Phages in a thermoreversible sustained-release formulation targeting E. faecalis in vitro and in vivo

Enterococcus faecalis is a key pathogen recovered from root canals when conventional treatment fails. Phage therapy has generated new interest in combating pathogens. A sustained-release formulation using specific phages against E . faecalis may offer an alternative approach. Objectives To evaluate the efficacy of anti- E . faecalis phages formulated in a thermo- sustained-release system against E . faecalis in vitro and in vivo . Methods EFDG1 and EFLK1 phages were formulated with poloxamer P407. Gelation time, phage survival, activity and toxicity were evaluated. Lytic activity was evaluated in vitro against E . faecalis at various growth phases, including anti-biofilm activity. Methods included viable bacterial count (CFU/mL), biofilm biomass determination and electron microscopy (live/dead staining). Further evaluation included infected incisors in an in vivo rat model. Anti -E . faecalis phage-cocktail suspension and sustained-release phage formulation were evaluated by viable bacterial count (CFU/mL), histology, scanning electron microscopy (SEM) and 16S genome sequencing of the microbiota of the root canal. Results Gelation time for clinical use was established. Low toxicity and a high phage survival rate were recorded. Sustained-release phages reduced E . faecalis in logarithmic (4 logs), stationary (3 logs) and biofilm (4 logs) growth phases. Prolonged anti-biofilm activity of 88% and 95% reduction in biomass and viable counts, respectively, was recorded. Reduction of intracanal viable bacterial counts was observed (99% of enterococci) also seen in SEM. Phage treatment increased Proteobacteria and decreased Firmicutes . Histology showed reduced periapical inflammation and improved healing following phage treatment. Conclusion Poloxamer P407 formulated with phages has an effective and long-lasting effect in vitro and in vivo targeting E . faecalis .