Biofilm Formation of Clinically Relevant Bacteria

In these experiments, P. aeruginosa PAO1 (ATCC^® BAA-47™) and S. aureus (ATCC^® 25923) were used as reference strains. In addition, saliva from 3 volunteers without active caries or periodontal disease was used to isolate salivary bacteria. In particular, Streptococcus spp., Bacteroides fragilis, and Staphylococcus epidermidis were identified by biochemical assays (API^®/ID32, bioMérieux, Grassina, Italy) and grown at 37°C in non-selective nutrient broth (NB; Oxoid, Basingstoke, Hants, UK). The biofilm was developed, as previously described, on sterile (gamma-irradiated) specimens in a 96-well polystyrene plate with some modifications [32 (link)]. Briefly, an overnight culture of each bacterial strain or inoculum was diluted with tryptic soy broth (TSB; BD Bioscience, Milan, Italy) to an OD_{600 nm} of 0.2, and then incubated statically at 37°C in a humid atmosphere for 48–72 hours until a mature biofilm was obtained.