     

番茄溃疡病菌单克隆抗体的制备与鉴定

为制备并鉴定番茄溃疡病菌(Clavibacter michiganensis subsp.michiganensis,Cmm)的单克隆抗体(McAbs),用全菌皮下免疫BALB/C小鼠,采用B细胞杂交瘤技术,经免疫、融合、间接ELISA筛选和克隆等,获得稳定分泌抗体的阳性杂交瘤细胞株,得到了抗番茄溃疡病菌的单克隆抗体。经免疫后获得3株单抗分别为1A4、1C3和1B7,经亚类鉴定分别是IgM、IgG1、IgG1;纯化腹水间接ELISA效价分别为1:3.2×106、1:8.1×105、1:3.2×106;与其他同属不同亚种无交叉反应。结果表明:3株单克隆抗体均具有较高特异性和敏感性,可作为番茄溃疡病菌的检测抗体,其中,1A4的效果最好。番茄溃疡病菌单克隆抗体的获得为进一步研发番茄溃疡病检测试剂盒奠定了基础。     

氯噻啉酶联免疫分析方法研究

建立了基于多克隆抗体的氯噻啉间接竞争酶联免疫分析(ic-ELISA)方法。设计合成了氯噻啉半抗原,将其分别与牛血清蛋白(BSA)和卵清蛋白(OVA)偶联制备免疫原和包被原。用免疫原免疫新西兰大白兔,制备多克隆抗体,抗体效价为2.56×106。通过对有机溶剂含量、离子强度和pH等影响因素的优化,确定了氯噻啉ic-ELISA的最佳检测条件(含体积分数10%甲醇的磷酸盐缓冲液、Na+浓度为0.14 mol/L、 pH 7.4),并建立了氯噻啉标准竞争曲线,该方法的抑制中浓度(IC50)为0.18 mg/L,最低检测限(IC10)为0.001 8 mg/L。所得多克隆抗体除与吡虫啉有较大的交叉反应外,与其他结构相似的化合物无明显交叉反应。在水、土壤和甘蓝中分别添加0.05~1 mg/kg的氯噻啉标准溶液,平均回收率为91.42% ~113.82%,相对标准偏差(RSD)为0.72% ~8.68%,符合农药残留检测要求。该ELISA方法可用于环境及农产品中氯噻啉残留检测。     

血清学检测的雄性特异性抗原Fab抗体的特异性鉴定

旨在研究血清学检测的雄性特异性抗原噬菌体Fab抗体的特异性。以从噬菌体Fab抗体库中筛选到具有较高雄性特异性结合活性的重组噬菌体克隆为基础,构建Fab抗体的可溶性表达噬质粒,并诱导和纯化该抗体片段,利用免疫荧光FITC/DAPI共染技术和ELISA分析其特异性。结果表明,该抗体在约49 ku处有条带出现。在Fab抗体和H-Y抗血清的细胞免疫荧光比较分析试验中发现,从雌、雄鼠脾细胞的阳性细胞数量和平均荧光强度的角度分析,Fab抗体的雄性特异性强于抗血清。石蜡切片免疫荧光定量分析显示,Fab抗体与雄鼠肝脏的结合活性明显高于对应雌鼠,差异极显著(t=20.73,P=0.002 3<0.01),而H-Y抗血清与雄鼠肝脏的结合活性略高于对应雌鼠,差异显著(t=7.11,P=0.019 2<0.05)。以C57BL/6雄鼠脾细胞、睾丸细胞作为抗原的ELISA分析显示,Fab抗体具有雄性特异性,但Fab抗体的OD值低于抗血清。结果提示,Fab抗体的雄性特异性高于H-Y抗血清,但其雄性特异性结合活性低于H-Y抗血清,Fab抗体在具备雄性特异性的同时,仍有少量雌性非特异性结合,Fab抗体的体外亲和力成熟可作为后续研究工作,以获...