Quantitative Evaluation of the Inhibitory Effects of Commercially Available Probiotics on Dual-Species Biofilms in...

Objectives: To quantitatively evaluate the inhibitory effects of commercially available probiotic formulations (Probien, Enterogermina, Reflor) applied as intracanal medicaments against mature dual-species biofilms of Enterococcus faecalis (E. faecalis) and Candida albicans (C. albicans) using a qPCR-based in vitro root canal model, with calcium hydroxide included as the reference intracanal medicament for comparison. Materials and Methods: Root canal specimens containing mature dual-species biofilms were medicated with probiotic-poloxamer gel formulations (Probien, Enterogermina, or Reflor) or calcium hydroxide (reference inhibitory control); infected but untreated canals served as the non-inhibitory control, and sterile non-inoculated specimens were included to confirm procedural sterility. After a 7-day intracanal application period, microbial loads were quantified at baseline and post-treatment by qPCR, and results were expressed as delta cycle threshold (ΔCt), colony-forming equivalents (CFE/mL), and percentage reduction values. Results: A total of 78 specimens (n = 13 per group) were analyzed. No significant intergroup differences were found in E. faecalis ΔCt or reduction percentages (p > 0.05), indicating its persistence despite intracanal medication. For C. albicans, differences among groups were significant (p < 0.001). Calcium hydroxide showed the strongest antifungal effect, producing marked ΔCt and CFE reductions versus probiotic and positive control groups, whereas probiotic formulations displayed only limited antifungal activity and no measurable inhibition against E. faecalis. Conclusions: Under the conditions of this in vitro model, the tested commercially available probiotic formulations-originally developed for gastrointestinal use-did not demonstrate significant antimicrobial effects against mature E. faecalis-C. albicans biofilms. These findings should be interpreted in the context of the absence of probiotic formulations specifically designed for intracanal use and the distinct ecological characteristics of the root canal system, which represents a closed, low-oxygen environment dominated by hard-tissue surfaces. Rather than excluding the potential of probiotics in endodontics, the present results highlight the need for root canal-adapted probiotic strains and delivery strategies tailored to intracanal conditions. Clinical Relevance: This in vitro study provides experimental insight into the limitations of directly applying commercially available gastrointestinal probiotic formulations within the root canal system. The findings highlight the importance of developing root canal-specific probiotic strains and delivery strategies tailored to the unique ecological conditions of the intracanal environment, thereby informing future translational and experimental research in biological endodontics.