Environmental and physico-chemical factors induce VBNC state in Listeria monocytogenes

Investigations of bacterial survival in natural environments have indicated that some organisms lose culturability on appropriate media under certain conditions and yet still exhibit signs of metabolic activity and thus viability. This reproducible loss of culturability in many bacterial species led to the description of a "Viable But Non Culturable" (VBNC) state. The purpose of this article is to determine environmental and physico-chemical factors which induce the VBNC state in a food-borne pathogen that has become a public concern: Listeria monocytogenes. The factors, i.e. inoculum size, natural sunlight, temperature (4 degrees C or 20 degrees C), NaCl concentration (0% or 7%) and pH (5 or 6) were studied on 4 strains (LO28, ATCC 19115, Scott A, CNL 895807). The culturability of the starved cell suspension was determined in each condition tested by the spread plate count, and the cell activity was determined by the Direct Viable Count technique and CTC-DAPI double staining. A strain effect was found in different test conditions. For the LO 28 and ATCC 19115 strains, the VBNC state was very transient in certain conditions. For the other strains tested (Scott A, CNL 895807), the VBNC state was maintained throughout the observation period. In the dark, the incubation temperature was the main factor in the production of VBNC forms in L. monocytogenes. However, natural sunlight rapidly produced the VBNC state in L. monocytogenes cells in microcosm water. We conclude that because of its ubiquity and the factors studied which are met in the food industry, the presence of VBNC L. monocytogenes cells could pose a major public health problem since they cannot be detected by traditional culturing methods. Further investigations are needed to establish virulence before and after resuscitation of VBNC L. monocytogenes cells.