Evaluation of the antimicrobial activity of Lactobacillus acidophilus LA-5 against Enterococcus faecalis, Candida albicans, and Staphylococcus aureus: An in-vitro study

Aim This study aimed to evaluate whether Lactobacilli probiotics cell-free supernatant (CFS) exhibit antibacterial activity comparable to chlorhexidine against Enterococcus faecalis ( E. faecalis ), Candida albicans ( C. albicans ) and Staphylococcus aureus ( S. aureus). Materials and Methods This study followed the Preferred Reporting Items for Laboratory Studies in Endodontology PRILE 2021 guidelines and used an in vitro agar well-diffusion assay to assess antimicrobial activity. Standardized 0.5 McFarland inocula of E. faecalis , C. albicans , and S. aureus were lawn-cultured on Mueller–Hinton agar. Lactobacillus acidophilus (LA-5) was cultivated to obtain cell-free supernatant (CFS), serving as the experimental biological antimicrobial agent. CFS solutions (300–50 mg/mL) were tested alongside chlorhexidine and sterile water controls and incubated for 18–24 h at 37°C. Zones of inhibition were measured in triplicate by two calibrated observers for statistical analysis. Results Intergroup analysis showed significant differences in antimicrobial activity across all groups (P < 0.05). E. faecalis and C. albicans exhibited no inhibition with any CFS concentration, while chlorhexidine produced large zones. S. aureus showed measurable inhibition with CFS at 300 and 200 mg/mL, though chlorhexidine remained most effective. Intragroup comparisons confirmed that only S. aureus responded to Lactobacillus CFS, demonstrating selective antibacterial activity. Conclusion Lactobacillus acidophilus CFS showed concentration-dependent antimicrobial activity, with the higher doses (300 and 200 mg/mL) producing measurable inhibition exclusively against S. aureus . No inhibitory effect was observed on E. faecalis or C. albicans , for which chlorhexidine remained significantly more effective. These findings indicate that CFS has limited but selective antibacterial potential, primarily evident at higher concentrations.