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微生物学报

kpsEkpsD双基因缺失显著降低肠道外致病性大肠杆菌的毒力
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国家自然科学基金(31272559,30972196,30771604,30471281);农业部公益性行业专项(201303044);中国博士后科学基金项目(2015M580477);江苏省博士后基金项目(1501076C);江苏省自然科学基金(BK20140485,BK20151308);江苏省高校自然科学基金(14KJB230001);江苏高校优势学科建设工程资助项目(PAPD);扬州大学科技创新培育基金项目(2014CXJ051,2015CXJ057)


Dual deletion of the kpsE and kpsD genes reduced the bacterial virulence of extraintestinal pathogenic Escherichia coli
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    摘要:

    [目的]探究荚膜对肠道外致病性大肠杆菌致病作用的影响。[方法]选取负责荚膜多糖转运的基因kpsEkpsD,利用λ Red重组系统构建APEC E058和UPEC U17荚膜缺失株E058ΔkpsED和U17ΔkpsED,并通过一系列的体内及体外试验对其生物学特性及致病性进行研究。[结果]双基因缺失株的生长速度较野生株没有明显差异,但缺失株抗血清补体杀菌能力和抗鸡巨噬细胞HD-11细胞吞噬能力显著下降。1日龄雏鸡LD50致病性试验结果显示,缺失株E058ΔkpsED和U17ΔkpsED对鸡失去致病力,而回复株毒力恢复至野生株水平;35日龄SPF鸡体内动态分布和竞争试验显示ΔkpsED缺失株在鸡体内定殖能力和竞争性生长能力显著下降,表明kpsED双基因的缺失能显著降低APEC E058和UPEC U17的致病力。[结论]荚膜与肠道外致病性大肠杆菌的致病性相关,是其重要的毒力因子。

    Abstract:

    [Objective] To study the role of capsule polysaccharide in pathogenesis of extraintesinal pathogenic Escherichia coli (ExPEC). [Methods] By using λ Red recombination system, we generated the capsular polysaccharide transport associated genes kpsE and kpsD double knockout mutants E058ΔkpsED and U17ΔkpsED. We then compared and analyzed the characteristics of the mutant strains and wild-type strains. [Results] The growth curves in Luria Bertani showed that the deletion of kpsED did not affect growth kinetics of the mutants. The abilities of resistance to serum and killing by chicken macrophages were significantly impaired. LD50 results showed that the double mutants completely abolished the virulence, whereas the complementation strains restored the virulence to resemble that of wild-type strains, and the colonization and coinfection model demonstrated that the deletion of kpsED led to attenuation of virulence, because the double mutant showed significantly decreased colonization compared with the wild-type strains in all organs tested in chickens. [Conclusion] These results indicated that the virulence factors encoded by capsule genes were important for the pathogenesis of ExPEC.

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高清清,夏乐,刘娟华,高崧,刘秀梵. kpsEkpsD双基因缺失显著降低肠道外致病性大肠杆菌的毒力. 微生物学报, 2016, 56(10): 1571-1582

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  • 收稿日期:2015-12-24
  • 最后修改日期:2016-03-05
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  • 在线发布日期: 2016-10-11
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