title: Resazurin metabolism assay for root canal disinfectant evaluation on dual-species biofilms.
abstract: BACKGROUND
  Endodontic infections are caused by polymicrobial biofilms. Therefore, novel root canal disinfectants should be evaluated not only on single-species biofilms but also on dual- or mixed-species biofilms. A simple, high-throughput assay is urgently needed for this. In this study, the application of the resazurin metabolism assay was investigated for the evaluation of a root canal disinfectant on dual-species biofilms.
  
  
  METHODS
  Enterococcus faecalis with or without Streptococcus mutans in biofilms were formed in an active attachment biofilm model for 24 hours. Subsequently, the biofilms were treated with various concentrations of NaOCl for 1 minute. After resazurin metabolism by both organisms was confirmed, treatment efficacies using 0.0016% resazurin were evaluated.
  
  
  RESULTS
  During NaOCl treatments, resazurin metabolism displays a clear dose response, not only in single-species E. faecalis (or S. mutans) biofilms but also in dual-species biofilms. Notably, the assay revealed that the resistance of dual-species biofilms to NaOCl was 30-fold higher than in single-species E. faecalis biofilms. Viability counts on a selected NaOCl treatment (0.004%) confirmed this result and showed the increased resistance of E. faecalis in dual-species biofilms.
  
  
  CONCLUSIONS
  Clearly, the high-throughput and low cost resazurin metabolism assay has a great potential for testing novel root canal antimicrobial agents in mixed-species biofilms.
authors.affiliation: Department of Cariology, Endodontology & Pedodontology, Academic Centre of Dentistry Amsterdam, University of Amsterdam and VU University, Amsterdam, The Netherlands. l.jiang@acta.nl
authors.forename: Lei-Meng
authors.initials: LM
authors.lastname: Jiang
authors.forename: Michel A
authors.initials: MA
authors.lastname: Hoogenkamp
authors.forename: Lucas W M
authors.initials: LW
authors.lastname: van der Sluis
authors.forename: Paul R
authors.initials: PR
authors.lastname: Wesselink
authors.forename: Wim
authors.initials: W
authors.lastname: Crielaard
authors.forename: Dong Mei
authors.initials: DM
authors.lastname: Deng
mesh_terms: D018441:Biofilms / Q000187:drug effects* / Q000254:growth & development; D018920:Coculture Techniques; D015169:Colony Count, Microbial; D013293:Enterococcus faecalis / Q000187:drug effects / Q000378:metabolism*; D057166:High-Throughput Screening Assays / Q000379:methods*; D007202:Indicators and Reagents / Q000493:pharmacokinetics*; D056265:Microbial Interactions / Q000187:drug effects; D010078:Oxazines / Q000493:pharmacokinetics*; D012388:Root Canal Irrigants / Q000494:pharmacology; D012973:Sodium Hypochlorite / Q000494:pharmacology; D013295:Streptococcus mutans / Q000187:drug effects / Q000378:metabolism*; D014966:Xanthenes / Q000493:pharmacokinetics*
chemical_list: D007202:Indicators and Reagents; D010078:Oxazines; D012388:Root Canal Irrigants; D014966:Xanthenes; C005843:resazurin; D012973:Sodium Hypochlorite