Establishment of a Dental Unit Biofilm Model Using Well-Plate

The water discharged from dental unit waterlines (DUWLs) is heavily contaminated with bacteria. The development of efficient disinfectants is required to maintain good quality DUWL water. The purpose of this study was to establish a DUWL biofilm model using well-plates to confirm the effectiveness of disinfectants in the laboratory. Bacteria were obtained from the water discharged from DUWLs and incubated in R2A liquid medium for 10 days. The bacterial solution cultured for 10 days was made into stock and these stocks were incubated in R2A broth and batch mode for 5 days. Batch-cultured bacterial culture solution and polyurethane tubing sections were incubated in 12-well plates for 4 days. Biofilm accumulation was confirmed through plating on R2A solid medium. In addition, the thickness of the biofilm and the shape and distribution of the constituent bacteria were confirmed using confocal laser microscopy and scanning electron microscopy. The average accumulation of the cultured biofilm over 4 days amounted to 1.15×107 CFU/cm2. The biofilm was widely distributed on the inner surface of the polyurethane tubing and consisted of cocci, short-length rods and medium-length rods. The biofilm thickness ranged from 2 μm to 7 μm. The DUWL biofilm model produced in this study can be used to develop disinfectants and study DUWL biofilm-forming bacteria.