Ozone ultrafine bubble water inhibits the early formation of Candida albicans biofilms

Abstract

This study aimed to investigate the effect of ozone ultrafine bubble water (OUFBW) on the formation and growth of Candida albicans (C. albicans) biofilms and surface properties of denture base resins. OUFBWs were prepared under concentrations of 6 (OUFBW6), 9 (OUFBW9), and 11 ppm (OUFBW11). Phosphate buffered saline and ozone-free electrolyte aqueous solutions (OFEAS) were used as controls. Acrylic resin discs were made according to manufacturer instructions, and C. albicans was initially cultured on the discs for 1.5 h. A colony forming unit (CFU) assay was performed by soaking the discs in OUFBW for 5 min after forming a 24-h C. albicans biofilm. The discs after initial attachment for 1.5 h were immersed in OUFBW and then cultured for 0, 3, and 5 h. CFUs were subsequently evaluated at each time point. Moreover, a viability assay, scanning electron microscopy (SEM), Alamar Blue assay, and quantitative real-time polymerase chain reaction (qRT-PCR) test were performed. To investigate the long-term effects of OUFBW on acrylic resin surface properties, Vickers hardness (VH) and surface roughness (Ra) were measured. We found that OUFBW9 and OUFBW11 significantly degraded the formed 24-h biofilm. The time point CFU assay showed that C. albicans biofilm formation was significantly inhibited due to OUFBW11 exposure. Interestingly, fluorescence microscopy revealed that almost living cells were observed in all groups. In SEM images, the OUFBW group had lesser number of fungi and the amount of non-three-dimensional biofilm than the control group. In the Alamar Blue assay, OUFBW11 was found to suppress Candida metabolic function. The qRT-PCR test showed that OUFBW down-regulated ALS1 and ALS3 expression regarding cell-cell, cell-material adhesion, and biofilm formation. Additionally, VH and Ra were not significantly different between the two groups. Overall, our data suggest that OUFBW suppressed C. albicans growth and biofilm formation on polymethyl methacrylate without impairing surface properties.