温度对副溶血弧菌T3SS中相关基因表达的影响  

Effect of Temperature on T3SS Gene Expression of Vibrio parahaemolyticus

作　　者：丁晓燕[1,2] 曲凌云[2,3] 王琛[1,2] 田欣欣[2] 孙承君[2] 王敏强[1] 

机构地区：[1]烟台大学生命科学学院,山东烟台264005 [2]国家海洋局第一海洋研究所,山东青岛266061 [3]海洋科学与技术国家实验室海洋渔业科学与食物产出过程功能实验室,山东青岛266237

出　　处：《海洋科学进展》2016年第2期250-259,共10页Advances in Marine Science

基　　金：国家科技支撑计划项目--海洋水产病害实用化检测及预警技术的建立与应用(2012BAD17B01);青岛海洋科学与技术国家实验室鳌山科技创新计划项目--近海健康养殖关键技术研究与新生产模式构建(2015ASKJ02-03);海洋公益性行业科研专项--重要鱼类增养殖区致病细菌基因芯片检测技术研究开发与示范(201105007-1);山东省自主创新及成果转化专项--重要水产养殖病害防治替代产品(2014ZZCX06205);泰山学者海外人才项目

摘　　要：III型分泌系统(type III secretion system,T3SS)是副溶血弧菌主要的毒力因子之一,能传递与致病有关的毒性因子来发挥病原菌的毒性,而温度是影响细菌基因表达等生命活动的关键生态因子。本文以ATCC17802和JA21作为实验菌株,分别提取其RNA,在不同温度和温度应激条件下,以pvuA为内参基因,以副溶血弧菌T3SS1的vcrD1、vopS、vopD1和T3SS2的vscC2β、vcrD2β、vopP2β作为目的基因,运用荧光定量PCR(RT-PCR)技术检测T3SS相关基因的表达下,用ΔΔCt法计算基因表达差异。结果表明:副溶血弧菌(Vibrio parahaemolyticus)ATCC17802菌株的T3SS1基因在16℃表达量最高,T3SS2基因在25℃表达量最高。副溶血弧菌JA21菌株的T3SS1基因和T3SS2基因均在25℃表达量最高。2株副溶血弧菌从不同的培养温度转入到应激温度37℃后,T3SS1和T3SS2基因的表达量均被显著诱导,并表现出时间依赖性,呈现先升高后降低的趋势。这为进一步探究环境因素与副溶血弧菌致病力的关系提供了科学依据。T3SS（Type III secretion system）is one of the major virulence factors of Vibrio parahaemolyticus,which could deliver pathogenic virulence factors or effectors to eukaryotic host cells,and temperature is one of the key ecological factors which could influence gene expression.We can compare the virulence of strains which in different culture temperature and stress conditionsby comparing the expression level of T3 SS genes,including vcrD1,vopS,vopD1,vscC2β,vcrD2β,vopP2β,which encode T3SS1 inner membrane protein,T3SS1 effector,T3SS1translocator,T3SS2 outer membrane protein,T3SS2 inner membrane protein and T3SS2 effector,respectively.In this paper,we extracted total RNA from two kinds of Vibrio parahaemolyticus ATCC17802 and JA21cultured under different temperature and stress condition.By means of established fluorescence quantitative RT-PCR method which using pvuA as the internal standard gene and theΔΔCt method as the calculation method,the transcription changes of the T3 SS mRNA were studied.The results showed that in V.parahaemolyticus 17802,T3SS1 exhibited highest expression at 16℃ while T3SS2 exhibited highest expression at 25℃.In V.parahaemolyticus JA21,both T3SS1 and T3SS2exhibited highest expression at 25℃.The expression level of T3 SS of the 2strains V.parahaemolyticus changed over time in response to stress condition,and were significantly increased compared to control group at 1hafter transporting from culture temperature into stress condition.After the peak point,the expression level dropped down.This research provides a scientific basis for the study of the relationship between the environmental factors and the pathogenicity of Vibrio parahaemolyticus.

关 键 词：副溶血弧菌 T3SS RT-PCR 基因表达 

分 类 号：X834[环境科学与工程—环境工程]