title: An in vitro biofilm model associated to dental implants: structural and quantitative analysis of in vitro biofilm formation on different dental implant surfaces.
abstract: OBJECTIVE
  The impact of implant surfaces in dental biofilm development is presently unknown. The aim of this investigation was to assess in vitro the development of a complex biofilm model on titanium and zirconium implant surfaces, and to compare it with the same biofilm formed on hydroxyapatite surface.
  
  
  METHODS
  Six standard reference strains were used to develop an in vitro biofilm over sterile titanium, zirconium and hydroxyapatite discs, coated with saliva within the wells of pre-sterilized polystyrene tissue culture plates. The selected species used represent initial (Streptococcus oralis and Actinomyces naeslundii), early (Veillonella parvula), secondary (Fusobacterium nucleatum) and late colonizers (Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans). The developed biofilms (growth time 1 to 120h) were studied with confocal laser scanning microscopy using a vital fluorescence technique and with low-temperature scanning electron microscopy. The number (colony forming units/biofilm) and kinetics of the bacteria within the biofilm were studied with quantitative PCR (qPCR). As outcome variables, the biofilm thickness, the percentage of cell vitality and the number of bacteria were compared using the analysis of variance.
  
  
  RESULTS
  The bacteria adhered and matured within the biofilm over the three surfaces with similar dynamics. Different surfaces, however, demonstrated differences both in the thickness, deposition of the extracellular polysaccharide matrix as well as in the organization of the bacterial cells.
  
  
  CONCLUSIONS
  While the formation and dynamics of an in vitro biofilm model was similar irrespective of the surface of inoculation (hydroxyapatite, titanium or zirconium), there were significant differences in regards to the biofilm thickness and three-dimensional structure.
authors.affiliation: Laboratory of Dental Research, University Complutense, Madrid, Spain.
authors.forename: M C
authors.initials: MC
authors.lastname: Sánchez
authors.affiliation: Laboratory of Dental Research, University Complutense, Madrid, Spain.
authors.forename: A
authors.initials: A
authors.lastname: Llama-Palacios
authors.affiliation: Laboratory of Dental Research, University Complutense, Madrid, Spain.
authors.forename: E
authors.initials: E
authors.lastname: Fernández
authors.affiliation: Laboratory of Dental Research, University Complutense, Madrid, Spain.
authors.forename: E
authors.initials: E
authors.lastname: Figuero
authors.affiliation: Laboratory of Dental Research, University Complutense, Madrid, Spain.
authors.forename: M J
authors.initials: MJ
authors.lastname: Marín
authors.affiliation: Dentaid SA, Barcelona, Spain.
authors.forename: R
authors.initials: R
authors.lastname: León
authors.affiliation: Dentaid SA, Barcelona, Spain.
authors.forename: V
authors.initials: V
authors.lastname: Blanc
authors.affiliation: ETEP (Etiology and Therapy of Periodontal Diseases) Research Group, University Complutense, Madrid, Spain.
authors.forename: D
authors.initials: D
authors.lastname: Herrera
authors.affiliation: ETEP (Etiology and Therapy of Periodontal Diseases) Research Group, University Complutense, Madrid, Spain. Electronic address: marianosanz@odon.ucm.es.
authors.forename: M
authors.initials: M
authors.lastname: Sanz
mesh_terms: D001419:Bacteria / Q000145:classification / Q000302:isolation & purification; D018441:Biofilms; D015921:Dental Implants / Q000382:microbiology*; D066298:In Vitro Techniques; D008855:Microscopy, Electron, Scanning; D013499:Surface Properties
chemical_list: D015921:Dental Implants
keywords: CLSM; Hydroxyapatite; LTSEM; Peri-implant biofilm; Titanium; Zirconium; qPCR