Determination of the amount of bacteria extruded apically during the shaping of root canals containing different biofilms

Objective: The aim of this study was to quantitatively evaluate apical microbial extrusion during root canal shaping in dentinal tubules colonized by Enterococcus faecalis( E. faecalis), Candida albicans (C. albicans), and a mixed biofilm formed by both microorganisms. Method: In this study, 54 extracted human mandibular premolars were used. After being sterilized in an autoclave, the teeth were contaminated with Enterococcus faecalis, Candida albicans, or a mixed culture containing both microorganisms. Following a seven day incubation period to allow biofilm formation, a subset of randomly selected teeth from each group was allocated as the control group, in which no root canal shaping was performed. The remaining specimens constituted the experimental group and underwent mechanical root canal shaping. Subsequently, all samples were irrigated with sterile distilled water, and the apically extruded fluid was collected for microbial culturing and CFU enumeration. In half of the specimens, OD (optical density) measurement was performed, while in the other half, biofilm morphology was evaluated using light microscopy following crystal violet staining. Results: In both the E. faecalis and mixed biofilm groups, significantly higher colony counts were observed compared to the C. albicans group (P < 0.05). When control and experimental groups were compared, all microbial species exhibited a statistically significant reduction in colony counts in the experimental group. Conclusion: Mechanical instrumentation was found to be effective in biofilm removal, with E. faecalis maintaining its dominance in both single and mixed biofilm models, while C. albicans exhibited lower growth. These findings indicate that further studies using multi-species biofilm models are essential to better understand resistance mechanisms in endodontic treatment. ii Keywords: biofilm, bacteria, dentinal tubule, apical extrusion, E. faecalis, C. albicans, mix biofilm