Quantification of Bacillus cereus stress responses.

The microbial stability and safety of minimally processed foods is controlled by a deliberate combination of preservation hurdles. However, this preservation strategy is challenged by the ability of spoilage bacteria and food-borne pathogens to adapt to stressing environments providing cell robustness. Bacillus cereus is a toxin-producing, spore-forming bacterium, and is able to survive minimal processing conditions. A quantitative approach was followed to gain insight in B. cereus’ stress adaptive behavior at population, individual cell and molecular level. B. cereus’ ability to adapt to salt stress and gain robustness towards subsequent heat challenge-stress exposure was quantified in detail using primary kinetics models. The adaptive salt stress response was influenced by the adaptation-stress concentration, the growth phase of the cells, strain diversity and the culturing temperature during adaptation-stress treatment. The nonlinear nature of the heat inactivation kinetics suggested heterogeneity within the population with respect to stress adaptive behavior. The direct-imaging-based Anopore technology was used to quantitatively describe the population heterogeneity of B. cereus upon mild and severe salt stress treatments and during low temperature growth. Fluorescent labeling of cells provided insights in the origin of stress-induced population heterogeneity. Then, to elucidate adaptive salt stress responses at molecular level, the genome-wide transcriptome profiles of mildly and severely salt-stressed cells were compared. Various transcriptome responses could be correlated to phenotypic features of salt stress-adapted cells. Comparison of the transcriptome profiles of salt stress-adapted cells to those that were exposed to mild heat, acid and oxidative stress, directed to potential cellular biomarkers for stress adaptation. The selected candidate-biomarkers  the transcriptional regulator B (activating general stress responses), catalases (removing reactive oxygen species), and chaperones and proteases (maintaining protein quality)  were measured upon adaptation-stress treatment at transcript, protein and/or activity level, and their induction was correlated to adaptation-stress induced robustness towards challenge-stress. Various candidate-biomarkers were suitable to predict the robustness level of adaptation-stress pretreated cells towards challenge-stress, and are therefore potential predictive cellular indicators for adaptation-stress induced robustness. The predictive potential of transcripts differed from that of proteins and activity level, underlining the significance to evaluate predictive potential of candidate-biomarkers at different functional cell levels. This quantitative understanding of B. cereus’ stress adaptive behavior provides mechanistic insights and opens up avenues to come to a mechanism-based approach for designing mild preservation strategies.