猪源牛病毒性腹泻病毒1型分离株的鉴定及培养特性研究  

Identification of bovine viral diarrhea virus type 1 isolate from pig origin and its characters on cellular culture

作　　者：陶洁[1,2] 李本强 程靖华[1,2] 马玉飞 刘惠莉[1,2] 

机构地区：[1]上海市农业科学院畜牧兽医研究所上海市农业遗传育种重点实验室,上海201106 [2]上海市种猪工程技术中心,上海201106

出　　处：《中国兽医科学》2017年第12期1555-1561,共7页Chinese Veterinary Science

基　　金：动物重大疫病新型广谱中和抗体疫苗研发项目(2017YFD0501102);上海市扬帆计划项目(15YF1410300);上海市农口青年成长计划项目[沪农青字(2016)第1-35号];上海市科技兴农重点推广项目[沪农科推字(2017)第1-11号];上海市科技兴农重点攻关项目[沪农科攻字(2013)第3-6号;沪农科攻字(2013)第5-5号;沪农科产字(2017)第6号]

摘　　要：利用RT-PCR方法从猪场腹泻样品中检测到牛病毒性腹泻病毒(BVDV)核酸阳性样品,测序证实为1型BVDV,命名为SHCM06。遗传进化分析表明,其与ZM-95亲缘性最近,同源性达98.3%;与BVDV-1经典毒株亲缘性较远,仅85.7%~89.8%。免疫荧光、Western-blot和荧光定量RT-PCR检测均表明,SHCM06毒株可感染MDBK、ST和PK15细胞,在MDBK和ST细胞上的增殖效率显著高于PK15细胞。进一步分析SHCM06感染对细胞中Ⅰ型干扰素表达的影响,结果显示,SHCM06感染均能使3种细胞中IFN-α和IFN-β表达量显著下降(P<0.05),且对MDBK细胞IFN-α的抑制效果显著高于对PK15细胞中IFN-α的抑制作用(P<0.05)。结果表明,从猪临床腹泻样品中分离1株BVDV1型,该病毒对MDBK、ST细胞适应性好于PK-15细胞,能显著抑制细胞中Ⅰ型IFN的产生,以发挥抵抗细胞抗病毒免疫效应,本试验为深入研究猪源BVDV的致病作用奠定了基础。In this study,a bovine viral diarrhea virus type I（BVDV-I） strain named SHCM06 was isolated from the porcine fecals identified by RT-PCR and sequencing. Phylogenetic analysis indicated that the virus was most closely to the BVDV-I strain ZM-95 with nucleotide homology of 98.3%,but shared a far distant with the classical strains in BVDV-I subgenotype （85.7%-89.8%）.Immune florescence assay,Western-blot and real-time RT-PCR detection confirmed that SHCM06 could proliferate in MDBK,ST and PK15 cells successfully and grew better in MDBK and ST cells rather than in PK15 cells. Besides,SHCM06 could inhibit the expression of IFN-α and IFN-β in the three kinds of cells（P〈0.05） with significant differences between MI）BK and PKI5 cells on IFN-α production. In conclusion,we identified one BVDV-I isolate from porcine fecals,which had good replication effective in MDBK and ST cells and could inhibit the type I interferonproduction significantly. Thisresearchwillbe helpful for its furtherpathogenicity studies.

关 键 词：猪源BVDV 同源性分析 生长特性 Ⅰ型干扰素 

分 类 号：S852.659.5[农业科学—基础兽医学]