Hypericin- and mTHPC-mediated photodynamic therapy for the treatment of cariogenic bacteria

Abstract

Abstract Objective Dental caries is one of the most common diseases in Western countries. Its pathoetiology is multifactorial, however, bacteria including Streptococcus mutans and the closely related Streptococcus sobrinus are regarded as key factors involved in this process. The fact that therapeutic approaches to eradicate these microorganisms are still limited prompted us to investigate the treatment potential of photodynamic therapy with the photoactive compounds hypericin (HYP) and meso-tetra(hydroxyphenyl)chlorin (mTHPC) in vitro. Material and methods S. mutans and S. sobrinus were cultivated under standard conditions and incubated with HYP (Invitrogen, Basel, Switzerland), the liposomal mTHPC derivative Foslipos (FOS, Biolitec, Jena, Germany), or a mixture of both at concentrations ranging between 0.625 and 10 μg/ml for various time points. Following a thorough washing step, bacteria were irradiated with a dental polymerization instrument (400–505 nm). All samples were subjected to serial dilutions and spiral plating on blood agar plates. Viable colony counts were determined after 48 h in culture. Photosensitizer fluorescence of bacteria was visualized by confocal microscopic techniques. Results One hundred percent of S. sobrinus could be killed by a 15 min incubation with as little as 2.5 μg/ml HYP, 5 μg/ml FOS or a mixture of 1.25 μg/ml of each photosensitizer followed by light activation of 120 s. In contrast to S. sobrinus, S. mutans displayed a significant dark toxicity for FOS (10–1.25 μg/ml) and no relevant PDT effects using HYP (10–0.625 μg/ml) under these conditions. HYP-mediated PDT effects (10 μg/ml) could be enhanced to more than 99.9% by prolonging photosensitizer incubation to 30 min and fractional illumination (2×120 s). Complete eradication of S. mutans was achieved by incubation for 15 min with a mixture of 0.625 μg/ml each of FOS and HYP and illumination for 120 s. Conclusion For both S. mutans and S. sobrinus, short PDT protocols with FOS and/or HYP could be established that completely eradicated these cariogenic bacteria in suspension. Our study, however, indicated that careful optimization of PDT conditions may be necessary for successful treatment of even closely related bacterial species. In multispecies microbial populations, the application of photosensitizer combinations for PDT may be useful.