Transcriptional profiling of the L. monocytogenes PrfA regulon identifies six novel putative PrfA-regulated genes.

The transcriptional activator Positive Regulatory Factor A (PrfA) regulates expression of genes essential for virulence in Listeria monocytogenes. To define the PrfA regulon, the 10403S wildtype (WT) strain, a constitutively active prfA* mutant, and an isogenic ∆prfA mutant were grown under PrfA-inducing conditions in a medium containing glucose-1-phosphate and pre-treated with 0.2% activated charcoal. RNA-seq-generated transcript levels were compared as follows: (i) prfA* and WT; (ii) WT and ∆prfA and (iii) prfA* and ∆prfA. Significantly higher transcript levels in the induced WT or constitutively active PrfA* were identified for 18 genes and 2 ncRNAs in at least one of the three comparisons. These genes included: (i) 10/12 of the genes previously identified as directly PrfA-regulated; (ii) 2 genes previously identified as PrfA-regulated, albeit likely indirectly; and (iii) 6 genes newly identified as PrfA-regulated, including one (LMRG_0 2046) with a σA-dependent promoter and PrfA box located within an upstream open reading frame. LMRG_0 2046, which encodes a putative cyanate permease, is reported to be downregulated by a σB-dependent anti-sense RNA. This newly identified overlap between the σB and PrfA regulons highlights the complexity of regulatory networks important for fine-tuning bacterial gene expression in response to the rapidly changing environmental conditions associated with infection.

Nevertheless, She Resisted - Role of the Environment on Listeria monocytogenes Sensitivity to Nisin Treatment in a Laboratory Cheese Model.

The growth of Listeria monocytogenes on refrigerated, ready-to-eat food products is a major health and economic concern. The natural antimicrobial nisin targets the bacterial cell wall and can be used to inhibit L. monocytogenes growth on cheese. Cell wall composition and structure, and therefore the efficacy of cell wall acting control strategies, can be severely affected by environmental and stress conditions. The goal of this study was to determine the effect of a range of pH and temperatures on the efficacy of nisin against several strains of L. monocytogenes in a lab-scale, cheese model. Cheese was made with or without the addition of nisin at different pH and then inoculated with L. monocytogenes; L. monocytogenes numbers were quantified after 1, 7, and 14 days of incubation at 6, 14, or 22°C. While our data show that nisin treatment is able to reduce L. monocytogenes numbers, at least initially, growth of this pathogen can occur even in the presence of nisin, especially when cheese is stored at higher temperatures. Several environmental factors were found to affect nisin efficacy against L. monocytogenes. For example, nisin is more effective when cheese is stored at lower temperatures. Nisin is also more effective when cheese is made at higher pH (6 and 6.5), compared to cheese made at pH 5.5, and this effect is at least partially due to the activity of cell envelope modification genes dltA and mprF. Serotype was also found to affect nisin efficacy against L. monocytogenes; serotype 4b strains showed lower susceptibility to nisin treatment compared to serotype 1/2 strains. Overall, our results highlight the importance of considering environmental conditions specific to a food matrix when developing and applying nisin-based intervention strategies against L. monocytogenes.

Environmental conditions and serotype affect Listeria monocytogenes susceptibility to phage treatment in a laboratory cheese model.

Listeria monocytogenes can survive and grow in a variety of environments, including refrigeration, making it difficult to control and highlighting the importance of optimizing control strategies against this pathogen. Listeria phages are attractive biocontrol agents because phages bind to specific wall teichoic acids (WTA) on the bacterial cell wall, inhibiting pathogens without disrupting the normal microbiota or structure of the food. Common stresses found on dairy products can affect cell wall composition and structure and subsequently affect the efficiency of control strategies that target the cell wall. The goal of this study was to determine the effect of a range of pH and temperatures on the effectiveness of a commercial phage cocktail treatment against several strains of L. monocytogenes in a cheese matrix. We developed a laboratory-scale cheese model that was made at different pH, treated with phage, and then inoculated with L. monocytogenes. Cheeses were incubated at 6, 14, or 22°C for 14 d, and bacterial counts were determined on d 1, 7, and 14. Our data show that phage treatment has a limited ability to reduce L. monocytogenes counts at each temperature tested; however, it was more effective on specific strains of L. monocytogenes when cheese was stored at higher temperatures. More specifically, the average counts of L. monocytogenes on phage-treated cheese stored at 22°C were significantly lower than those on phage-treated cheese stored at 6 or 14°C. Similarly, phage treatment was significantly more effective at inhibiting L. monocytogenes on cheese made at higher pH (6 and 6.5) compared with counts on cheese made at pH 5.5, where L. monocytogenes did not grow. Furthermore, serotype was found to affect the susceptibility of L. monocytogenes to phage treatment; serotype 1/2 strains showed significantly higher susceptibility to phage treatment than serotype 4b strains. Overall, our results suggest the importance of considering the efficacy of phage under conditions (i.e., temperature and pH) specific to a given food matrix when applying interventions against this important foodborne pathogen.