SYMBIOTIC INTERACTION BETWEEN BACILLUS SUBTILIS AND STREPTOCOCCUS MUTANS: DUAL SPECIES BIOFILM FORMATION FACILITATED BY CELL WALL ASSOCIATED PROTEIN TASA

Bacterial interactions often result in formation of complex structural communities termed biofilms. Biofilm in most microbial ecosystems contains large number of genetically distinct microorganisms, thus represents multispecies system. Up until now, Bacillus subtilis and Streptococcus mutans have been both extensively studied individually as model bacteria for robust biofilm formation. Whereas, the aim of this research is to investigate their interrelationship during dual species biofilm formation.

Single cell force spectroscopy using atomic force microscopy was performed to measure the initial interaction between B. subtilis cells and S. mutans biofilm. B. subtilis cells showed 5-fold higher adhesion force to S. mutans biofilm compared to glass or B. subtilis bundle. Since TasA protein is a cell wall associated protein which play crucial role in B. subtilis biofilm formation, we examined its possible role in the attachment of B. subtilis cells in the dual species biofilm. As opposed to the WT cells of B. subtilis, ΔtasA cells showed very weak interaction with S. mutans biofilm. Furthermore, our results were confirmed by CLSM visualization. B. subtilis cells appeared in deeper layers of the biofilm surrounded by the EPS associated with S. mutans` dextran. The amount of ΔtasA mutant cells was notably reduced in the dual species biofilm compared to WT B. subtilis cells. Also, visualization of the mono and dual species biofilms using SEM showed that the dual biofilm has more developed and complex structures than the mono-species biofilm. Moreover, our observation indicates about tight interaction between rod shape cells of B. subtilis and the cocci shape cells of S. mutans attached to each other and surrounded by EPS.

Acknowledgments: This work was partially supported by the COST ACTION FA1202 BacFoodNet and by the Israel Dairy Board grant 421-0270-16. Danielle Duanis-Assaf is a recipient of Scholarship of Excellency for outstanding Ph.D. students from The ARO.