Modification of oral biofilms adhering to dental hard tissues could lead to new treatment approaches in cariology and periodontology. In this study the impact of DNase I and/or proteinase K on the formation of a simulated supragingival biofilm was investigated in vitro. Six-species biofilms were grown anaerobically in the presence of DNase I and proteinase K. After 64 h biofilms were either harvested and quantified by culture analysis or proceeded to staining followed by confocal laser scanning microscopy. Microbial cells were stained using DNA-dyes or fluorescent in situ hybridization. Exopolysaccharides, eDNA and exoproteins were stained with Calcofluor, anti-DNA-antibody, and SyproTM Ruby, respectively. Overall, results showed that neither DNase I nor proteinase K had an impact on total colony-forming units (CFUs) compared to the control without enzymes. However, DNase I significantly suppressed the growth of Actinomyces oris, Fusobacterium nucleatum, Streptococcus mutans, Streptococcus oralis and Candida albicans. Proteinase K treatment induced significant increase in S. mutans and S. oralis CFUs (p < 0.001), whereas C. albicans and V. dispar showed lower CFUs compared to the control. Interestingly, confocal images visualized the biofilm degradation caused by DNase I and proteinase K. Thus, enzymatic treatment should be combined with conventional antimicrobial agents aiming at both bactericidal effectiveness and biofilm dispersal.
Keywords: CLSM; DNase I; FISH; extracellular polymeric substances (EPS); proteinase K; supragingival in vitro biofilm.