Oral Biofilm Architecture on Natural Teeth

Periodontitis and caries are infectious diseases of the oral cavity in which oral biofilms play a causative role. Moreover, oral biofilms are widely studied as model systems for bacterial adhesion, biofilm development, and biofilm resistance to antibiotics, due to their widespread presence and accessibility. Despite descriptions of initial plaque formation on the tooth surface, studies on mature plaque and plaque structure below the gum are limited to landmark studies from the 1970s, without appreciating the breadth of microbial diversity in the plaque. We used fluorescent in situ hybridization to localize in vivo the most abundant species from different phyla and species associated with periodontitis on seven embedded teeth obtained from four different subjects. The data showed convincingly the dominance of Actinomyces sp., Tannerella forsythia, Fusobacterium nucleatum, Spirochaetes, and Synergistetes in subgingival plaque. The latter proved to be new with a possibly important role in host-pathogen interaction due to its localization in close proximity to immune cells. The present study identified for the first time in vivo that Lactobacillus sp. are the central cells of bacterial aggregates in subgingival plaque, and that Streptococcus sp. and the yeast Candida albicans form corncob structures in supragingival plaque. Finally, periodontal pathogens colonize already formed biofilms and form microcolonies therein. These in vivo observations on oral biofilms provide a clear vision on biofilm architecture and the spatial distribution of predominant species

Chlorhexidine Substantivity on Salivary Flora and Plaque-Like Biofilm: An In Situ Model

This work was supported by project FIS PI11/01383 from Carlos III Institute of Health (Ministry of Economy and Competitiveness, Madrid, Spain

Polyethyleneimine nanoparticles incorporated into resin composite cause cell death and trigger biofilm stress in vivo

Incorporation of cross-linked quaternary ammonium polyethylenimine (QPEI) nanoparticles in dental resin composite has a long-lasting and wide antimicrobial effect with no measured impact on biocompatibility in vitro. We hypothesized that QPEI nanoparticles incorporated into a resin composite have a potent antibacterial effect in vivo and that this stress condition triggers a suicide module in the bacterial biofilm. Ten volunteers wore a removable acrylic appliance, in which two control resin composite specimens and two resin composite specimens incorporating 1% wt/wt QPEI nanoparticles were inserted to allow the buildup of intraoral biofilms. After 4 h, the specimens were removed and tested for bacterial vitality and biofilm thickness, using confocal laser scanning microscopy. The vitality rate in specimens incorporating QPEI was reduced by > 50% (p < 0.00001), whereas biofilm thickness was increased (p < 0.05). The ability of the biofilm supernatant to restore bacterial death was tested in vitro. The in vitro tests showed a 70% decrease in viable bacteria (p < 0.05). Biofilm morphological differences were also observed in the scanning electron microscope micrographs of the resin composite versus the resin composite incorporating QPEI. These results strongly suggest that QPEI nanoparticles incorporated at a low concentration in resin composite exert a significant in vivo antibiofilm activity and exhibit a potent broad spectrum antibacterial activity against salivary bacteria

Effect of fluoride and chlorhexidine digluconate mouthrinses on plaque biofilms

Objective. To develop a model in which to investigate the architecture of plaque biofilms formed on enamel surfaces in vivo and to compare the effects of anti-microbial agents of relevance for caries on biofilm vitality. Materials and Methodology : Enamel discs mounted on healing abutments in the pre-molar region were worn by three subjects for 7 days. Control discs were removed before subjects rinsed with 0.1% chlorhexidine digluconate (CHX) or 0.2% sodium fluoride (NaF) for 1 minute. Biofilms were stained with Baclight Live/Dead and z-stacks of images created using confocal scanning laser micoscopy. The levels of vital and dead/damaged bacteria in the biofilms, assessed as the proportion of green and red pixels respectively, were analysed using ImageTrak(®) software. Results : The subjects showed individual differences in biofilm architecture. The thickness of the biofilms varied from 28-96µm although cell density was always the greatest in the middle layers. In control biofilms, the overall levels of vitality were high (71-98%) especially in the area closest to the enamel interface. Rinsing with either CHX or NaF caused a similar reduction in overall vitality. CHX exerted an effect throughout the biofilm, particularly on the surface of cell clusters whereas NaF caused cell damage/death mainly in the middle to lower biofilm layers. Conclusion : We describe a model that allows the formation of mature, undisturbed oral biofilms on human enamel surfaces in vivo and show that CHX and NaF have a similar effect on overall vitality but differ in their sites of action