title: A novel approach to antibiofilm susceptibility testing using a thermo-reversible matrix.
abstract: OBJECTIVE
  Biofilm microorganisms are known to have a much higher tolerance to antimicrobials compared to their planktonic equivalents. Therefore, traditional antimicrobial susceptibility testing may not extrapolate to clinical treatment of infections of biofilm origin, and as a result, there is a need to not only develop antimicrobials with antibiofilm activity, but also suitable in vitro testing methods for their evaluation. In this study, we report on a novel method of antibiofilm testing using a thermo-reversible matrix (poloxamer 407), coupled with live/dead staining of bacteria cultured from the matrix.
  
  
  METHODS
  Pseudomonas aeruginosa (NCIMB 8626) was cultured in medium containing poloxamer 407 at 37°C for 24 hours to generate biofilms. The preparation was cooled to liquefy the poloxamer and allow recovery of the biofilm cells, which were then stained with SYTO9 to determine viability following exposure to four antimicrobials: polyhexanide, octenadine dihydrochloride, povidone-iodine and silver carbonate. Over an 8-minute time period, fluorescence levels were spectrophotometrically measured and compared with bacterial controls, cultured in the absence of poloxamer and without antimicrobial.
  
  
  RESULTS
  Untreated cells showed no reduction in viability over this period. Importantly, planktonic cells were more susceptible to test agents compared with those of a 'biofilm' phenotype cultured in poloxamer. Antibiofilm activity was evident for all of the test agents, with highest relative activity seen with octenadine dihydrochloride.
  
  
  CONCLUSIONS
  In summary, a novel and relatively rapid approach to screen compounds for antibiofilm activity has been described. The method uses standard laboratory equipment and can be readily adapted to test a wide range of microorganisms and other antibiofilm compounds.
  
  
  BACKGROUND
  This research was, in part, supported by Advanced Medical Solutions in the form of a Knowledge Transfer Project. Mr J. Nosworthy was employed by Advanced Medical Solutions. There are no other conflicts of interests to declare.
authors.affiliation: Oral and Biomedical Sciences, School of Dentistry, Cardiff University, Cardiff.
authors.forename: B J
authors.initials: BJ
authors.lastname: Taylor
authors.affiliation: Oral and Biomedical Sciences, School of Dentistry, Cardiff University, Cardiff.
authors.forename: L L
authors.initials: LL
authors.lastname: Marsh
authors.affiliation: Advanced Medical Solutions Ltd, 33 Premier Park, Winsford, Cheshire.
authors.forename: J O
authors.initials: JO
authors.lastname: Nosworthy
authors.affiliation: Oral and Biomedical Sciences, School of Dentistry, Cardiff University, Cardiff.
authors.forename: D W
authors.initials: DW
authors.lastname: Williams
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology* / Q000627:therapeutic use*; D000890:Anti-Infective Agents / Q000494:pharmacology* / Q000627:therapeutic use*; D001424:Bacterial Infections / Q000188:drug therapy*; D001645:Biguanides / Q000494:pharmacology / Q000627:therapeutic use; D018441:Biofilms / Q000187:drug effects*; D002254:Carbonates / Q000627:therapeutic use; D006801:Humans; D007097:Imines; D008826:Microbial Sensitivity Tests; D011206:Povidone-Iodine / Q000494:pharmacology / Q000627:therapeutic use; D011550:Pseudomonas aeruginosa / Q000187:drug effects*; D011725:Pyridines / Q000494:pharmacology / Q000627:therapeutic use; D018030:Silver Compounds / Q000627:therapeutic use
chemical_list: D000900:Anti-Bacterial Agents; D000890:Anti-Infective Agents; D001645:Biguanides; D002254:Carbonates; D007097:Imines; D011725:Pyridines; D018030:Silver Compounds; C031233:polihexanide; D011206:Povidone-Iodine; C034213:octenidine; C033260:silver carbonate
keywords: Pseudomonas aeruginosa; SYTO9; antibiofilm agents; biofilm modelling; poloxamer