牛源性大肠杆菌O26热休克-酸应激响应

Response of Bovine-borne Escherichia coli O26 to Heat Shock-Acid Stress

为探讨热休克-酸应激对大肠杆菌O26存活及相关基因表达的影响,以本实验收集的22株牛源性O26为对象,首先进行乳酸和盐酸耐受实验,进而选取乳酸耐受性能不同的菌株混合进行热休克-酸应激存活实验,最后选取1株代表菌株采用实时聚合酶链式反应分析应激2 h和4 h时一般应激基因rpoS、酸应激基因(asr、ycfR、gadA)、热应激基因(rpoH、dnaK、clpB和groEL)的表达差异。结果表明,乳酸或盐酸处理2 h后22株O26存活菌数均显著下降(P<0.05),耐受程度呈现菌株差异,且同一菌株对乳酸、盐酸的耐受有差异。与正常胰蛋白胨大豆肉汤对照组相比,5株O26混菌酸应激和热休克-酸应激均导致存活菌数逐渐下降,而热休克-酸应激组存活菌数高于酸应激组,表明热休克发挥交叉保护效应,增强了O26抗酸能力。酸应激导致菌株G10Z1应激相关基因表达基本下调,而热休克-酸应激组与酸应激组相比上述基因的表达基本上调,表明热休克的交叉保护作用与上述基因表达水平的增加有关。提示食品实际生产加工中,当非致死性热处理与酸化联合使用时应注意交叉保护可能导致的食品安全风险增加的现象。

The aim of this study was to investigate the effects of heat shock-lactic acid stress on the survival of and the expression of related genes in Escherichia coli O26.First,22 bovine-borne O26 strains were collected and their tolerance to hydrochloric acid and lactic acid were compared.Then,strains with different tolerance to lactic acid were selected to evaluate their survival under heat shock-acid stress conditions.Finally,one representative strain was further selected to examine the differential expression of the general stress response gene rpoS,acid stress response genes(asr,ycfR,gadA),and heat stress response genes(rpoH,dnaK,clpB,and groEL)at 2 and 4 h of acid stress treatment using real-time polymerase chain reaction(real-time PCR).Results showed that the survival of all 22 strains significantly decreased(P<0.05)after treatment with lactic acid or hydrochloric acid for 2 h.Nevertheless,acid tolerance varied among strains and the tolerance of each strain to lactic and hydrochloric acid was also different.Compared with the control group cultured in normal tryptic soy broth(TSB),the survival rate of a mixed culture of five O26 strains gradually decreased in the acid stress and heat shock-acid stress groups,being higher in the latter than in the former,indicating that heat shock exerted a cross-protective effect and enhanced the acid resistance of O26.Real-time PCR detection showed that acid stress induced the down-regulation of stress response gene expression,whereas the expression of these genes increased significantly in heat shock-acid stress group compared with the acid stress group,indicating that the cross-protection of heat shock was related to the increased expression of the genes.It is suggested that when the combination of non-lethal heat treatment and acidification is applied in food processing,more attention should be paid to the potential of cross-protection to cause increased food safety risk.