Structural organization and dynamics of exopolysaccharide matrix and microcolonies formation byStreptococcus mutansin biofilms

Abstract

Aims: To investigate the structural organization and dynamics of exopolysaccharides (EPS) matrix and microcolonies formation by Streptococcus mutans during the biofilm development process. Methods and Results: Biofilms of Strep. mutans were formed on saliva‐coated hydroxyapatite (sHA) discs in the presence of glucose or sucrose (alone or mixed with starch). At specific time points, biofilms were subjected to confocal fluorescence imaging and computational analysis. EPS matrix was steadily formed on sHA surface in the presence of sucrose during the first 8 h followed by a threefold biomass increase between 8 and 30 h of biofilm development. The initial formation and further development of three‐dimensional microcolony structure occurred concomitantly with EPS matrix synthesis. Tridimensional renderings showed EPS closely associated with microcolonies throughout the biofilm development process forming four distinct domains (i) between sHA surface and microcolonies, (ii) within, (iii) covering and (iv) filling the spaces between microcolonies. The combination of starch and sucrose resulted in rapid formation of elevated amounts of EPS matrix and faster assembly of microcolonies by Strep. mutans, which altered their structural organization and susceptibility of the biofilm to acid killing (vs sucrose‐grown biofilms; P < 0·05). Conclusions: Our data indicate that EPS modulate the development, sequence of assembly and spatial distribution of microcolonies by Strep. mutans. Significance and Impact of the Study: Simultaneous visualization and analysis of EPS matrix and microcolonies provide a more precise examination of the structural organization of biofilms than labelling bacteria alone, which could be a useful approach to elucidate the exact mechanisms by which Strep. mutans influences oral biofilm formation and possibly identify novel targets for effective antibiofilm therapies.

Extracellular Polysaccharides Matrix — An Often Forgotten Virulence Factor in Oral Biofilm Research

“…The production of exopolysaccharides by Gtfs acting in concert at different sites is a highly efficient mechanism by which S. mutans colonize the surface and maintain the microcolony structure, allowing them to persist on the tooth surface for prolonged period. This EPS-dependent mechanism could explain why S. mutans, either alone or mixed with other species, can only build up microcolonies when sucrose is available (Kreth et al, 2008;Xiao and Koo, 2009); strongly suggesting that sucrose may increase the competitiveness of this bacterium in a mixed species environment. Considering that bacterial fitness in biofilms may be linked with microcolonies formation, EPS may modulate the colonization and survival of S. mutans within complex biofilms in a context not previously considered.…”

“…The elevated amounts of EPS, which may involve up-regulation of gtf genes in response to pH and carbohydrate availability (Li and Burne, 2001), increase the accumulation of bacterial cells on the surface and the bulk and stability of the biofilm, enhancing its virulence (Bowen, 2002). Recently, we examined the structural relationships between the bacterial cells and EPS using a novel fluorescence imaging technique (Klein et al, 2009;Xiao and Koo, 2009), which allows for visualization and quantification of exopolymers and bacterial cells simultaneously within intact biofilms (see Figure 1). The rendered images showed EPS closely associated with bacterial cells and microcolonies throughout the biofilm development process (1) serving as a matrix holding bacterial cells on the surface allowing for the initial clusterization, and further development into microcolonies, and (2) providing supporting frame for continuous growth of the microcolonies (by enmeshing and filling the spaces between bacterial cells).…”

S. mutans biofilm model to evaluate antimicrobial substances and enamel demineralization

“…4,5 S. mutans are considered the most cariogenic microorganisms in dental biofilm due to their capacity to use dietary carbohydrates such as sucrose, to synthesize extracellular polysaccharides (EPS) and because of their acidogenic and aciduric properties. 4 EPS are important virulence factors of S. mutans because they promote bacterial adherence to the tooth surface, 6,7 contribute to the structural integrity of dental biofilms, 8,9 change the porosity of the biofilm 10 and consequently increase enamel demineralization. 11 Therefore, S. mutans biofilms have been used to evaluate their cariogenic properties due to difficulties of developing in vivo studies in controlled cariogenic situations.…”

Influence of a polymerizable eugenol derivative on the antibacterial activity and wettability of a resin composite for intracanal post cementation and core build-up restoration

“…S.Mutans virulence factors include its ability to synthesise adhesive glucans [55] and generate acids that result in the demineralisation of dental tissues, thereby initiating dental caries. Studies in literature reported that eugenol can effectively suppress the virulence of S. mutans in vitro by reducing the total mass of microorganisms and by virtue of its anti-adherence against this bacterium [56].…”

“…Studies in literature reported that eugenol can effectively suppress the virulence of S. mutans in vitro by reducing the total mass of microorganisms and by virtue of its anti-adherence against this bacterium [56]. Eugenol at sub-MIC concentrations has the ability to inhibit the formation of adhesive glucans synthesised by glucosyltransferases (GTFs) that provide specific binding sites for bacterial colonisation on the tooth surface and binding to each other [55].…”