Method: The strains Enterococcus faecalis JH2-2, Streptococcus gordonii DL1 and Streptococcus mutans NG8 were grown from glycerol stocks in closed tubes containing Todd Hewitt Broth (THB) for 24 hours. Mono-species biofilms were grown in 96-well microtitre plates for 72 hours before exposure to antimicrobial agents. Dilution series of the synthetic antimicrobial peptide BM2 (D-NH2-RRRFWWFRRR-CONH2) and the widely-used endodontic disinfectants sodium hypochlorite (NaOCl) and saturated calcium hydroxide (Ca(OH)₂) were prepared as aqueous solutions. The efficacy of BM2, NaOCl and Ca(OH)₂ at causing biofilm disruption was measured using a crystal violet assay after 24, 48 and 72 h exposure. More clinically relevant biofilms formed on sterile root canal surfaces was exposed to BM2, NaOCl and Ca(OH)₂, stained using a LIVE/DEAD assay™ kit and visualized using confocal laser scanning microscopy (CLSM).
Result: Both the crystal violet assay and CLSM showed BM2 had antimicrobial activity against all microorganisms tested. BM2 at 40µg/mL was as effective as NaOCl (1%) and saturated Ca(OH)₂ against S. gordonii and S. mutans (P < 0.05). After 72 h mean biofilm mass was reduced ≥75% and the remaining cells mass was ≤50% viable. BM2 at 40µg/mL showed antimicrobial activity similar to 0.25% NaOCl and saturated Ca(OH)₂ in disrupting the E. faecalis biofilm.
Conclusion: BM2 at 40µg/mL showed robust antimicrobial activity against biofilms of major endodontic pathogens comparable to current endodontic disinfectants. BM2 should be further investigated as an alternative antimicrobial agent in endodontic treatment.