The Cooperative and Interdependent Roles of GerA, GerK, and Ynd in Germination of Bacillus licheniformis Spores

When nutrients are scarce, Bacillus species form metabolically dormant and extremely resistant spores that enable survival over long periods of time under conditions not permitting growth. The presence of specific nutrients triggers spore germination through interaction with germinant receptors located in the spore's inner membrane. Bacillus licheniformis is a biotechnologically important species, but it is also associated with food spoilage and food-borne disease. The B. licheniformis ATCC 14580/DSM13 genome exhibits three gerA family operons (gerA, gerK, and ynd) encoding germinant receptors. We show that spores of B. licheniformis germinate efficiently in response to a range of different single L-amino acid germinants, in addition to a weak germination response seen with D-glucose. Mutational analyses revealed that the GerA and Ynd germination receptors function cooperatively in triggering an efficient germination response with single L-amino acid germinants, whereas the GerK germination receptor is essential for germination with D-glucose. Mutant spores expressing only GerA and GerK or only Ynd and GerK show reduced or severely impaired germination responses, respectively, with single L-amino acid germinants. Neither GerA nor Ynd could function alone in stimulating spore germination. Together, these results functionally characterize the germination receptor operons present in B. licheniformis. We demonstrate the overlapping germinant recognition patterns of the GerA and Ynd germination receptors and the cooperative functionalities between GerA, Ynd, and GerK in inducing germination. IMPORTANCE To ensure safe food production and durable foods, there is an obvious need for more knowledge on spore-forming bacteria. It is the process of spore germination that ultimately leads to food spoilage and food poisoning. Bacillus licheniformis is a biotechnologically important species that is also associated with food spoilage and food-borne disease. Despite its importance, the mechanisms of spore germination are poorly characterized in this species. This study provides novel knowledge on germination of B. licheniformis spores. We characterize the germinant recognition profiles of the three germinant receptors present in B. licheniformis spores and demonstrate that the GerA germinant receptor cooperates with the Ynd and GerK germinant receptors to enable an effective germination response to L-amino acids. We also demonstrate that GerK is required for germination in response to the single germinant glucose. This study demonstrates the complex interactions between germinant receptors necessary for efficient germination of B. licheniformis spores.