Mukamolova G V, Yanopolskaya N D, Kell D B, Kaprelyants A S
Bakh Institute of Biochemistry, Russian Academy of Sciences, Moscow, Russia.
Antonie Van Leeuwenhoek. 1998 Apr;73(3):237-43. doi: 10.1023/a:1000881918216.
It has been found previously that a significant number of Micrococcus luteus cells starved in a prolonged stationary phase (up to 2 months) and then held on the bench at room temperature without agitation for periods of up to a further 2-7 months can be resuscitated in liquid media which contained (statistically) no initially-viable (colony-forming) cells but which were fortified with sterile supernatant from the late logarithmic phase of batch growth. Here it was found that such resuscitation can be done only within a defined time period after taking the first sample from such cultures, necessarily involving agitation of the cells. The duration of this period depends on the age of the starved culture: cells kept on the bench for 3 months possess a 2 month period of resuscitability while cells starved for 6 months can be resuscitated only within 10 days after the beginning of sampling. It is suggested that the input of oxygen to the starved cultures while they are agitated may exert a negative influence on the cells, since cultures stored in anaerobic conditions (under nitrogen) had a more prolonged 'survival' time. The cells which experienced between 10 and 60 days of starvation on the bench could be resuscitated, although the number of resuscitable cells depended strongly on the concentration of yeast extract in the resuscitation medium. This concentration for cells stored on the bench for more than 2 months was 0.05% while '1-month-old' cells displayed a maximum resuscitability in the presence of 0.01% of yeast extract. Application of the fluorescent probe propidium iodide revealed the formation of cells with a damaged permeability barrier if resuscitation was performed by using concentrations of yeast extract of 0.1% and above. Thus the successful resuscitation of bacterial cultures under laboratory conditions may need rather strictly defined parameters if it is to be successfully performed for the majority of cells in a population.
先前已经发现，大量在延长的稳定期（长达2个月）饥饿培养的藤黄微球菌细胞，然后在室温下置于实验台上不搅拌长达2至7个月，可在液体培养基中复苏，该液体培养基（从统计学角度）最初不含活的（可形成菌落的）细胞，但添加了分批培养对数后期的无菌上清液。在此发现，这种复苏只能在从此类培养物中首次取样后的特定时间段内进行，且细胞必须搅拌。该时间段的长短取决于饥饿培养物的培养时间：在实验台上放置3个月的细胞具有2个月的复苏期，而饥饿6个月的细胞只能在取样开始后的10天内复苏。有人认为，在搅拌饥饿培养物时向其输入氧气可能会对细胞产生负面影响，因为储存在厌氧条件（氮气下）的培养物“存活”时间更长。在实验台上经历10至60天饥饿的细胞可以复苏，尽管可复苏细胞的数量在很大程度上取决于复苏培养基中酵母提取物的浓度。对于在实验台上存放超过2个月的细胞，该浓度为0.05%，而“存放1个月”的细胞在含有0.01%酵母提取物时显示出最大复苏能力。如果使用0.1%及以上浓度的酵母提取物进行复苏，荧光探针碘化丙啶的应用显示会形成具有受损渗透屏障的细胞。因此，在实验室条件下要成功复苏细菌培养物中的大多数细胞，可能需要相当严格定义的参数。
