title: Colonisation of gingival epithelia by subgingival biofilms in vitro: role of "red complex" bacteria.
abstract: OBJECTIVE
  Biofilm formation on tooth surface results in colonisation and invasion of the juxtaposed gingival tissue, eliciting strong inflammatory responses that lead to periodontal disease. This in vitro study investigated the colonisation of human gingival multi-layered epithelium by multi-species subgingival biofilms, and evaluated the relative effects of the "red complex" species (Porphyromonas gingivalis, Tannerella forsythia and Treponema denticola).
  
  
  METHODS
  The grown biofilm consisted of Fusobacterium nucleatum, Campylobacter rectus, Veillonella dispar, P. gingivalis, Prevotella intermedia, T. forsythia, T. denticola, Actinomyces oris, Streptococcus anginosus and Streptococcus oralis, or its variant lacking the "red complex". After 48h in co-culture with the gingival epithelia, the bacterial species in the biofilm were quantified, whereas their localisation on the cell surface was investigated by combining confocal-laser scanning microscopy (CLSM) and fluorescence in situ hybridisation (FISH), as well as by scanning electron microscopy (SEM).
  
  
  RESULTS
  Exclusion of the "red complex" quantitatively affected S. oralis, but not other species. The "red-complex" species were all able to colonise the gingival epithelial cells. A co-localisation trend was observed between P. gingivalis and T. denticola, as determined by FISH. However, in the absence of all three "red complex" bacteria from the biofilm, an immense colonisation of streptococci (potentially S. oralis) was observed on the gingival epithelia, as confirmed by both CLSM and SEM.
  
  
  CONCLUSIONS
  While the "red complex" species synergise in colonizing gingival epithelia, their absence from the biofilm enhances streptococcal colonisation. This antagonism with streptococci reveals that the "red complex" may regulate biofilm virulence, with potential implications in periodontal pathogenesis.
authors.affiliation: Oral Microbiology and Immunology, Institute of Oral Biology, Center of Dental Medicine, University of Zürich, Switzerland. Electronic address: thomas.thurnheer@zzm.uzh.ch.
authors.forename: Thomas
authors.initials: T
authors.lastname: Thurnheer
authors.affiliation: Oral Microbiology and Immunology, Institute of Oral Biology, Center of Dental Medicine, University of Zürich, Switzerland.
authors.forename: Georgios N
authors.initials: GN
authors.lastname: Belibasakis
authors.affiliation: Oral Translational Research, Institute of Oral Biology, Center of Dental Medicine, University of Zürich, Switzerland.
authors.forename: Nagihan
authors.initials: N
authors.lastname: Bostanci
mesh_terms: D000190:Actinomyces; D001438:Bacteroidaceae / Q000502:physiology*; D018441:Biofilms; D044883:Campylobacter rectus; D018920:Coculture Techniques; D004848:Epithelium / Q000382:microbiology*; D016967:Fusobacterium nucleatum; D005881:Gingiva / Q000382:microbiology*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D066298:In Vitro Techniques; D059013:Microbial Consortia; D018613:Microscopy, Confocal; D008855:Microscopy, Electron, Scanning; D016966:Porphyromonas gingivalis / Q000502:physiology*; D018713:Prevotella intermedia; D034366:Streptococcus anginosus; D019533:Streptococcus oralis; D048373:Treponema denticola / Q000502:physiology*; D014678:Veillonella; D014774:Virulence
keywords: CLSM; FISH; Oral biofilm; Periodontal disease