毒力基因调控蛋白PrfA促进单核细胞增生李斯特菌生物被膜的形成

The virulence regulator PrfA plays a significant role in the Listeria monocytogenes biofilm formation

摘要:

单核细胞增生李斯特菌(Listeria monocytogenes, LM)是重要的革兰氏阳性食源性致病菌, 易在食品以及各种食品加工、运输和保藏设备的接触面形成生物被膜, 从而具有更强的抗逆性而难以彻底清除, 因此成为食品卫生安全的重要隐患。PrfA是LM毒力基因转录表达的重要调控因子, 通过比较研究LM野生株(EGD和EGDe)、PrfA缺失株(EGD?prfA和EGDe?prfA)、无害李斯特菌(Listeria innocua, LI)、携带组成性表达PrfA蛋白的重组无害李斯特菌(LI-pERL3-prfA*)以及重组单核细胞增生李斯特菌(EGDe?prfA-pERL3-prfA*)生物被膜形成能力的差异, 探讨LM重要的毒力调控蛋白PrfA对生物被膜形成的影响。实验结果显示: LM野生株具有较强的生物被膜形成能力, 而LI形成生物被膜的能力最弱; PrfA的缺失能降低LM生物被膜的形成能力; 组成性高量表达PrfA蛋白可以回复EGDe?prfA的生物被膜形成能力, 但对LI没有增强作用。以上实验结果表明: PrfA在LM生物被膜形成中具有重要的促进作用。

Abstract:

The ability of the foodborne pathogen Listeria monocytogenes to develop biofilm in food-processing environment is a major concern for the food safety, because formation of biofilm facilitates bacteria to survive in the adverse environment and resist desiccation, UV light and treatment with antimicrobial and sanitizing agents. However, the molecular mechanism of biofilm formation in L. monocytogenes has not been fully understood. PrfA is a key transcriptional activator that positively regulates most of the known listerial virulence genes expression. In order to explore the role of PrfA on Listeria biofilm development, we compared the abilities of biofilm formation in this study for L. monocytogenes wild type strains (EGD and EGDe) and their prfA deletion mutants (EGD??prfA and EGDe??prfA), nonpathogenic Listeria innocua, as well as the recombinant strains that can constitutively express PrfA in L. innocua (LI-pERL3-prfA*) and in EGDe??prfA (EGDe?prfA-pERL3-prfA*). Our results showed that the wild types of L. monocytogenes had strong abilities to develop “a network of knitted chains” biofilm structures on polyvinyl chloride microtiter plates, while unstructured biofilm was observed in L. innocua. Biofilm formation was reduced in L. monocytogenes mutants lacking PrfA and rescued in the strain with constitutive expression of PrfA. However, PrfA had no impact on L. innocua biofilm formation. Our results suggest that PrfA plays a significant role only in the L. monocytogenes biofilm formation but not in L. innocua. PrfA might indirectly regulate expression of certain genes involving in L. monocytogenes biofilm formation.