Biofilms in Dental Unit Waterlines: Ultrastructural and Cytochemical Analysis

Dental unit waterlines are heavily colonized by bacteria which contaminate the water used to perform intraoral, and sometimes invasive, procedures. There is little information on the nature of the biofilm colonizing the flexible plastic tubing used to supply water to the different handpieces. We have therefore undertaken quantitative microbial analysis and ultrastructural studies of these biofilms by direct counting with the epifluorescent filter technique and by transmission and scanning electron microscopy after staining with ruthenium red and periodic acid-thiosemicarbazide-silver proteinate to visualize the composition and the distribution of exopolysaccharides and intracellular polysaccharides. The biofilm was revealed as a non-uniform structure characterized by an uneven cell distribution in an extracellular matrix. The microbial community of the biofilm was composed of metabolically active Gram-negative, short rod and coccoid bacteria that formed microcolonies embedded in a multilayer exopolysaccharides envelope. Our results indicate that even though the biomass of dental unit waterlines biofilms increases with time there is no direct correlation with the concentration of free floating bacteria in the water. It can be concluded that the biofilm structure described herein is responsible for the high level of bacteria in some medical devices. The consequences are of clinical significance as it is known that this extracellular material limits the action of biocides and host defense.