Synergistic biocidal effects of metal oxide nanoparticles-assisted ultrasound irradiation: Antimicrobial sonodynamic therapy against Streptococcus mutans biofilms

Abstract

BackgroundAntimicrobial sonodynamic therapy (aSDT) is an adjunctive modality, which uses ultrasound irradiation to kill microbial cells by the activation of a sonosensitizer. The aim of this study was to evaluated the synergistic biocidal effects of zinc oxide nanoparticles (ZnO NPs) and titanium dioxide nanoparticles (TiO2 NPs) as the metal oxide nanoparticles (MONPs)-assisted ultrasound irradiation against Streptococcus mutans biofilms. Materials and methodsFollowing preparation and characterization of MONPs, cellular uptake and generation of intracellular reactive oxygen species (ROS) were assessed. After determination of the sub-significant reduction (SSR) doses of ZnO NPs, TiO2 NPs, ZnO/TiO2 NPs, and ultrasound intensity against S. mutans, anti-biofilm effects of aSDT were assessed using colorimetric assay, plate counting, and field emission scanning electron microscope (FESEM) analysis. Also, the metabolic activity of S. mutans and the expression levels of glucosyltransferase B (gtfB) as a main virulence factor of S. mutans were evaluated by XTT assay and quantitative real-time polymerase chain reaction following ZnO/TiO2 NPsSSR- mediated aSDT. ResultsThe finding of this study showed that an incubation time of 5 min was sufficient to achieve maximal uptake of MONPs. The ROS production following aSDT using ZnO NPs, TiO2 NPs, and ZnO/TiO2 NPs were ~ 4.1-, 5.6-, and 11.7-fold increase, respectively. The dose-dependent reduction in cell viability of S. mutans was revealed by increasing the concentrations of ZnO NPs, TiO2 NPs, ZnO/TiO2, as well as ultrasound intensities. According to the data, 1.5 µg/mL, 3.1 µg/mL, 25 µg/mL, and 0.75 W/cm2 were considered as the SSR doses of ZnO/TiO2 NPs, ZnO NPs, TiO2 NPs, and ultrasound intensity, respectively (P>0.05). ZnO/TiO2 NPsSSR-mediated aSDT showed a significantly higher biofilm inhibitory activity than the other treatment groups (P<0.05). Based on the FE-SEM analysis, aSDT based on the ZnO/TiO2 NPsSSR had a strong anti-biofilm effect against preformed biofilms of S. mutans on the enamel slabs. Also, the metabolic activity of S. mutans and the expression levels of gtfB were significantly decreased to 85.5% and 12.3-fold, respectively following ZnO/TiO2 NPsSSR-mediated aSDT (P<0.05). No considerable difference was observed in anti-biofilm activity between ZnO/TiO2 NPsSSR- mediated aSDT and 0.2% CHX (P>0.05). ConclusionThe results revealed anti-metabolic and anti-biofilm potential activities of ZnO/TiO2 NPs-mediated aSDT against S. mutans with the highest cellular uptake and ROS generation.