Dormancy in Non-Sporulating Bacteria: Its Significance for Environmental Monitoring

Abstract

In natural ecosystems, the total cell count obtained microscopically typically exceeds the viable count on non-selective media by orders of magnitude. The question therefore arises as to whether the “invisible”, apparently nonculturable cells are dead, are killed by our isolation media, or are merely in a dormant state from which we might in principle be able to resuscitate them if only we knew how. In particular the suggested “viable-but-nonculturable” (VBNC) bacteria have been invoked to explain phenomena as divergent as the epidemiology of some infections and the persistence of genetically marked organisms in the environment (e.g. the failure to isolate Vibrio cholerae and Campylobacter jejuni from clearly implicated sources or reservoirs of infection could be accounted for on the basis of their being present in a VBNC or dormant state). Application of flow cytometry may be a useful tool to visualize bacteria without their growing and to discriminate between dead and dormant bacteria. We found that dormant Micrococcus luteus can be physically separated from dead cells by flow cytometry sorter after cell staining with rhodamine 123. Resuscitation of dormant M.luteus cells in liquid medium does not proceed in the absence of a culture supernatant from batch-grown cells. This suggests that viable cells can excrete a pheromone-like substance necessary for the resuscitation of dormant cells.