副溶血性弧菌生物被膜动态形成机制的转录组分析

Transcriptomic Analysis of the Dynamic Formation Mechanism of Vibrio parahaemolyticus Biofilm

为探究副溶血性弧菌生物被膜形成机制,在菌株生理特性分析的基础上,结合转录组测序技术,对生物被膜形成过程中的基因表达调控规律进行探究。实验测定了2株被膜差异菌株(ATCC 17802和VP-0)的胞外多糖、胞外蛋白、信号分子AI-2、细胞渗透性和生物被膜微观形态等生理指标,研究被膜差异菌株的动态形成过程。结果显示:供试菌株生物被膜形成过程,0~12 h为可逆黏附阶段,12~48 h为不可逆黏附和微菌落形成阶段,48~72 h为成熟阶段,72~144 h为解离阶段。ATCC 17802胞外多糖和蛋白在生物被膜成熟期分泌量分别是VP-0的1.67倍和2.3倍,在生物被膜形成过程中信号分子含量相差不显著(P>0.05),激光共聚焦显微镜显示ATCC 17802呈现更聚集状态。根据转录组测序结果分析可知,3个处理组(72 h vs 8 h、48 h vs 8 h和72 h vs 48 h)分别鉴别出802、1061个和267个显著性差异表达基因,其中分别有506、655个和96个差异基因下调,296、406个和171个差异基因上调。这些差异主要体现在能量代谢、鞭毛系统、转运系统等与生物被膜形成有关的方面。本实验详细划分了副溶血性弧菌生物被膜的形成阶段,也为生物被膜动态调控中的基因调控过程提供了理论基础。

To investigate the mechanism of biofilm formation in Vibrio parahaemolyticus,its physiological characteristics were examined and the regulation of gene expression during biofilm formation was studied transcriptomic sequencing.Two strains of V.parahaemolyticus(ATCC 17802 and VP-0)that differ in their biofilms were evaluated for their physiological indexes such as extracellular polysaccharides,extracellular proteins,the signaling molecule autoinducer-2(AI-2),cell permeability,and biofilm microscopic morphology,and the dynamic formation process of biofilm was investigated.The results showed that the biofilm formation process of the tested strains involved reversible adhesion(from 0 to 12 h),irreversible adhesion and micro-colony formation(from 12 to 48 h),maturation(from 48 to 72 h),and dissociation(from 72 to 144 h).The secretion rates of extracellular polysaccharides and proteins at the biofilm maturation stage of ATCC 17802 were 1.67 and 2.3 times higher than those of VP-0,respectively.The difference in signaling molecule content during biofilm formation was not significant(P>0.05),and laser confocal scanning microscopy(LCSM)showed that ATCC 17802 had a greater degree of aggregation.Based on the results of transcriptomic sequencing,802,1061,and 267 significantly differentially expressed genes were identified in three treatment groups(72 vs 8 h,48 vs 8 h,and 72 vs 48 h),of which 506,655 and 96 differentially expressed genes were down-regulated and 296,406 and 171 differentially expressed genes were up-regulated,respectively.These differences were mainly involved in energy metabolism,flagellar system,transport system,and other aspects related to biofilm formation.The findings obtained in this study provide a theoretical basis for gene expression regulation during the dynamic formation of V.parahaemolyticus biofilm.