Moderate High-Pressure Superdormancy in $\textit{Bacillus}$ Spores: Properties of Superdormant Spores and Proteins Potentially Influencing Moderate High-Pressure Germination

Resistant bacterial spores are a major concern in industrial decontamination processes. An approach known as pressure-mediated germination-inactivation strategy aims to artificially germinate spores by isostatic pressure to mitigate their resistance to inactivation processes. The successful implementation of such a germination-inactivation strategy relies on the germination of all spores. However, germination is heterogeneous, with some “superdormant” spores germinating extremely slowly or not at all. The present study investigated potential underlying reasons for moderate high-pressure (150 MPa; 37°C) superdormancy of Bacillus subtilis spores. The water and dipicolinic acid content of superdormant spores was compared with that of the initial dormant spore population. The results suggest that water and dipicolinic acid content are not major drivers of moderate high-pressure superdormancy. A proteomic analysis was used to identify proteins that were quantified at significantly different levels in superdormant spores. Subsequent validation of the germination capacity of deletion mutants revealed that the presence of protein YhcN is required for efficient moderate high-pressure germination and that proteins MinC, cse60, and SspK may also play a role, albeit a minor one.