Resuscitation and characterization of VNC vibrio cholerae

我们构建了一种在不同条件下不可培养的霍乱弧菌(活的但不可培养的：VNC)形式，以阐明作为感染源的病原菌在环境中的生活方式，并对其生理特性进行了分析。此外，我们还试图建立一种能持续复苏霍乱弧菌在琼脂上生长的方法。将霍乱弧菌悬浮在M9盐中并低温保存，培养约1个月后细菌进入VNC状态，发现进入VNC的条件不仅与霍乱弧菌菌株不同，而且与细菌周围的各种条件也不同。尤其是霍乱弧菌的菌体浓度和菌体生长阶段都会影响菌体的VNC时间，处于低菌体浓度和中等对数生长期的细菌迅速进入VNC状态。霍乱弧菌在含有许多杂质(如灰尘或生物)的水中比在…中存活的时间更长更多的放在干净的水里。这再次证实了霍乱弧菌在环境中附着在浮游生物上并能够长期存活的事实。我们证实了VNC霍乱弧菌是通过在氨离子存在下进行热处理而复苏形成菌落的。这种复苏方法对环境中的霍乱弧菌是有效的，因为我们可以复苏孟加拉国达卡附近采集的水样中的VNC霍乱弧菌。我们认为，这种复苏方法对霍乱弧菌以外的细菌都是有用的。然而，VNC的复苏速度并不是恒定的，我们必须探索更复杂的复苏方法。从我们获得的数据来看，很难认为VNC状态是“主动的生存机制”，相反，VNC状态似乎是由于细胞成分或功能的各种损伤的积累而导致“细胞死亡”的过程的初始阶段。我们现在正在研究这种修复机制是否可能是由热处理等刺激诱导的，并修复了VNC霍乱弧菌的损伤。较少

We constructed a nonculturable (viable but nonculturable : VNC) form of Vibrio cholerae under various conditions to clarify mode of life of pathogenic bacteria as infectious sources in the environment and analized their physiological characterizations. Furthermore, we tried to establish the method that could constantly resuscitate the VNC V.cholerae to grow on agar medium.When V.cholerae was suspended into M9 salt and was kept at low temperature, the bacteria entered into a VNC state after about 1 month incubation, It was found that the terms for entering into the VNC were different from not only strains of V.cholerae but also various conditions surrounding the bacteria. Especially, both cell concentrations of V.cholerae and stages of cell growth affected the time for VNC.The bacteria in low cell concentration and middle log phase have quickly entered into the VNC state. V.cholerae showed longer survival in the water containing many impurities such as dust or living things rather than … More in clean water. This indicated reconfirmation of the fact that V.cholerae attached to the planktons in the environment and was able to survive for long time.We confirmed that the VNC V.cholerae was resuscitate to form colonies by heat treatment in the presence of ammonium ions. This resuscitation method was effective to V.cholerae in the environment, since we could resuscitate VNC V.cholerae in the water sample collected near Dhaka in Bangladesh. We think that the resuscitation method will be useful to the bacteria other than V.cholerae. The rate of resuscitation from VNC, however, was not constant and we have to grope for the more sophisticated method for resuscitation.From our data obtained, it is difficult to consider the VNC state "active mechanism for Survival", rather the VNC state seems to be the initial stage of process to "cell death" caused by the accumulation of various damages to cell components or functions. We are now investigate the possibility that the repair mechanism is induced by the stimuli such as heat treatments and repair the damages in the VNC V.cholerae. Less