Synergistic efficacy of high-intensity ultrasound and chlorine dioxide combination for Staphylococcus aureus biofilm control

This study investigated the synergistic efficacy of combined high-intensity ultrasound (HIU, 20 kHz, 60 W) and chlorine dioxide (ClO2, 4 mg/L) on the inactivation of viable cells and clearance of Staphylococcus aureus biofilms followed by comparing with those when treated by either HIU and ClO2. The bactericidal rate after the ClO2 treatment at 10 min was 82.62%; however, half of S. aureus remained sublethal or injured instead of fully dead as proved by flow cytometric assay. The bactericidal rate of the HIU treatment was even worse (18.72%). Inactivation rate of viable S. aureus cells after the combined treatment was 99.03% at 10 min and all bacteria were completely inactivated. Therefore, a strongly synergistic effect between HIU and ClO2 treatment was determined with a synergistic value of 1.24 (>1.15 as the critical limit); and the synergistic effect is mainly attributed to mechanical oscillation and cavitation generated by HIU which promote penetration and diffusion of ClO2 into the biofilm structure. Furthermore, transcriptional changes of biofilm-related genes in S. aureus indicate that the combined treatment reduced the biofilm formation through mediating quorum sensing (QS) signaling molecules, cell-cell adhesion, and polysaccharide intercellular adhesin (PIA) production; meanwhile, the secretion of enterotoxin in S. aureus was also suppressed after the combined treatment. These findings indicate that the HIU-ClO2 combination synergistically reduced the number of sublethal cells and efficiently detached the biofilm. Lastly, surface properties, especially roughness of food contact materials showed a significantly negative correlation with the clearance rate of biofilms after the combined treatment. This result suggests that the efficacy of biofilm detachment by using the combined treatment should be further evaluated based on the surface properties of food contact materials.

Automated summary

The combination of high-intensity ultrasound and chlorine dioxide treatment showed synergistic effects in inactivating viable cells and clearing Staphylococcus aureus biofilms, with the ultrasound promoting penetration and diffusion of ClO2. The treatment also reduced biofilm formation and toxin secretion, leading to a decrease in sublethal cells. Surface properties of food contact materials, particularly roughness, negatively correlated with biofilm clearance efficiency after the combined treatment, suggesting the need for further evaluation.