青海省牛羊蓝舌病血清学调查

从青海省玉树、共和、格尔木、果洛、大通、祁连、德令哈、同德8个地区采集牛羊血清样品413份,应用琼脂免疫扩散法对蓝舌病进行了血清学调查。结果在被检的209份羊血清中,共检出阳性血清样品4份,平均阳性率为1.91%;而在被检的204份牛血清中则未检测到阳性样品。     

内蒙古地区奶牛病毒性腹泻/黏膜病血清流行病学调查

应用ELLSA试验对来自内蒙古地区17个大﹑中﹑小型奶牛场的2391份牛血清样品进行了牛病毒性腹泻/黏膜病抗体检测,并对其中222份抗体阴性牛应用ELISA试验进行牛病毒性腹泻/黏膜病的抗原检测。结果表明：17个奶牛场均检出BVDV抗体阳性,共检出阳性血清2125份,阳性率最高达100%,最低为46.8%,平均为88.9%。对14个奶牛场进行了BVDV抗原检测,在5个奶牛场检出BVDV抗原阳性,阳性率为3.6%（8/222）。表明内蒙古地区奶牛场普遍存在牛病毒性腹泻/黏膜病感染,感染率较高,并且牛群中存在持续性感染（PI）牛。     

祁连山北麓中部地区牦牛常见病的血清学调查

[目的]为了掌握祁连山北麓中部地区牦牛常见病的感染情况。[方法]对祁连山北麓中部地区的牦牛常见病开展了血清学检测。共检测280份血清样品,分别检测了布病、口蹄疫、牛传染性鼻气管炎、弓形虫、牛副结核病的感染抗体。[结果]检出布病阳性样品3份,平均阳性率1.2%;检出口蹄疫病毒非结构蛋白3ABC抗体阳性样品31份,平均阳性率11.1%;检出牛传染性鼻气管炎阳性样品14份,平均阳性率5.0%;检出弓形虫抗体阳性样品87份,平均阳性率31.1%;检出牛副结核抗体阳性样品1份,平均阳性率0.4%。[结论]通过血清学检测,初步掌握了5种疫病的感染情况,为科学防控提供了科学依据。     

民和县某一养殖合作社羊布病疫情应急处置

对来自青海省海东市民和县的244份牛羊血清样品(某合作社羊133份、牛7份,相邻散养户羊104份),采用虎红平板凝集试验和试管凝集试验进行了布病特异性抗体检测。结果在合作社的血清样品中检出阳性25份,阳性率为18.8%,牛的血清样品中未检出;在散养户的血清样品中检出阳性1份,阳性率为0.96%。结合该羊场饲养员确诊已患布病,经综合分析后,认为该羊群存在布病感染,对检出的26只布病阳性羊进行了扑杀等无害化处理。     

规模猪场种猪布鲁氏菌病血清学调查

猪布鲁氏菌病是由猪布鲁氏杆菌（B.suis）引起的流产、睾丸炎等为特征的人畜共患病，危害畜牧业健康与公共卫生安全。为了了解延平区规模猪场种猪布鲁氏菌病感染情况，2013年对23个规模猪场901份血清样品采用琥红平板疑集试验和试管疑集试验进行初检和确诊。结果表明：被检的23个规模猪场有1个猪场检出阳性，被检血清中1份样品阳性，猪场阳性率为4.3%，样品阳性率为0.1%。     

富宁县蓝舌病血清学调查

为了解富宁县牛羊蓝舌病的感染情况,2015—2016年,连续两年从新华、板仑、归朝、谷拉、者桑等13个乡(镇)的养殖户采集羊血清样品212份、牛血清样品478份,应用B-ELISA方法检测蓝舌病抗体。在被检的212份羊血清中,检出阳性样品86份,阳性率为40.57%;478份牛血清中,检出阳性样品232份,阳性率为48.54%。调查结果表明富宁县牛、羊中存在蓝舌病病毒感染的情况。     

青海省海晏县绵羊布氏杆菌病的调查

用布氏杆菌试管凝集试验(SAT),对海晏地区的绵羊进行了布氏杆菌血清抗体的检测,共检397份血清样品,检出阳性血清2份,阳性率为0.50%;其中:成年母羊血清233份,检出阳性1份,阳性率为0.43%;后备母羊血清91份,检出阳性1份血清,阳性率为1.10%;羯羊血清52份,阳性率为0%;种公羊21份,也未检出阳性。     

牦牛病毒性腹泻病PCR诊断方法的建立

采用阻断ELISA方法对来自天峻县的327份牦牛血清进行了牛BVDV特异性抗体的检测,共检出231份血清阳性,血清阳性率为70.64%。其中:在135份野牦牛血清中检出110份阳性血清,阳性率为81.48%;在192份家牦牛血清中检出阳性121份,阳性率为63.02%。从231份ELISA检测阳性血清中随机抽取了20份,采用RT-PCR方法进行检测,结果共检出18份样品为BVDV/MV RT-PCR阳性,阴性血清未扩增出E0基因,RT-PCR方法检测结果与ELISA方法符合率为90%。     

