Figure 5

Effect of quercetin on biofilm formation. (a) P. gingivalis suspension was grown in BHI supplemented with quercetin for 48 h at 37 °C in a 96-well polystyrene plates. After washing three times with sterile PBS, the adherent biofilms were incubated with methanol for 15 min followed by staining with 0.04% (w/v) crystal violet to determine overall biomass of the biofilm at wavelength of 550 nm. (b) The biofilm was formed as described for the above method. MTT (0.5 mg/mL) was added to detect the metabolic activity of the biofilms at wavelength of 590 nm. (c–g) CLSM images of P. gingivalis biofilm treated with quercetin. P. gingivalis biofilms were formed as described above assay for 48 h on glass coverslips with different concentrations of quercetin. Bacterial cells were washed three times with saline to remove unbound cells and stained with the LIVE/DEAD BacLigh Bacterial Viability Kit containing SYTO 9 dye and propidium iodide. (c) 0 μM, (d) 12.5 μM, (e) 25 μM, (f) 50 μM, (g) 100 μM, Bar = 50 μm. (h) Biofilm thickness. Bars denoted by (**), (***) and (****) indicate significant difference at p < 0.01, p < 0.001 and p < 0.0001, respectively by one-way analysis of variance (ANOVA) with Dunnett's post hoc test.