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Biofilms that form on abiotic surfaces promote microbial persistence, increase tolerance to sanitizers and antibiotics, and contribute to dispersion in adverse environments. Ozone, a potent, highly reactive oxidizing agent that leaves no residue, has been investigated as a promising and sustainable alternative for surface decontamination in the food industry. In this study, the application of ozonated mist (OM) was evaluated to quantify the inactivation of adherent Staphylococcus aureus ATCC 25923 cells on AISI 304 stainless steel. Biofilms (n = 40) were formed for 72 h at 35 °C. OM was generated by an ultrasonic humidifier coupled to an ozone generator, operating with a gas flow of 2 L/min and an inlet concentration of 34.76 mg/L. The coupons were exposed to OM for 0.5, 1, 2, 3, 4, and 5 min. As a control, non-ozonated mist was used under the same operating conditions. The adhered cells were recovered and quantified in Soy Tryptone Agar. There was an average reduction of 0.86 and 1.70 LogCFU/cm² for each exposure time to OM, with a total reduction of 1.1 and 1.52 LogCFU/cm². Studies on biofilm eradication on abiotic surfaces use chemical sanitizers to reduce the sessile fraction of pathogens, with typical reductions between 0.5 and 3.0 log CFU·cm⁻². Under the conditions evaluated, the counts observed ranged from 5.38 to 4.88 log CFU·cm⁻², resulting in 4.48 ± 0.51 log CFU·cm⁻² at the end. The OM promoted significant inactivation of S. aureus in a short contact time. Compared to the non-ozonated mist, the OM showed equal or greater reductions, characterizing it as a technically viable alternative for the control of biofilms on abiotic surfaces.
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