西藏那曲地区蓝舌病血清流行病学调查

为了解那曲地区牛羊蓝舌病的流行情况,从双湖、尼玛、嘉黎、安多和聂荣等5个县采集羊血清样品904份、牛血清样品739份,应用ELISA抗体检测法对蓝舌病进行血清学调查。在被检的904份羊血清中,检出阳性样品8份,平均阳性率为0.88%,在牛血清中未检测出阳性样本。调查结果表明那曲地区羊群中存在蓝舌病病毒的感染。     

北京地区规模化奶牛场牛病毒性腹泻病血清学调查

对北京郊区6个未进行牛病毒性腹泻病（BVD）免疫牛场的546份奶牛血清样品,使用牛病毒性腹泻抗体ELISA试剂盒进行检测,并对其中3个牛群进行牛病毒性腹泻病毒（BVDV）血清抗原筛查。共检出阳性血清514份,总阳性率94.1%,其中5个牛场的场内血清抗体阳性率在95%以上。牛病毒性腹泻持续性感染牛（PI牛）筛查的3个牛群均有阳性牛检出。结果表明,北京地区规模化奶牛场存在牛病毒性腹泻感染和接触史,应采取净化措施进行控制。     

蚌埠市某奶牛场IBRV和BVDV的血清学调查

本试验对蚌埠市一家未进行牛传染性鼻气管炎及病毒性腹泻免疫的规模化奶牛场的94份奶牛血清样品,分别使用牛传染性鼻气管炎及病毒性腹泻抗体ELISA试剂盒进行检测,共检出牛传染性鼻气管炎阳性血清65份,阳性检出率为69．15％;病毒性腹泻阳性血清83份,阳性检出率为88．30％。结果表明,该规模化奶牛场存在牛传染性鼻气管炎及病毒性腹泻的感染和接触史,应采取净化措施进行控制。     

Detection of Infectious Bovine Rhinotracheitis and Bovine Viral Diarrhea Viruses in the Nasal Epithelial Cells by the Direct Immunofluorescence Technique

Nasal epithelial cells were collected by cotton swabs for the diagnosis in experimental and field cases of infectious bovine rhinotracheitis and field cases of bovine viral diarrhea in calves. A portion of the cells was washed twice in phosphate buffered saline and a 25 µL drop was placed on microscope slides. The cells were dried, fixed and stained according to the direct fluorescent antibody technique. Another portion of the same specimen was inoculated onto primary bovine skin cell cultures for virus isolation. In the experimental studies for infectious bovine rhinotracheitis, 29/35 specimens were positive by fluorescent antibody technique and 32/35 by cell culture and in the field cases, 22/119 were positive by fluorescent antibody technique and 19/119 by cell culture. In the field cases of bovine viral diarrhea, 28/69 samples were positive by fluorescent antibody technique and 14/69 by cell culture. When fluorescent antibody technique was performed on inoculated cell cultures a total of 24/69 specimens were positive for bovine viral diarrhea. The sensitivity of fluorescent antibody technique was thus comparable to that of cell culture method for infectious bovine rhinotracheitis and bovine viral diarrhea.     

Effect of maternal antibody upon vaccination with infectious bovine rhinotracheitis and bovine virus diarrhea vaccines.

This report presents the normal rate of decay of maternal antibody and the influence of maternal antibody on responses to a single vaccination with modified-live bovine virus diarrhea and infectious bovine rhinotracheitis virus vaccines at 196 days of age and on response to vaccinations with the same vaccines given twice at 84 and 196 days of age. Passive immunity decreased to near zero over the first six months of life for both bovine virus diarrhea and infectious bovine rhinotracheitis controls. All calves seroconverted to bovine virus diarrhea vaccine at 84 days of age, even though high levels (greater than 1:32) of maternal antibodies were present. These calves did not seroconvert to infectious bovine rhinotracheitis vaccine at 84 days of age when high levels (less than 1:16) of maternal antibodies were present. Calves responded well to bovine virus diarrhea and infectious bovine rhinotracheitis vaccines given only once at 196 days of age after passive immunity disappeared. Calves which were revaccinated with infectious bovine rhinotracheitis seroconverted showing a more rapid response than the single vaccinates. Those revaccinated with bovine virus diarrhea showed an immediate response of small magnitude.     

