A new model of in vitro dentin intratubular contamination for Fusobacterium nucleatum: Validation by confocal laser scanning microscopy

Abstract

Objectives: To test and validate a new protocol for in vitro contamination of dentinal tubules using Fusobacterium nucleatum (F. nucleatum) by confocal laser scanning microscopy (CLSM), in addition to evaluating the effectiveness of conventional endodontic irrigants such as sodium hypochlorite (NaOCl) and chlorhexidine (CLX) on this biofilm. Material and methods: Thirty lower premolars were contaminated with F. nucleatum (ATCC 51190) for 7 days under anaerobic conditions using the proposed new model. The specimens were divided into a control group and experimental groups, according to the irrigants: NaOCl 2.5% and CLX 2%. Then, the samples were submitted for analysis by CLSM and the LIVE/DEAD technique to quantify bacterial viability. Data normality was verified by the Shapiro-Wilk test. Intragroup and intergroup comparisons were performed using the Kruskal-Wallis test, followed by Dunn's post-test. Results: The CLSM images obtained demonstrated the effectiveness of the proposed new contamination protocol, with a high percentage of viable bacteria in relation to the treated groups (p < 0.05). Lower viability values were observed for the 2.5% NaOCl group. Conclusion: The new contamination protocol resulted in a high and homogeneous percentage of viable bacteria in the dentinal tubules in all specimens evaluated. Both irrigating solutions proved to be effective in reducing the intratubular microbiota, especially 2.5% NaOCl.

Keywords

• Fusobacterium nucleatum
• Confocal microscopy
• Contamination
• Dentin