Monitoring analysis of VBNC state of Escherichia coli O157 in soil

土壤中大肠杆菌O157 VBNC状态监测分析

• 批准号: 16510018
• 负责人: KASAHARA Yasuhiro
• 金额: $ 2.5万
• 依托单位: HOKKAIDO UNIVERSITY (2005)Ibaraki University (2004)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2004
• 资助国家: 日本
• 起止时间: 2004 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16510018/
• 关键词: Escherichia coli; VBNC; Proteome analysis; 大腸菌O157

Many pathogenic or non pathogenic bacteria enter the "viable but nonculturable (VBNC) state". In recent years, a great amount of research has revealed what occurs in cells as they enter and exist in this state, physiologically and morphologically. An aim of this study is to identify genes involved in the VBNC state of Escherichia coli O157 by mainly proteome analysis. At the first time, the experimental condition to enter the VBNC state was constructed, that is the incubation in sterile water at 4℃. The culturable cells decreased from 10^7 cells/ml to 10^2 cells/ml during 56 days incubation, but viable cells were constant at 10^7 cells/ml. The protein profiles of cells between VBNC 0 day and 56 days incubation were compared using SDS-PAGE and two dimensional gel electrophoresis, but both profiles were not observed significant difference. From proteome analysis with LC/MS/MS, about 1000 genes were detected, but they could not be found out the relation to VBNC state. Using "KO collection", that is all genes knock-out mutants in E.coli, VBNC genes were tried to identify. For 480 out of 3800 mutant strains, their VBNC states were examined. The genes have not been detected yet.

许多致病性或非致病性细菌进入“活的但不可培养（VBNC）状态”。近年来，大量的研究揭示了细胞在进入和存在于这种状态时在生理和形态上发生了什么。本研究的目的是通过蛋白质组学分析来鉴定大肠杆菌O 157 VBNC状态相关基因。首次建立了进入VBNC状态的实验条件，即在4℃无菌水中孵育。在56天的孵育期间，可培养细胞从10^7个细胞/ml降至10^2个细胞/ml，但活细胞恒定在10^7个细胞/ml。用SDS-PAGE和双向凝胶电泳比较VBNC培养0天和56天的细胞蛋白质谱，但两种蛋白质谱无显著差异。通过LC/MS/MS蛋白质组分析，共检测到约1000个基因，但未发现与VBNC状态相关的基因。利用“KO收集”即大肠杆菌中所有基因的敲除突变体，尝试鉴定VBNC基因。对于3800个突变菌株中的480个，检查了它们的VBNC状态。基因还没有被发现。