Viability of Listeria monocytogenes and Salmonella Typhimurium after isochoric freezing

Isochoric freezing, different from isobaric (conventional) freezing, allows for storage below freezing temperatures without significant damage from ice formation. While several types of tissues have been successfully stored in sub-zero isochoric conditions, it is unknown how isochoric freezing affects pathogenic microorganisms. Thus, the objective of this study was to investigate the survival of Salmonella Typhimurium and Listeria monocytogenes at below freezing storage (<0°C) in isochoric conditions. Tested conditions included storage at ?4, ?7, and ?15°C for 24 hr and at ?15°C for 1, 2, 3, 6, 12, and 24 hr. A comparison of bacterial survival during isobaric freezing was included with every trial. Additionally, bacterial cells were examined for morphological damage using transmission electron and field-emission scanning electron microscopes. Isochoric freezing at ?15°C for 24 hr reduced both species of bacteria down to unrecoverable levels and maximum efficacy achieved after the 6 hr timepoint for L. monocytogenes and the 12 hr timepoint for S. Typhimurium. When viewed using electron microscopy, S. Typhimurium cells were noticeably disfigured with regions of cytosol separated from the cell wall. The results of this study demonstrate that isochoric freezing is capable of substantial levels of pathogen reduction. Unlike conventional nonthermal interventions, isochoric freezing does not require additional devices such as elevated pressure machines or pulsed electric fields and can be achieved with simple, inexpensive, rigid closed volume containers such as household freezers or commercial cold storage facilities.