Oil-in-water emulsion and acid concentrations impact Bacillus weihenstephanensis KBAB4 growth and culturability

Organic acids have long been used to limit microbial growth and preserve food products from spoilage and pathogens. However, the food complexity can affect the efficiency of preservatives in inhibiting bacterial growth. The inhibitory activity of acetic, lactic, caprylic and lauric acid in nutrient broth and in a model food system, i.e. an oil-in-water (O/W) emulsion at two pH levels, were compared using the plate count method. The fate of B. weihenstephanensis KBAB4 cells incubated in O/W emulsions revealed a loss of culturability on Brain Heart Infusion agar (BHI agar) medium, occurring at different times according to the tested conditions. Cell viability in O/W emulsion was assessed through flow cytometry measurements which confirmed that a large majority of cells possessed an intact membrane, despite only 3 % being able to recover on BHI agar. Decreasing the pH of the emulsions delayed the time at which cells lost their culturability as well as the time at which it was restored. Using acetic, caprylic and lauric acid in the emulsion impacted strain recovery on BHI agar whereas using lactic acid enhanced the cell recovery from the emulsion exposure, on BHI agar surface. The culturability of B. weihenstephanensis KBAB4 cells incubated in an O/W emulsion without acid has been restored using other recovery media, highlighting ROS involvement and limited strain catalase activity in the recovery ability. This study put in evidence the susceptibility of this strain to oxidative stress and challenges the use of given media for enumerating spore-forming bacteria in food and other complex matrices.