胸膜肺炎放线杆菌relA基因缺失株的构建及其生物学特性的研究

发布时间：2018-05-07 15:07

本文选题：胸膜肺炎放线杆菌 + relA基因　；参考：《中国预防兽医学报》2017年12期

【摘要】：为探究信号分子4或5磷酸鸟苷[(p)ppGpp]对胸膜肺炎放线杆菌(APP)适应性及致病性影响,本研究以APP血清7型S8为亲本株,通过同源重组、卡那抗性及蔗糖负向筛选的方法,将(p)ppGpp的合成调控基因relA敲除,构建缺失突变株S8△relA。生物学特性鉴定结果表明,S8△relA具有良好的遗传稳定性,与亲本株增殖速度变化差异不显著,但平台期营养限制环境下,S8△relA存活能力降低,而且在Gly、Ile、Val营养缺失时,S8△rel生长能力显著抑制;S8△relA的持留菌形成能力、生物膜形成能力以及体外抗肺泡巨噬细胞(PAM)吞噬能力均有不同程度降低;对小鼠致病性试验结果显示,S8△relA相比亲本株S8毒力降低2.7倍。研究结果表明(p)ppGpp在环境胁迫下参与APP适应性调控,并对其毒力有一定的影响。本研究首次证明了(p)ppGpp作为一种信号分子,同样可以调控APP的生长,并且在宿主肺部弱营养环境条件下,APP与致病力相关的表型均受到影响,这为进一步阐明APP的致病机制提供实验依据。
[Abstract]:In order to investigate the effects of signal molecule 4 or 5 guanosine phosphate on the adaptability and pathogenicity of Actinobacillus pleuropneumoniae (APP), the APP serotype 7 S8 was used as parent strain, which was screened by homologous recombination, kana-resistance and sucrose negative screening. RelA knockout was used to construct the deletion mutant S8 rela. The results of biological identification showed that S8 relA had good genetic stability, and there was no significant difference between S8 relA and its parents, but the survival ability of S8 relA was decreased under the condition of platform nutrition restriction. In addition, the growth ability of S8 rel significantly inhibited the retention of S8 relA, the ability of biofilm formation and the ability of anti-PAM phagocytosis of alveolar macrophages in vitro were decreased in different degree. The pathogenicity test of mice showed that the virulence of S8 relA was 2. 7 times lower than that of parent strain S8. The results showed that PppGPP participated in the adaptive regulation of APP under environmental stress and had a certain effect on its virulence. This study demonstrated for the first time that PPP, as a signaling molecule, can also regulate the growth of APP, and that the phenotypes of app associated with pathogenicity are affected under the condition of weak nutritional environment of the host lung. This provides experimental basis for further elucidating the pathogenesis of APP.
【作者单位】：东北林业大学野生动物资源学院;中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/动物细菌病创新团队;黑龙江八一农垦大学动物科技学院;
【基金】：国家自然科学基金青年项目(31302091) 公益性行业(农业)科研专项(201303034)
【分类号】：S852.61

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