Sporulation modulates viscoelastic development through extracellular matrix restructuring in Bacillus subtilis...

Biofilm formation-the establishment of cellular communities within a self-secreted polymer matrix-is a behavior exhibited by almost all species of bacteria. Bacterial biofilms are well described as viscoelastic materials, with properties that grant physical advantages, such as elastic resilience and viscous adaptability. Additionally, many biofilm-forming species produce a subpopulation of highly resilient spores. In the model biofilm- and spore-forming species Bacillus subtilis , sporulation and matrix production are regulated by a common gene pathway, provoking the question of how sporulation influences biofilm viscoelastic properties. Investigating this point will facilitate the management of both salutary and deleterious biofilms, especially through manipulating their establishment and dispersion. Here, we investigate the interplay of sporulation and viscoelastic properties using rheological measurements of B. subtilis biofilms with varied matrix and spore production. We find that, to a significantly greater extent than matrix production, biofilm physical development is strongly altered by activation of sporulation through the spo0A regulation pathway. Spore-deficient strains initially establish physically robust biofilms with linear viscoelastic properties and recovery behavior distinct from spore-producing biofilms. However, spore-deficient biofilms fail to maintain their physical properties over time. In contrast, spore-producing wild-type and matrix-mutant biofilms exhibit more stable physical properties. Our results demonstrate that cell-level gene regulation is associated with macroscopic mechanical transitions in cellular communities, with sporulation granting not only a cellular-level survival advantage, but also greater community-level physical stability.

Biofilms are living materials with physical properties that influence their resilience. It is known that extracellular matrix composition largely determines biofilm physical properties. However, the role of cellular development pathways on biofilm extracellular matrix remains underexplored. Here, we examine the role of the cellular development pathway of sporulation-the production of resilient spores-in the development of biofilm physical properties. We find that spore-producing biofilms are initially softer and store less stress than spore-deficient biofilms. However, spore-producing biofilms maintain their physical properties over time whereas spore-deficient biofilms become softer and recover more poorly. Our results suggest sporulation benefits biofilms by maintaining their physical properties at the cost of decreased stiffness, which has important implications for their survival and persistence.