Changes of ultrastructure and stress tolerance of Vibrio parahaemolyticus upon entering viable but nonculturable state

This study examined the change of ultrastructure and stress tolerance of the marine foodborne pathogen, Vibrio parahaemolyticus 1137, when incubated under viable but nonculturable (VBNC) state induction conditions for different time intervals. The rod-shaped V. parahaemolyticus cells in the exponential phase became coccoid cells in the VBNC state, with aberrantly shaped cells formed in the initial stage. In the aberrantly shaped cells, the cell wall was loosened, flexible and allowed the cell to bulge, and the formation of new and thin cell wall or the expansion of cell wall was also discerned primarily at the polar position, enclosing an empty cellular space. The thickness of the cell wall increased with the VBNC induction time, and was increased in cultures that were removed from the induction conditions and whose temperature was upshifted to 25 °C for 1 or 2 days. The incubation of V. parahaemolyticus under the VBNC induction conditions significantly enhanced its tolerance to heat, H2O2 and low salinity, but sensitized it to bile salts. Tolerance to heat, bile salts and low salinity was significantly higher in the temperature upshifted cultures than in the corresponding unheated cultures, and the heated cultures were also more susceptible to H2O2. The V. parahaemolyticus cultures that were incubated in the VBNC state induction conditions and the corresponding temperature-upshifted cultures exhibited unique changes in ultrastructure and tolerance to various stresses, unlike the nutrient-starved cells.

► First report on the complete change of ultrastructure of cells entering VBNC state ► Distinct features suggesting the mechanism for formation of aberrantly shaped cells ► Cell wall thickening found in cells entering VBNC state/heat recovered cells ► Stress tolerance enhanced in cells entering VBNC state/heat recovered cells ► Stress tolerance in the VBNC cultures different from the response of starvation