Streptococcus mutans adhesion force sensing in multi-species oral biofilms

Abstract

Bacteria utilize chemical and mechanical mechanisms to sense their environment, to survive hostile conditions. In mechanical sensing, intra-bilayer pressure profiles change due to deformation induced by the adhesion forces bacteria experience on a surface. Emergent properties in mono-species Streptococcus mutans biofilms, such as extracellular matrix production, depend on the adhesion forces that streptococci sense. Here we determined whether and how salivary-conditioning film (SCF) adsorption and the multi-species nature of oral biofilm influence adhesion force sensing and associated gene expression by S. mutans. Hereto, Streptococcus oralis, Actinomyces naeslundii, and S. mutans were grown together on different surfaces in the absence and presence of an adsorbed SCF. Atomic force microscopy and RT-qPCR were used to measure S. mutans adhesion forces and gene expressions. Upon SCF adsorption, stationary adhesion forces decreased on a hydrophobic and increased on a hydrophilic surface to around 8 nN. Optical coherence tomography showed that triple-species biofilms on SCF-coated surfaces with dead S. oralis adhered weakly and often detached as a contiguous sheet. Concurrently, S. mutans displayed no differential adhesion force sensing on SCF-coated surfaces in the triple-species biofilms with dead S. oralis, but once live S. oralis were present S. mutans adhesion force sensing and gene expression ranked similar as on surfaces in the absence of an adsorbed SCF. Concluding, live S. oralis may enzymatically degrade SCF components to facilitate direct contact of biofilm inhabitants with surfaces and allow S. mutans adhesion force sensing of underlying surfaces to define its appropriate adaptive response. This represents a new function of initial colonizers in multi-species oral biofilms.

Fig. 1. Examples of OCT images of S. mutans UA159 mono-species biofilm and different triple-species biofilms.

The triple-species biofilms included live or dead S. oralis J22 and A. naeslundii T14V-J1 in addition to live S. mutans UA159 after 5 h and 24 h growth on different substratum surfaces. OCT images of S. mutans mono-species biofilms on glass and silicone rubber surfaces in the absence of an adsorbed salivary-conditioning film (SCF) are taken from Wang et al.. Scale bars indicate 50 µm.

Fig. 2. The percentage composition of different triple-species biofilms containing S. mutans UA159, in combination with live or dead S. oralis J22 and A. naeslundii T14V-J1.

The percentage composition was expressed relative to the total number of CFU/cm² in a biofilm. Error bars represent SDs over triplicate experiments with separate bacterial cultures.

Fig. 3. Gene expression in S. mutans UA159 mono-species biofilms and different triple-species oral biofilms.

Different triple-species biofilms were comprised of S. mutans UA159 (S. m.) in combination with live or dead S. oralis J22 (S. o.) and A. naeslundii T14V-J1 (A. n.) in the absence and presence of an adsorbed salivary-conditioning film on different substratum surfaces. a brpA gene expression. b gbpB gene expression. c comDE gene expression. Error bars represent SDs over triplicate experiments with separately grown biofilms. Data on brpA, gbpB, and comDE expressions in S. mutans UA159 mono-species biofilms on glass and silicone rubber surfaces in the absence of an adsorbed salivary-conditioning film were taken from Wang et al..