内蒙古地区部分奶牛场牛传染性鼻气管炎的血清学调查

为了解内蒙古地区牛传染性鼻气管炎在奶牛场的流行情况,对内蒙古地区的16个大、中、小型奶牛养殖场的2 321头奶牛,应用ELISA方法进行牛传染性鼻气管炎的血清流行病学调查。结果显示,5个大型奶牛场牛传染性鼻气管炎的阳性率在68.5%~100%之间,5个中型奶牛场的阳性率在34.0%~84.2%之间,6个小型奶牛场的阳性率在32.6%~89.3%之间,平均阳性率为75.7%。     

牦牛病毒性腹泻/黏膜病和传染性鼻气管炎血清学调查

为掌握青海省大通种牛场和海晏县牦牛病毒性腹泻/黏膜病、传染性鼻气管炎的感染和流行情况,2010年3月至5月在大通种牛场和海晏地区采集252份牦牛血清样品,应用定量酶联免疫吸附试验（ELISA）对牦牛病毒性腹泻/黏膜病和传染性鼻气管炎的进行了血清抗体检测。结果显示,大通种牛场牦牛群中牛病毒性腹泻/黏膜病阳性率为23.42%,传染性鼻气管炎阳性率为65.45%;海晏县牦牛群中牛病毒性腹泻/黏膜病阳性率为19.86%,传染性鼻气管炎阳性率为4.96%。     

套式RT-PCR检测牛呼吸道合胞体病毒的研究

为了检测牛呼吸道合胞体病毒(BRSV),根据已发表的融合蛋白(F)基因序列,设计了套式RT-PCR引物,初步建立了BRSV的套式RT-PCR检测方法。对138份牛鼻腔棉拭子进行了检测,结果检测到了BRSV阳性样品54份,总的阳性检出率为39.1%。对大部分BRSV阳性样品的F基因扩增产物进行了序列测定与分析。用该套式RT-PCR对牛传染性鼻气管炎病毒、牛副流感病毒3型、牛病毒性腹泻病毒等进行了检测,结果无交叉反应,表明该检测方法具有良好的特异性。本研究首次用套式RT-PCR技术证实了我国部分省的牛群中存在BRSV感染。     

牛传染性鼻气管炎病毒gE基因的截短克隆与表达

以牛传染性鼻气管炎病毒Baaha Nu/67株的DNA作为模板，用PCR扩增gE基N并克隆至pGEM-T Easy裁体，再以此质粒作为模板将gE基因分成6个片段，分别插入原核表达载体pET32a并在大肠杆菌中进行了表达。蛋白电泳结果表明6个片段中有2个片段以可溶形式表达，1个片段以包涵体形式表达，另外3个片段没有表达。采用固定化金属离子亲和层析法在非变性条件下对两个可溶性片段进行了纯化。经免疫印迹试验，间接ELISA和交叉试验证明，两个纯化的重组蛋白均与牛传染性鼻气管炎阳性血清样品发生反应，而与牛传染性鼻气管炎阴性血清无任何反应，显示其具有良好的抗原性和特异性，可用于牛传染性鼻气管炎gE-ELISA诊断方法的建立。     

牛病毒性腹泻的鉴定以及治疗

为了鉴定甘肃武威地区某牛场中牛病毒性腹泻病的发病情况,分别采集了13只病牛的血液样品、粪便样品配合诊断,通过血清学诊断方法和PCR鉴定的方法对样本进行了检测,结果显示血清学方法中的13份血液样本有8份为阳性,粪便样本提取RNA,进行RT-PCR同样可以检测到8份样本中扩增得到片段大小为267bp的条带,检测的13份样本有8份样本的血清学检测和粪便PCR检测均为阳性,本次检测的阳性率为61.54%,结果表明本次检测的甘肃武威地区某养殖场的牛病毒性腹泻阳性率相对较高,需要加强牛病毒性腹泻的防控,采取科学的手段治理该类疾病降低其对养牛产业的损失。     

四川部分地区牦牛传染性鼻气管炎血清学检测报告

[目的]为了解四川省牦牛传染性鼻气管炎感染的流行情况。[方法]应用酶联免疫吸附试验对在2012年间采自对四川省甘孜藏族自治州和红原县共377份牦牛血清进行牛传染性鼻气管炎病毒抗体检测。[结果]检出阳性血清样品共105份,阳性率为27.9%。[结论]表明四川省牦牛群中均存在牛传染性鼻气管炎病毒的感染。该研究为该地区牦牛传染性鼻气管防治提供依据。     

Viral RNA extraction using an automatic nucleic acid extractor with magnetic particles and genetic characterization of bovine viral diarrhea virus in Tokachi Province,Japan, in 2016–2017

In this study, the viral genome extraction performance of automatic nucleic acid extractors and manual nucleic acid extraction kits was compared. We showed that compared with manual kits, the automatic extractors showed superior genome extraction performance using bovine viral diarrhea virus (BVDV) genome-positive cattle sera and bovine coronavirus/infectious bovine rhinotracheitis virus-spiked cattle nasal swabs. In addition, the subgenotyping of BVDV strains detected in Tokachi Province in Japan during 2016–2017 was performed. Results showed that most of these BVDV strains belonged to subgenotype 1b, while few strains belonged to subgenotypes 1a and 2a. This study showed the high applicability of automatic nucleic acid extractors in extracting multiple viral genomes and the dominant subgenotype of BVDV in Tokachi.