不同温度条件下副溶血性弧菌tdh基因表达及其代谢组响应

Metabolome response to tdh expression of pathogenic Vibrio parahaemolyticus induced by different temperatures

摘要:

摘要:【目的】采用RT-PCR的方法分析致病性副溶血性弧菌毒力基因表达，并应用代谢组学的方法研究毒力基因不同表达水平下致病性副溶血性弧菌代谢组的响应。【方法】本文以致病性副溶血性弧菌(Vibrio parahaemolyticus)ATCC33846为材料，分别提取不同温度(4、25 和37℃)下菌体总RNA和代谢组。采用相对定量的方法检测副溶血性弧菌tdh基因在不同温度条件下的表达差异，同时应用超高压液相色谱-四级杆飞行时间质谱联用仪(UPLC/Q-TOF-MS)系统为工作平台检测其代谢组。采用主成分分析法(principal component analysis，PCA)比较副溶血性弧菌代谢组轮廓差异，并通过皮尔森和斯皮尔曼相关性分析法分析代谢组与tdh基因表达之间相关性。【结果】结果表明，不同温度条件下tdh基因表达强弱的排列顺序25℃＞4℃＞37℃;在tdh基因不同表达水平下发生显著性(P＜0.05)变化的主要代谢物是有机酸、氨基酸、醇、酮、酯;共得到11种代谢物与tdh基因表达高度相关(相关性系数︱r︱= 1，P＜0.05)，其中3种为负相关，8种为正相关，且醇类代谢物与tdh基因表达的正相关性最显著。【结论】本研究发现副溶血性弧菌代谢组与毒力基因表达存在一定的相关性，有望为副溶血性弧菌致病机理的深入探究提供一定的理论支持。

Abstract:

Abstract:［Objective］We studied the metabolome responses to different relative virulence gene expression of pathogenic Vibrio parahaemolyticus based on reverse transcription real-time PCR (RT-PCR) and metabolomics.［Methods］ We extracted total RNA and metabolites of V．parahaemolyticus ATCC33846 cultured at different temperatures (4，25 and 37℃)．We applied relative quantification to analyze the tdh expression and Ultra Performance Liquid Chromatography ＆ Quadrupole-Time-of-Flight Mass Spectrometry (UPLC/Q-TOF-MS) platform to determine the low-molecular-weight metabolites．Then，we compared the metabolic profiling of V．parahaemolyticus by principal components analysis．The correlation between relative gene expression and metabolome was obtained by Pearson and Spearman correlation coefficient (r) analysis.［Results］The expression level of tdh at different temperatures ranked 25℃＞4℃＞37℃．The main metabolites that changed significantly (P＜0.05) were organic acids，amino acids，alcohols，ketones，and esters．The results from correlation coefficient suggested that 11 metabolites were highly correlated with tdh expression，including 3 negative correlations and 8 positive correlations，at the threshold of |r|=1，P＜0.05．Further analysis revealed that alcohols were highly significant correlated with tdh expression．［Conclusion］ The results suggested that there were significant correlation between metabolome and virulence genes expression，which might contribute to understanding the pathogenic mechanism of V．parahaemolyticus.