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迟缓爱德华氏菌两个外膜蛋白的功能及其亚单位疫苗的研究
金成东
Subtype硕士
Thesis Advisor李墨非
2021-05-17
Degree Grantor中国科学院大学
Place of Conferral中国科学院海洋研究所
Degree Name工程硕士
Keyword迟缓爱德华氏菌 外膜蛋白 毒力 疫苗
Abstract

迟缓爱德华氏菌(Edwardsiella tarda)是一种重要的鱼类病原菌,能引发多种养殖鱼类的严重病害,给水产养殖业造成巨额的经济损失。细菌外膜蛋白是细菌细胞的重要组成部分。本研究以迟缓爱德华氏菌的外膜蛋白为切入点,分析并研究了两个迟缓爱德华氏菌的外膜蛋白功能(命名为:Omp1EtOmp2Et)。研究发现Omp1EtOmp2Et位于迟缓爱德华氏菌的外膜表面,推测是与毒力相关的外膜蛋白。重组表达的Omp1EtOmp2Et蛋白(rOmp1EtrOmp2Et)能够与牙鲆外周血淋巴细胞结合。为了进一步研究这两个外膜蛋白的功能,我们利用缺失敲除的方式,构建了两株迟缓爱德华氏菌的基因敲除株,即基因Omp1敲除株TX01Δomp1Omp2基因敲除株TX01Δomp2。敲除株TX01Δomp1TX01Δomp2的毒力与野生株相比,在致死率,细胞侵染和黏附,运动性及其生物膜形成能力方面显著降低。与野生株相比,敲除株TX01Δomp1侵染组织的能力明显下降;TX01Δomp1TX01Δomp2在过氧化氢中的存活率显著降低 TX01Δomp1在酸性条件下的存活率显著降低。利用rOmp1EtrOmp2Et构建的亚单位疫苗免疫牙鲆,发现rOmp1EtrOmp2Et亚单位疫苗能够增强牙鲆产生的免疫反应,帮助牙鲆抵御迟缓爱德华氏菌的感染。综上所述,研究表明Omp1EtOmp2Et是位于迟缓爱德华氏菌外膜表面的毒力相关因子,并且Omp1Et作为亚单位疫苗,能诱导牙鲆免疫系统产生良好的免疫反应。本研究首次证实了Omp1EtOmp2Et是位于迟缓爱德华氏菌外膜表面的毒力相关因子,构建的Omp1EtOmp2Et亚单位疫苗对于牙鲆感染迟缓爱德华氏菌有良好的免疫保护效应。

Other Abstract

Edwardsiella tarda is a vital bacterial pathogen to a wide range of aquaculture animals, causing severe lesions and economic loss. The bacterial outer membrane proteins are important components of bacterial cell. In our study, we focused on two outer membrane proteins of E. tarda. We analyzed and examined the function of two outer membrane proteins of E. tarda (named Omp1Et and Omp2Et). The Omp1Et and Omp2Et were founded to localize on the surface of E. tarda and were predicted as virulence-related proteins. The recombinant Omp1Et and Omp2Et were interacted with the peripheral blood leukocytes of Paralichthys olivaceus. We knockout gene Omp1 or Omp2, and constructed the mutant strains TX01Δomp1 or TX01Δomp2. Compared with the wild type, Omp1 knockout TX01Δomp1 and Omp2 knockout TX01Δomp2 were significantly reduced in motility, biofilm formation, attachment and invasion into host cells and inducing fish mortality. Furthermore, compared with the wild type, TX01Δomp1 was significantly impaired in tissue dissemination. Meanwhile, compared with the wild type, the survival ability of TX01Δomp1 and TX01Δomp2 were significantly reduced  under oxidizing conditions while the survival ability of TX01Δomp1 were significantly reduced under low pH. When introduced into flounder as a subunit vaccine, rOmp1Et and rOmp2Et subunit vaccines induced strong immune protection against E. tarda challenge. Taken together, our studies indicated that Omp1Et and Omp2Et were surface-located virulence factors and rOmp1Et can induce effective immune-protection as a subunit vaccine. Our study firstly indicated that Omp1Et and Omp2Et were surface-located virulence factors, and constructed Omp1Et and Omp2Et subunit vaccines could induce effective immune-protection to Paralichthys olivaceus from infection of E. tarda.

Subject Area生物学
MOST Discipline Catalogue工学 ; 工学::生物工程
Funding ProjectNational Natural Science Foundation of China[31972831] ; National Key Research and Development Program of China[2018YFD0900501] ; Huiquan Young Scholar Program of Institute of Oceanology, CAS ; Youth Innovation Promotion Association of the Chinese Academy of Sciences[2017249] ; Taishan Scholar Program of Shandong Province ; Taishan Scholar Program of Shandong Province ; Youth Innovation Promotion Association of the Chinese Academy of Sciences[2017249] ; Huiquan Young Scholar Program of Institute of Oceanology, CAS ; National Key Research and Development Program of China[2018YFD0900501] ; National Natural Science Foundation of China[31972831]
Language中文
Document Type学位论文
Identifierhttp://ir.qdio.ac.cn/handle/337002/170648
Collection实验海洋生物学重点实验室
Recommended Citation
GB/T 7714
金成东. 迟缓爱德华氏菌两个外膜蛋白的功能及其亚单位疫苗的研究[D]. 中国科学院海洋研究所. 中国科学院大学,2021.
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