Viable but nonrecoverable stage of Salmonella enteritidis in aquatic systems

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Abstract
An environmental isolate (13-1BB) of S. enteritidis serogroup C1 was inoculated into sterile Potomac River [USA] water microcosms to observe survival and culturability of the organism by employing acridine orange direct count, fluorescent antibody direct count, direct viable count, plate count on veal infusion agar and xylose lysine decarboxylase agar, and indirect enumeration by the most-probable-number method (MPN), using media selective for Salmonella. Loss of culturability on laboratory media was observed within 48 h. Cultures could be resuscitated and cultured on solid media, following addition of nutrients to the microcosms. Cells, resuscitated 4 days after apparent die-off (0 colony-forming units [cfu]/ml) using plate count techniques, yielded numbers of cfu in the same order of magnitude as was observed before the onset of nutrient limitation. Microscopic techniques for direct viable counting indicated that viability is maintained for as long as 60 days after depletion of nutrients, although attempts to culture these cells, by addition of nutrient, after 21 days yielded apparently sterile plates. Longer periods of dormancy appear to require conditions other than simple nutrient addition for resumption of cell growth and division.