Evaluation of the Antimicrobial Efficacy of Sodium Hypochlorite, Ozone, Diode Laser, and Photodynamic Therapy Against Enterococcus faecalis and Streptococcus mutans: An In-Vitro Study

Purpose: Persistent bacterial contamination of the root canal system, particularly by Enterococcus faecalis (E. faecalis) and Streptococcus mutans (S. mutans), remains a major obstacle in endodontic therapy. Sodium hypochlorite (NaOCl) is widely recognized as the gold-standard irrigant due to its broad-spectrum antimicrobial properties. However, its cytotoxicity has prompted the exploration of alternative or adjunctive disinfection methods, including ozone therapy, diode lasers, and photodynamic therapy (PDT). This study aimed to evaluate and compare the antimicrobial efficacy of NaOCl, ozone, diode laser, and PDT—used alone or in combination with NaOCl—against E. faecalis and S. mutans in vitro. Materials and Methods: Standard strains (E. faecalis ATCC 29212 and S. mutans ATCC 25175) were inoculated into 96-well microplates and exposed to the designated treatments following standardized protocols. In the combination groups, NaOCl was applied first, followed immediately by the secondary modality without rinsing. Colony-forming units (CFUs) were quantified by plating on Brain Heart Infusion agar and incubating under appropriate conditions. Statistical significance was assessed using the Kruskal–Wallis test with post-hoc pairwise comparisons (p < 0.05). Results: NaOCl, both alone and in combination, completely eradicated both bacterial species. Among the alternative methods, ozone gas and PDT statistically significantly reduced S. mutans counts but were less effective against E. faecalis. Diode laser and ozonated water exhibited minimal antimicrobial activity. No synergistic enhancement was observed in the combination groups. Conclusions: NaOCl remains the most effective agent for root canal disinfection. Although ozone and PDT showed moderate efficacy—particularly against S. mutans—they may serve as adjunctive options when NaOCl use is limited. Further research is warranted to optimize these alternative approaches for clinical implementation.