Novel Antibiofilm Chemotherapy Targets Exopolysaccharide Synthesis and Stress Tolerance in Streptococcus mutans To Modulate Virulence ExpressionIn Vivo

Abstract

ABSTRACTFluoride is the mainstay of dental caries prevention, and yet current applications offer incomplete protection and may not effectively address the infectious character of the disease. Therefore, we evaluated the effectiveness of a novel combination therapy (CT; 2 mM myricetin, 4 mMtt-farnesol, 250 ppm of fluoride) that supplements fluoride with naturally occurring, food-derived, antibiofilm compounds. Treatment regimens simulating those experienced clinically (twice daily for ≤60 s) were used bothin vitroover a saliva-coated hydroxyapatite biofilm model andin vivowith a rodent model of dental caries. The effectiveness of CT was evaluated based on the incidence and severity of carious lesions (compared to fluoride or vehicle control). We found that CT was superior to fluoride (positive control,P< 0.05); topical applications dramatically reduced caries development in Sprague-Dawley rats, all without altering theStreptococcus mutansor total populations within the plaque. We subsequently identified the underlying mechanisms through which applications of CT modulate biofilm virulence. CT targets expression of keyStreptococcus mutansgenes during biofilm formationin vitroandin vivo. These are associated with exopolysaccharide matrix synthesis (gtfB) and the ability to tolerate exogenous stress (e.g.,sloA), which are essential for cariogenic biofilm assembly. We also identified a unique gene (SMU.940) that was severely repressed and may represent a potentially novel target; its inactivation disrupted exopolysaccharide accumulation and matrix development. Altogether, CT may be clinically more effective than current anticaries modalities, targeting expression of bacterial virulence associated with pathogenesis of the disease. These observations may have relevance for development of enhanced therapies against other biofilm-dependent infections.