Biofilm formation by Bacillus cereus was assessed using 56 strains of B. cereus, including the two sequenced strains, ATCC 14579 and ATCC 10987. Biofilm production in microtiter plates was found to be strongly dependent on incubation time, temperature, and medium, as well as the strain used, with some strains showing biofilm formation within 24 h and subsequent dispersion within the next 24 h. A selection of strains was used for quantitative analysis of biofilm formation on stainless steel coupons. Thick biofilms of B. cereus developed at the air-liquid interface, while the amount of biofilm formed was much lower in submerged systems. This suggests that B. cereus biofilms may develop particularly in industrial storage and piping systems that are partly filled during operation or where residual liquid has remained after a production cycle. Moreover, depending on the strain and culture conditions, spores constituted up to 90% of the total biofilm counts. This indicates that B. cereus biofilms can act as a nidus for spore formation and subsequently can release their spores into food production environments.

The gene encoding the alternative sigma factor B in Listeria monocytogenes is induced upon exposure of cells to several stresses. In this study, we investigated the impact of a sigB null mutation on the survival of L. monocytogenes EGD-e at low pH, during high-hydrostatic-pressure treatment, and during freezing. The survival of ⌬sigB mutant exponential-phase cells at pH 2.5 was 10,000-fold lower than the survival of EGD-e wild-type cells. Moreover, the ⌬sigB mutant failed to show an acid tolerance response. Upon preexposure for 1 h to pH 4.5, the survival at pH 2.5 was 100,000-fold lower for the ⌬sigB mutant than for the wild type. The glutamate decarboxylase (GAD) acid resistance system is important in survival and adaptation of L. monocytogenes in acidic conditions. The B dependence of the gad genes (gadA, gadB, gadC, gadD, and gadE) was analyzed in silico. Putative B -dependent promoter sites were found upstream of the gadCB operon (encoding a glutamate/ ␥-aminobutyrate antiporter and a glutamate decarboxylase, respectively) and the lmo2434 gene (gadD, encoding a putative glutamate decarboxylase). Reverse transcriptase PCR revealed that expression of the gadCB operon and expression of gadD are indeed B dependent. In addition, a proteomics approach was used to analyze the protein expression profiles upon acid exposure. Although the GAD proteins were not recovered, nine proteins accumulated in the wild type but not in the ⌬sigB strain. These proteins included Pfk, GalE, ClpP, and Lmo1580. Exposure to pH 4.5, in order to preload cells with active B and consequently with B -dependent general stress proteins, also provided considerable protection against high-hydrostatic-pressure treatment and freezing. The combined data argue that the expression of B -dependent genes provides L. monocytogenes with nonspecific multiple-stress resistance that may be relevant for survival in the natural environment as well as during food processing.Listeria monocytogenes is a gram-positive, nonsporulating, facultatively anaerobic microorganism. It is the causative agent of listeriosis, a serious illness for which the young, elderly, and immunocompromised are especially at risk (31). L. monocytogenes is of particular concern for the food industry due to the severity of the illness, as well as the wide distribution of the pathogen in the environment and consequently its presence on raw and minimally processed foods. Moreover, L. monocytogenes is known for its ability to survive and proliferate in adverse environmental conditions, including acidic conditions, refrigeration temperatures, and high osmolarity (up to 10%) (4, 36). These characteristics of L. monocytogenes make it a food-borne pathogen that is problematic for certain kind of foods, especially minimally processed, ready-to-eat foods.Adaptation to (sudden) adverse conditions in the environment of a bacterium requires the ability to respond rapidly. Such a response of a bacterium to environmental changes involves activation of existing enzymes and enhanced rates of transcripti...

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