Dual-light photodynamic therapy administered daily provides a sustained antibacterial effect on biofilm and prevents Streptococcus mutans adaptation

Abstract

IntroductionAntibacterial photodynamic therapy (aPDT) and antibacterial blue light (aBL) are emerging treatment methods auxiliary to mechanical debridement for periodontitis. APDT provided with a near infrared (NIR) light in conjunction with an indocyanine green (ICG) photosensitizer has shown efficacy in several dental in-office-treatment protocols. In this study, we tested Streptococcus mutans biofilm sensitivity to either single-light (aPDT or aBL) or dual-light aPDT (simultaneous aPDT and aBL) exposure. Materials and MethodsBiofilm was cultured by pipeting diluted Streptococcus mutans suspension with growth medium on the bottom of well plates. Either a single-light aPDT (810-nm aPDT or 405-nm aBL) or a dual-light aPDT (simultaneous 810-nm aPDT and 405nm aBL) was applied, in both cases together with the ICG photosensitizer, while keeping the total light energy constant at 100J/cm2. Single-dose light exposures were given after one-day, four-day or fourteen-day biofilm incubations. Also, a repeated daily dose of the same light energy was applied during biofilm incubations on four-day and fourteen-day biofilms. Finally, antibacterial action of the dual-light aPDT with different relative ratios of 810 nm and 405 nm of light energy was examined on the single-day and four-day protocols. Biofilms were scraped, diluted into ratios between 1:1 to 1:100 000, and plated. After re-incubation, colony-forming units (CFUs) were counted, and confocal 3D biofilm imaging was performed. ResultsOn a one-day biofilm, dual-light aPDT was significantly more efficient than single-light aBL or aPDT, although all modalities were bactericidal. On a four-day maturated biofilm, a single exposure of aPDT or dual-light aPDT was more efficient than aBL, resulting in a four logarithmic scale reduction in bacterial counts. Surprisingly, when the same amount of aPDT was repeated with a daily dosing on a four-day or a fourteen-day biofilm, bacterial viability improved significantly. A similar but milder response of improved bacterial viability was seen after repetitive aBL application. The viability improvement was eliminated when dual-light aPDT was applied. By changing the relative light energy ratios in dual-light aPDT, a relative increase in aBL improved the antibacterial action of dual-light aPDT when the biofilm was older. ConclusionWhen aPDT is administered repeatedly to S. mutans biofilm, a single wavelength-based aBL or aPDT leads to a significant biofilm adaptation and increased S. mutans viability. The combined use of aBL light in synchrony with aPDT arrests the adaptation and provides significantly improved and sustained antibacterial efficacy.