Monitoring the live to dead transition of bacteria during thermal stress by a multi-method approach

Rapid microbiological methods to assess cell physiological properties of microorganisms are gaining interest in the elucidation of the effect of antimicrobial agents or physical inactivation. Fluorescent probes combined with flow cytometry or microplate assays provide information about cellular targets of chemical or physical stressors and help to clarify the underlying mode of action. In this work we exemplarily monitored the bacterial response of Listeria innocua, Staphylococcus aureus, Salmonella enterica and Escherichia coli to a mild thermal treatment by applying various methods to illustrate bacterial vital functions like the redox activity, membrane potential, esterase activity, efflux activity, glucose uptake, membrane integrity and plate counts. It was observed that some cellular properties are affected earlier than others. Respiration, glucose-uptake and pump activity were the most sensitive parameters, followed by the loss of membrane potential and membrane integrity. Unspecific esterase was found to be relatively resistant to mild heat exposure. This study shows that such a multi-method approach is a suitable tool to monitor the impact of inactivation treatments on bacteria, providing information about the mode of action, the heterogeneity of populations, species-specific differences to stressors and valuable insight in vital functions beyond pure culturability.