Listeria monocytogenes Sublethal Injury and Viable-but-Nonculturable State Induced by Acidic Conditions and Disinfectants.

The dormancy continuum hypothesis states that in response to stress, cells enter different stages of dormancy ranging from unstressed living cells to cell death, in order to ensure their long-term survival under adverse conditions. Exposure of Listeria monocytogenes cells to sublethal stressors related to food processing may induce sublethal injury and the viable-but-nonculturable (VBNC) state. In this study, exposure to acetic acid (AA), hydrochloric acid (HCl), and two disinfectants, peracetic acid (PAA) and sodium hypochlorite (SH), at 20°C and 4°C was used to evaluate the potential induction of L. monocytogenes strain Scott A into different stages of dormancy. To differentiate the noninjured subpopulation from the total population, tryptic soy agar with 0.6% yeast extract (TSAYE), supplemented or not with 5% NaCl, was used. Sublethally injured and VBNC cells were detected by comparing plate counts obtained with fluorescence microscopy and by using combinations of carboxyfluorescein and propidium iodide (viable/dead cells). Induction of sublethal injury was more intense after PAA treatment. Two subpopulations were detected, with phenotypes of untreated cells and small colony variants (SCVs). SCVs appeared as smaller colonies of various sizes and were first observed after 5 min of exposure to 5 ppm PAA at 20°C. Increasing the stress intensity from 5 to 40 ppm PAA led to earlier detection of SCVs. L. monocytogenes remained culturable after exposure to 20 and 30 ppm PAA for 3 h. At 40 ppm, after 3 h of exposure, the whole population was considered nonculturable, while cells remained metabolically active. These results corroborate the induction of the VBNC state.