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The potential anti-cariogenic effect of blue light was evaluated using an oral biofilm model. Two species, Streptococcus mutans and Streptococcus sanguinis, were cultivated ex vivo on bovine enamel blocks for 24 h, either separately or mixed together, then exposed to blue light (wavelengths 400-500 nm) using 112 J/cm 2 . Twenty four or 48 h after exposure to light the biofilm structure and biomass were characterized and quantified using SEM and qPCR, respectively. Bacterial viability was analyzed by CLSM using live/dead bacterial staining. Gene expression was examined by RT-qPCR. After exposure to light, S. mutans biomass in mono-species biofilm was increased mainly by dead bacteria, relative to control. However, the bacterial biomass of S. mutans when grown in mixed biofilm and of S. sanguinis in mono-species biofilm was reduced after light exposure, with no significant change in viability when compared to control. Furthermore, when grown separately, an upregulation of gene expression related to biofilm formation of S. mutans, and downregulation of similar genes of S. sanguinis, were measured 24 h after exposure to blue light. However, in mixed biofilm, a downregulation of those genes in both species was observed, although not significant in S. mutans. In conclusion, blue light seems to effectively alter the bacterial biomass by reducing the viability and virulence characteristics in both bacterial species and may promote the anti-cariogenic balance between them, when grown in a mixed biofilm. Therefore, exposure of oral biofilm to blue light has the potential to serve as a complementary approach in preventive dentistry.

In prior studies, exposure of Streptococcus mutans in biofilm to blue light using high fluences of up to 680 J/cm(2) did not interfere with bacterial capability to reform an initial biofilm; however, a delayed antibacterial effect was observed. Our aim was to determine the sustained effecttts of blue light-emitting diode (LED) curing light on the pathogenicity of the newly formed biofilm. S. mutans were grown to form biofilm that was exposed to blue light (wavelengths, 460-480 nm) for 1, 3, and 7 min (equivalent to 37, 112, and 262 J/cm(2), respectively). Then, bacteria were suspended and allowed to regrow into new biofilms. The regrown biofilms were assessed for bacterial quantification by optical density (OD) measurement and quantitative polymerase chain reaction (qPCR), bacterial viability and extracellular polysaccharide production by fluorescent staining using confocal scanning laser microscopy, acid production by bacteria (acidogenicity), and bacterial survival at low pH (aciduricity) using qPCR. Bacterial growth in the regrown biofilms was increased when samples were previously exposed to light; however, under the confocal microscopy, a higher proportion of dead bacteria and a reduction in polysaccharide production were observed. The acidogenicity from the regrown biofilm was lowered as fluences of light increased. The aciduricity of the regrown biofilm was decreased, meaning less growth of bacteria into biofilm in low pH with increasing fluences. Blue light has sustained effects on S. mutans bacteria grown into new biofilm. Although bacterial growth in biofilm increased, bacterial viability and virulence characteristics were impaired. The cariogenic potential over time of S. mutans previously exposed to blue light when grown on tooth surfaces is yet to be determined.