DEPTH OF PENETRATION AND BIOVOLUME OF ENTEROCOCCUS FAECALIS AND FUSOBACTERIUM NUCLEATUM BIOFILM INTO DENTINAL TUBULES USING CONFOCAL MICROSCOPE AND CO-AGGREGATION ASSAY

Hannah Rosaline*, Kandaswamy Deivanayagam, J. Kalyani and Benedict Paul

ABSTRACT

Introduction: Formation of biofilms in dentinal tubules is the major cause for the bacterial survival and virulence. The purpose of the study was to evaluate the depth of penetration and a biovolume of a mono and dual species of gram positive facultative anaerobe Enterococcus. faecalis and gram negative obligate anaerobic Fusobacterium. nucleatum into the dentinal tubules under Confocal microscope and coaggregation of bacteria using visual coaggregation assay. Methods: Root segment with a length of about 7 to 8 mm were prepared from Forty five caries-free single rooted lower premolar. Root canal was enlarged to a size of a Gates Glidden bur #2 (70mm).Irrigation protocol followed. The teeth were washed and was split in to two half Segments. The specimens were sterilized by autoclave for 20 minutes at 121°C. The strains F. nucleatum American Type Culture Collection (ATCC) 25586 and E.Faecalis ATCC 29212 were anaerobically incubated to an optical density at 600 nm (OD600) of 0.2 with 2 × 107 colony-forming units (CFU) per milliliter. Bacterial suspension was inoculated as mono and dual species into the root canal for 21 days with sterile thioglycolate medium changed every 48 hours in an anaerobic condition. Depth of penetration and biovolume of mono(control group A and group B –fusobacterium nucleatum) and dual species biofilm (group C) into dentinal tubules was evaluated. Visual Coaggregation assays were performed and a coaggregation score was given and confirmed with a bacterial smear. Results: The depth of penetration and biovolume of dual species biofilm into the dentinal tubules was statistically higher than monospecies biofilm((p <0.1). Coaggregation assay showed maximum coaggregation (+4) of E. faecalis and F. nucleatum. Conclusion: Dual species biofilm penetration and biovolume is dentinal tubules can be one of the major factors in failure of root canal treatment.

Keywords: Enterococcus. Faecalis, Fusobacterium. Nucleatum.