Factors affecting biofilm formation and transition of Listeria monocytogenes to the long-term-survival phase and their possible roles in persistence in food processing plants

Our earlier research showed that Listeria monocytogenes changes its cellular morphology from bacilli to cocci and increases its resistance to heat and high pressure during the transition to the long-term-survival (LTS) phase. The goal of this thesis project was to understand factors affecting biofilm formation and transition to the LTS phase and their relationships to persistence in the food processing environment. The transition to the LTS phase was significantly affected by both initial cell density and pH ( P < 0.001) with initial cell density being the main determining factor. Control of population growth/death kinetics appeared to be consistent with the Logistic Equation and under the control of L. monocytogenes , not external environmental factors (e.g., loss of nutrients). After 30-d incubation, the mean cell density was 4.3 +/- 1.1 x 10 8 CFU/ml and there was no significant difference between any of the initial cell density and pH treatment combinations (P > 0.05).;To understand the transition of L. monocytogenes to the LTS phase on a gene expression level, the transcriptomic profiles of L. monocytogenes at different growth stages in tryptic soy broth with yeast extract (TSBYE) were compared using a whole-genome DNA microarray. A total of 225 differentially expressed genes (≥ 4-fold, P < 0.05) were identified during the transition to the LTS phase. Genes related to cell envelope structure, energy metabolism and transport were most significantly upregulated in the LTS phase. The upregulation of compatible solute transporters may lead to accumulation of cellular solutes, lowering intracellular water activity and thus increasing bacterial stress resistance in the LTS phase. The downregulation of genes associated with protein synthesis may indicate a status of metabolic dormancy in the LTS phase. When cells in the LTS phase were inoculated into fresh TSBYE the transcriptomic profiles resembled those of log-phase cells (r = 0.94) and decreased baro- and thermotolerance were observed.;The LTS phase may help L. monocytogenes persist over a long period of time within harborage sites in food plants and subsequently transmit to food products during processing. Specific strains of L. monocytogenes are known to persist in food processing plants for years and cause contamination; however, there is a lack of understanding as to why specific strains persist in different processing plants that process different foods. Thus, we investigated the effects of different L. monocytogenes strains and different types of food-conditioning films (FCFs) on cell attachment, growth, and biofilm and cocci formation, which may help explain the persistence of specific strains in food processing plants. Type of FCF, strain and their interaction significantly affected cell density after 2-d incubation (P < 0.001). Meat and poultry FCFs showed significantly higher cell densities, as compared to the control without FCF (P < 0.05). All strains showed medium to very high densities on the respective foods from which they were isolated, except that the strain J1703 (isolated from turkey) showed very low cell density on Wegman's Brand turkey deli but very high densities on all other brands of turkey deli meat. Strains lacking the comK prophage showed lower cell densities than those containing the prophage on all four meat and poultry FCFs (P < 0.05). Biofilms were only formed by strains containing the comK prophage. Cocci were formed by all strains on all FCFs after 2-weeks incubation. The ability of specific strains of L. monocytogenes to form biofilms on specific FCFs and subsequently control their entry into the LTS phase may explain why specific strains persist in different food processing plants and cause contamination of foods manufactured in those plants.