低温寡营养条件下副溶血弧菌形成活的非可培养状态及其复苏研究  

STUDIES OF VIABLE BUT NONCULTURABLE VIBRIO PARAHAEMOLYTICUS AT LOW TEMPERATURE UNDER POOR NUTRITION CONDITIONS AND ITS RESUSCITATION

作　　者：杜萌[1] 陈吉祥[1] 孙丰蓉[1] 栾晓燕[1] 王丹[1] 李筠[1] 张晓华[1] 

机构地区：[1]中国海洋大学生命学院,青岛266003

出　　处：《水生生物学报》2008年第2期178-183,共6页Acta Hydrobiologica Sinica

基　　金：国家自然科学基金项目(30371108);上海市科技兴农项目(沪农科攻字(2005)第4-2号)资助

摘　　要：为研究在低温寡营养条件下副溶血弧菌(Vibrio parahaemolyticus)能否进入活的非可培养状态(VBNC),将浓度为1×1010CFU/mL的副溶血弧菌HW799在陈海水中4℃保存,每隔5天取样分别用吖啶橙染色荧光显微镜直接计数法(AODC)、活菌直接计数法(DVC)和涂布平板法(PC)测定细菌总数、活细菌数和可培养细菌数。在第30天时总细菌数基本不变,仍保持在109CFU/mL,活菌数为106CFU/mL,比总菌数低了约三个数量级,可培养细菌数为零,表明绝大部分副溶血弧菌HW799进入了VBNC状态;用扫描电镜、流式细胞仪对副溶血弧菌HW799活的非可培养状态、正常状态以及复苏后的细胞形态的研究表明进入VBNC状态后副溶血弧菌HW799形状变为球状,体积比正常状态明显变小,活细胞数也略有减少;采用在培养液中添加营养物质升温培养的方法,使VBNC状态的副溶血弧菌细胞在48h内复苏为可培养状态,复苏后的副溶血弧菌HW799与正常状态的细菌形态相似。Vibrio parahaemolyticus, a halophilic Gram negative bacterium, causes acute gastroenteritis in humans. This bacterium is a prevalent foodborne pathogen isolated from coastal water＇ and from a variety of seafood, including fish. Most clinical isolates are hemolytic on Wagatsuma agar （Kanagawa-positive, KP ＋ ） and produce a major virulence factor, thermostable direct hemolysin （TDH）. Seawater represents a nutrient-limiting environment for V. parahaemolyticus. During its persistence in seawater, V. parahaemolyt/cus is exposed to a variety of environmental stresses, including fluctuations in temperature and cold stress. Many bacteria have developed strategies for metamorphosis into more or less sophisticated survival forms in response to such harsh environmental conditions. The formation of （Viable but nonculturable） VBNC cells of bacteria has been proposed as a survival strategy to adverse conditions. The VBNC cell is a metabolically one although cells are incapable of undergoing the sustained cellular division required to form a colony on regular agar media. This state has been demonstrated in other human and fish bacterial pathogens, including Vibrio vulnificus , Salmonella enteritidis , enterotoxigenic Escherichia coli , Helicobacter pylori , Campylobacter jejuni , LegioneUa pneumophila, Shigella dysenteriae and Aeromonas hydrophila. It has been demonstrated that VBNC cells are active in metabolism and can nevertheless be able to recover from this dormant state, which becoming metabolically active and fully cuhurable. Although the loss caused by V. parahaemolyticus occurs over a long period of time in the year, V. parahaemolyticus is focusing on high temperature months, and is seldom isolated when the temperature of the water is low. A suggestion had been made that V. parahaemolyticus can survive for long period and may become nonculturable in the aquatic environment. In order to investigate the viable but noncuhurable state of Vibrio parahaemolyticus at low temperature under poor nutrition

关 键 词：副溶血弧菌 活的非可培养状态 复苏 扫描电镜 流式细胞仪 

分 类 号：Q93-335[生物学—微生物学]