活的非可培养状态大肠杆菌O157:H7的复苏研究

Resuscitation of Viable but Non-culturable E. coli O157:H7

为研究活的非可培养状态(viable but nonculturable state,VBNC)的Escherichia coli O157:H7的复苏能力,将E.coli O157:H7分别置于LB液体培养基和生理盐水在-18℃低温条件下诱导生产活的非可培养状态E.coli。结合吖啶橙荧光显微镜计数法、活菌直接计数法和平板计数法对样本细菌检测验证,分别在第15天和第18天得到VBNC样本。采用将培养液直接升温到37℃、加入体积分数为25%酵母浸膏(25℃)和加入体积分数为8%吐温-80(37℃)3种方法进行复苏研究。同时利用荧光定量PCR技术监测从诱导到复苏整个过程中的菌数变化。结果显示:3种方法均能使VBNC状态E.coli O157:H7在2d内复苏,复苏后菌株与正常菌株形态类似;细菌进入VBNC状态15d后,采用相同的方法则无法复苏。

In order to explore the resuscitative capability of viable but nonculturable （VBNC） Escherichia coli O157：H7, the bacteria were induced into LB broth and physiological saline at low temperatures （-18 ℃） respectively. Plate count, scanning electron microscope and qPCR technique were used to detect the VBNC cell populations on the 15th and 18th days, respectively. Resuscitation of VBNC samples within 2 d was achieved by direct heating to 37 ℃, addition of 25% yeast extract at 25℃, or addition of 8% Tween-80 at 37 ℃. The pattern of the resuscitated VBNC cells was similar to that of the normal cells. However, the bacteria could not resuscitate after the VBNC state was kept for 15 d.