IADR Abstract Archives
Fusobacterium nucleatum Counts Increase After Biofilm Treatment With DNase I
Objectives: Bacterial biofilm matrix mainly contains a scaffold of high molecular weight exopolysaccharides, which incorporates more carbohydrates, proteins, lipids, and nucleic acids. Targeting and eliminating nucleid acids or proteins could enable an efficient control of biofilms. The objective of this study was therefore to test the effect of deoxyribonuclease I (DNase I) and proteinase K on the structural stability of oral biofilms.
Methods: Six-species biofilms consisting of Streptococcus mutans, Streptococcus oralis, Actinomyces oris, Fusobacterium nucleatum, Veillonella dispar and Candida albicans were grown anaerobically on pellicle-coated hydroxyapatite disks. At specific intervals (0h, 16h, 40h), biofilms were exposed to two different concentrations of DNase I and proteinase K (0.001 mg/ml, 0.002 mg/ml) for 1h. Afterwards, biofilms were harvested and the number of colony forming units (CFU) were determined. The visualization of the treated biofilms was aided by immunofluorescence (IF) and confocal laser scanning microscopy (CLSM) using anti-DNA antibodies, streptavidin (Cy3), calcofluor, SYPRO Ruby and YoPro-1/Sytox. The results were analysed using ANOVA, followed by the Tukey test at a 5% significance level.
Results: The biofilm treatment induced a significant increase of 1.5 logs in F. nucleatum CFU counts, while the growth of all other species was not affected. No significant differences were found between different concentrations of DNase I and proteinase K independent of the time of their addition into the biofilm mass. Interestingly, the confocal images revealed a significant reduction of the biofilm thickness from 23µm (control) to 12 µm after treatment with 0.002 mg/ml DNAse I due to integrity decrease in matrix.
Conclusions: Besides favoring growth of F. nucleatum, treatment either with DNase I or proteinase K reduces matrix integrity, and thus biofilm volume.
Methods: Six-species biofilms consisting of Streptococcus mutans, Streptococcus oralis, Actinomyces oris, Fusobacterium nucleatum, Veillonella dispar and Candida albicans were grown anaerobically on pellicle-coated hydroxyapatite disks. At specific intervals (0h, 16h, 40h), biofilms were exposed to two different concentrations of DNase I and proteinase K (0.001 mg/ml, 0.002 mg/ml) for 1h. Afterwards, biofilms were harvested and the number of colony forming units (CFU) were determined. The visualization of the treated biofilms was aided by immunofluorescence (IF) and confocal laser scanning microscopy (CLSM) using anti-DNA antibodies, streptavidin (Cy3), calcofluor, SYPRO Ruby and YoPro-1/Sytox. The results were analysed using ANOVA, followed by the Tukey test at a 5% significance level.
Results: The biofilm treatment induced a significant increase of 1.5 logs in F. nucleatum CFU counts, while the growth of all other species was not affected. No significant differences were found between different concentrations of DNase I and proteinase K independent of the time of their addition into the biofilm mass. Interestingly, the confocal images revealed a significant reduction of the biofilm thickness from 23µm (control) to 12 µm after treatment with 0.002 mg/ml DNAse I due to integrity decrease in matrix.
Conclusions: Besides favoring growth of F. nucleatum, treatment either with DNase I or proteinase K reduces matrix integrity, and thus biofilm volume.