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2025年5月16日 星期五
农业生物技术学报  2019, Vol. 27 Issue (2): 330-336    DOI: 10.3969/j.issn.1674-7968.2019.02.016
  研究论文与报告 本期目录 | 过刊浏览 | 高级检索 |
肠炎沙门菌sipC缺失株构建及部分生物学特性分析
张钰1,2, 陈阳1, 徐琪1, 朱国强2, 陈国宏1*
1 扬州大学 动物科学与技术学院,扬州225009;
2 扬州大学 江苏省人兽共患病学重点实验室,扬州225009
Construction and Partial Biological Characterization Analysis of the sipC Deletion Strain of Salmonella enteritidis
ZHANG Yu1,2, CHEN Yang1, XU Qi1, ZHU Guo-Qiang2, CHEN Guo-Hong1*
1 College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China;
2 Jiangsu Key Laboratory of Zoonosis, Yangzhou University, Yangzhou 225009, China
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摘要 肠炎沙门菌(Salmonella enteritidis, SE)可长期定植于家禽的肠道与生殖道组织,造成禽蛋的直接污染和垂直传播,难于清除与预防,不仅影响产蛋量,还造成环境污染,危害人类健康。Ⅲ型分泌系统(type secretion system, T3SS)的效应蛋白在肠炎沙门菌与宿主细胞表面黏附时至关重要。为研究肠炎沙门菌T3SS1主要效应蛋白SipC (type secretion system-1 effector SipC, T3SS1-SipC)在感染中的作用,本研究利用λRed同源重组系统构建肠炎沙门菌MY1的sipC基因缺失株MY1ΔsipC,同时将sipC基因克隆至载体质粒pBR322,构建回补株MY1ΔsipC/psipC。比较分析了野生株MY1、缺失株MY1ΔsipC以及回补株MY1ΔsipC/psipC在生长特性、对人(Homo sapiens)结肠细胞(human colorectaladeno carcinoma cell line, Caco-2)、鸭(Anas platyrhynchos)卵泡颗粒细胞(duck granulosa cells, dGCs)的黏附和侵袭能力,以及雏鸭半数致死量的差异。结果显示,缺失sipC基因后,肠炎沙门菌生长特性不变;感染2 h后,对Caco-2黏附和侵袭数量分别下降44.9%和34.5%;对dGCs黏附和侵袭数量分别下降37.9%和30.4%;对雏鸭的致病性显著降低(P<0.05)。本研究初步证实sipC参与肠炎沙门菌的致病力,为进一步探索sipC在肠炎沙门菌黏附宿主的分子机制提供实验材料。
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张钰
陈阳
徐琪
朱国强
陈国宏
关键词 肠炎沙门菌sipC基因缺失株毒力    
AbstractSalmonella enteritidis (SE) always colonized in the intestinal and reproductive tracts of the poultry, causing direct contamination and vertical transmission of eggs that were difficult to eradicate and prevent. It not only affected the egg production, but also caused egg pollution as well as environmental pollution and imperiled human health. The effect proteins of the type secretion system (T3SS) played a critical role when SE attached to the host cell. In order to study the role of the T3SS1 major effect protein SipC during SE infection, the sipC deletion mutant strain of SE MY1 were constructed by using λRed homologous recombination system, and a complementary strain MY1ΔsipC/psipC was constructed by transformation of the a plasmid pBR322 carrying the sipC gene cloned from MY1. The growth curve, the ability of adhesion and invasive to human (Homo sapiens) colorectaladeno carcinoma cell line (Caco-2) and duck (Anas platyrhynchos) granulosa cell (dGC) of the wild-type strain MY1, mutants MY1ΔsipC and MY1ΔsipC/psipC in vitro were measured. And the strains were orally inoculated to 7-day-old shaoxing ducklings separately to study the pathogenicity of sipC in vivo. The results showed that the mutants MY1ΔsipC and MY1ΔsipC/psipC were successfully constructed and SE MY1 contained a sipC gene with 100% identity to the S. typhimurium. No significant difference was observed between the parental strain MY1 and sipC mutant strains in growth curve. However, after 2 h of infection, the ability of SE MY1ΔsipC adhering to Caco-2 and dGCs were decreased about 44.9% and 34.5%, respectively, as compared with the wild-type adherence, and the invasive ability of SE MY1ΔsipC to Caco-2 and dGCs were also reduced 37.9% and 30.4%, respectively. The virulence of the sipC mutant was significantly reduced in a 7-day-old duckling model of SE disease (P<0.05), as determined by quantifying the lethal dose 50% of the bacterial strains. Collectively, a novel function of sipC in contributes to SE virulence was covered and experimental reference of the interaction between sipC and host genes were offered for further study.
Key wordsSalmonella enteritidis    sipC    Deletion strain    Virulence
收稿日期: 2018-09-16     
ZTFLH:  S852.6  
基金资助:国家自然科学基金(No.31702107)和江苏省人兽共患病学重点实验室资助项目(No.R1806)
通讯作者: *,ghchen@yzu.edu.cn。   
引用本文:   
张钰, 陈阳, 徐琪, 朱国强, 陈国宏. 肠炎沙门菌sipC缺失株构建及部分生物学特性分析[J]. 农业生物技术学报, 2019, 27(2): 330-336.
ZHANG Yu, CHEN Yang, XU Qi, ZHU Guo-Qiang, CHEN Guo-Hong. Construction and Partial Biological Characterization Analysis of the sipC Deletion Strain of Salmonella enteritidis. 农业生物技术学报, 2019, 27(2): 330-336.
链接本文:  
http://journal05.magtech.org.cn/Jwk_ny/CN/10.3969/j.issn.1674-7968.2019.02.016     或     http://journal05.magtech.org.cn/Jwk_ny/CN/Y2019/V27/I2/330
 
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