用相同量猪肺炎支原体(Mhyo)和不同量猪圆环病毒2型(PCV2)混合后的4组疫苗分别免疫BALB/c小鼠、新西兰大耳白兔和健康仔猪,测定小鼠、兔和仔猪血清中PCV2和Mhyo抗体水平,并在仔猪二免后进行Mhyo济南系强毒攻击.结果:小鼠和兔免疫疫苗后产生的Mhyo抗体水平随着疫苗中PCV2抗原含量的增加而提高.仔猪免疫疫苗后,ELISA与IHA检测结果都显示Myho抗体水平随着疫苗中PCV2抗原含量的增加而提高;Mhyo攻毒后肺部病变程度也与疫苗中PCV2抗原浓度呈负相关.以上结果说明PCV2、Mhyo二联疫苗中PCV2全病毒抗原对Mhyo免疫有增强作用,且随着PCV2全病毒抗原含量的增加,Mhyo的免疫保护效果也随之增强,这个结果提示了 PCV2抗原对Mhyo抗原的免疫效果有促进作用.

本研究通过整理分析2012～2018年国内获得临床试验批件的兽用生物制品的数量、分类、治疗靶动物、申报单位、成功转化新兽药情况等内容,挖掘兽用生物制品领域的研究开发趋势和方向.

猪瘟是严重危害我国和世界养猪业发展的重要传染病.我国研制的猪瘟兔化弱毒疫苗(即C株)在世界范围内的猪瘟免疫防控和最终根除过程中起到了极其重要的作用.尽管如此,目前我国养猪业仍在承受该病的困扰,因种种原因造成的疫苗免疫力下降或免疫失败现象时有发生,每年都让养殖业蒙受巨大的经济损失.文章就我国猪瘟流行特点、现有猪瘟疫苗特点及合理使用猪瘟ST传代细胞苗、科学免疫猪瘟等问题作一概述和探讨分析.

猪圆环病毒2型(Porcine circovirus 2,PCV2)是引起猪圆环病毒病(Porcine circovirus disease,PCVD)的主要病原.该病毒可破坏动物机体免疫系统,引起严重的免疫抑制,最终导致猪的生产性能下降、进而造成巨大的经济损失.目前,国内共有6种不同的PCV2疫苗获得了新兽药注册证书.文章就目前国内外PCVD的流行状况、我国当前PCV2疫苗特点及PCVD的科学免疫等方面作一简述,以期为PCVD的疫苗选择和科学防控等提供参考和借鉴.

To obtain the monoclonal antibodies of Mycoplasma hyopneumoniae(Mhp),Mhp 168 strain was used as immunogen to immunize BALB/c mice.With the hybridoma technology,three hybridoma cell lines which steadily secret monoclonal antibody against DnaK protein were obtained,separately named 3A8,5D10 and 3E2.The isotype of the 3E2,3A8 and 5D10 was IgG1,κ chain.Western blot showed that all of the McAbs could react with Mhp168 isolates and the DnaK recombinant protein,and had no reaction activity with vector protein and Mycoplasma hyorhinitis(Mhr) except for 3A8.The McAbs of 3E2 and 5D10 also specifically reacted with 168 strain F105,J strain,XLW-1 isolate,XLW-2 isolate,XLW-3 isolate,XLW-4 isolate,AH isolate,and WX isolate of Mhp.Indirect ELISA revealed the titers of the ascites of the three monoclonal antibodies against DnaK recombinant protein and Mhp 168 protein were 1∶1 000 000.3A8 was the monoclonal antibody which reacted with the common antigen epitopes of Mhp and Mhr;3E2 and 5D10 were the specific monoclonal antibodies against Mhp,both of them with high titers.

Balb/c mice were immunized with Mycoplasma hyopneumoniae(Mhp) strain 168,and the splenocytes of the immunized mice were fused with SP2/0 cells.After subcloning for 3 times,four hybridoma clones which produced Monoclonal Antibodies(McAbs) steadily were screened by ELISA,named 2F5,2G7,4F5,and 5E9.The result of Western-blot indicated that 2G7 and 4F5 both strongly specifically reacted with the Mhp strain 168.Both of the McAbs(2G7 and 4F5) belonged to IgG1 isotype,and their light chains were κ type.However,2F5 and 5E9 belonged to IgG2a isotype,and their light chains were also κ type.They all reacted strongly with Mhp strain 168 in immunofluorescence assay(IFA).The McAbs could be used for the detection and pathogenic mechanism research of Mhp.

反向疫苗学是通过对病原基因组测序和计算机预测,筛选出可能编码疫苗抗原的基因,通过高通量克隆、表达纯化技术及免疫原性的检测,制备具有保护力的疫苗,为传统方法无法制备疫苗的病原微生物提供了一种新的研究思路与方法.本文介绍了反向疫苗学的基本流程,并对近几年反向疫苗学在猪病病原微生物方面的研究进展进行了综述.

Mycoplasma hyopneumoniae （Mhp） is the etiologic agent of mycoplasma pneumonia in swine. In this study the DnaK gene of strain NJ is amplified by gene splicing with overlap extension PCR using a pair of inside-mutation primers. The PCR product is inserted into expression plasmid pET-28a （＋）. The recombinant plasmid is transformed into competent cells of BL21（DE3） and it contains correct object fragments by sequencing. It is induced to express by 1 mmol/L IPTG at 37 ~C for 5 hours. The dose of the recombinant protein is nearly 14. 1~ of the total product. The expression product of interest and its biological activities are characterized with Western blotting and ELISA analysis. This study provides the basis for diagnose agent and future preparation of new vaccine a~ainst Mvconlasma hvonneurnnniae.

本研究将附红细胞体的MSG1蛋白基因按大肠杆菌密码子偏嗜性改造后进行全基因人工合成,合成的基因连接入pBAD/HisB载体后导入大肠杆菌Top10感受态细胞中,进行MSG1诱导表达,确定诱导表达的最佳时间和最佳诱导剂浓度。对表达的蛋白应用His单抗进行了特异性鉴定,对鉴定后的蛋白应用Ni-NTA纯化试剂进行了不同条件下的蛋白纯化。试验结果表明,诱导蛋白的分子量在45 ku左右,加入0.2%的L-阿拉伯糖,诱导2 h后所表达的蛋白量最高。经Western-blot鉴定,该蛋白可以被His单抗特异性识别,经纯化后可以得到较高纯度的MSG1蛋白。表达的MSG1蛋白经纯化后免疫家兔制备多克隆抗体,Western blot证明制备的抗体具有高度的特异性。MSG1蛋白的表达和多抗的制备可以为后续研究附红细胞体的吸附机制、致病机理,以及附红细胞体病的治疗等提供理论基础。