文章摘要
枯草芽孢杆菌表达柱状黄杆菌lip基因口服疫苗对草鱼免疫保护效果的研究*
Oral vaccination of grass carp Ctenopharyngodon idella against Flavobacterium columnare using Bacillus subtilis expressing lip gene
投稿时间:2020-07-17  修订日期:2020-09-08
DOI:
中文关键词: 柱状黄杆菌  枯草芽孢杆菌  草鱼  口服疫苗
英文关键词: Flavobacterium colunmare  Bacillus subtilis  Grass carp  Oral vaccine
基金项目:国家大宗淡水鱼产业技术体系建设专项(CARS-45-33) ;天津市自然科学基金(16JCYBJC30000) ;水产增养殖安徽省重点实验室开放课题(AHSC20190203);天津市水产研究所创新团队项目(scyjs201904)。
作者单位E-mail
罗璋 水产增养殖安徽省重点实验室 luozhang2010@163.com 
张振国 天津市水产研究所
天津市水产研究所 
 
郝爽 天津市水产研究所  
陈红莲 水产增养殖安徽省重点实验室  
冯守明① 天津市动物疫病预防控制中心
天津市动物疫病预防控制中心 
smfeng65@163.com 
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中文摘要:
      枯草芽孢杆菌是一种水产益生菌,最近也被用作口服疫苗的投递载体。本研究利用前期筛选到的柱状黄杆菌保护性抗原基因lip与pBE2R载体连接后,以质粒的方式转入枯草芽孢杆菌WB800中构建了重组菌株WB800(pBE2R-lip)。通过聚丙酰胺电泳和免疫印迹实验证明枯草芽孢杆菌WB800(pBE2R-lip)能表达lip蛋白,其分子量大小约为32 KDa。利用WB800(pBE2R-lip)口服免疫草鱼,通过酶联免疫吸附实验在受免草鱼血清中检测到特异性抗体效价的上升,在人工感染实验中受免草鱼的免疫保护率为52.4%。本研究结果表明利用枯草芽孢杆菌能有效表达柱状黄杆菌的lip基因,以其作为口服疫苗能引起草鱼的免疫应答和提供较好的免疫保护效果。
英文摘要:
      Bacillus subtilis, being consumed as probiotics in aquacuture, have been explored as live carriers for expression and oral delivery of antigen proteins. InSaSpreviousSstudy, we found a protective antigen gene lip of Flavobacterium columnare, the pathogen of gill-rot disease in freshwater fish. In this paper, lip gene was inserted into a shuttle expression vecter pBE2R , and then expressed as a recombinant fusion protein in B. subtilis WB800. The expressed protein of 32 KDa, as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western blotting, was consistent with the molecular weight deduced from the amino acid sequence. Grass carp Ctenopharyngodon idella was immunized by oral delivery of B. subtilis WB800 (pBE2R-lip). Then, the effect of protection and the immune responses were evaluated. The results of ELISA showed that lip-specific antibody in the sera of immunized fish can be detected when compared to the control group. The outcome of fish immunized with B. subtilis WB800 (pBE2R-lip) provided a relative percent survival (RPS) of 52.4% against F. columnare. Our study demonstrated that the oral administration of B. subtilis expressing lip could cause an effective immune response and offer good resistance to F. columnare.
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