Temperature and growth-phase effects on Aeromonas hydrophila survival in natural seawater microcosms: role of protein synthesis and nucleic acid content on viable but temporarily nonculturable response

The behaviour of Aeromonas hydrophila in nutrient-poor filter-sterilized seawater was investigated at 23 and 5 degrees C with respect to its growth phase. At both temperatures, the culturable A. hydrophila population declined below the detection level (0.1 c.f.u. ml(-1)) after 3-5 weeks, depending on the initial physiological state of the cells. During the first week, starved A. hydrophila cells appeared more resistant to the seawater stress at 5 degrees C than cells initially in the exponential growth phase. This difference was not observed at 23 degrees C, where de novo protein synthesis seemed to be required for long-term adaptation of cells from the exponential growth phase. Over the duration of the experiments, intact and total cell concentrations were not significantly affected, indicating that bacteria had entered a so-called viable but nonculturable state (VBNC). However, the incubated bacteria rapidly became heterogeneous with respect to their nucleic acid content, and their cell size decreased faster at 23 than at 5 degrees C. Resuscitation of VBNC cells was attempted by a temperature shift from 5 to 23 degrees C without exogenous nutrient addition. Comparison of the growth rates of the stressed population and of the untreated bacteria growing in the same autoclaved initial cell suspension showed significantly faster growth for the stressed cells, suggesting that in addition to growth of the few culturable stressed cells, a proportion of injured cells became culturable.