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The emergence and spread of antimicrobial resistance (AMR) has become a persistent problem globally. In this study, an ozone treatment facility was established for an advanced hospital wastewater treatment in a core hospital facility in an urban area in Japan to evaluate the inactivation of antimicrobial-resistant bacteria and antimicrobials. Metagenomic DNA-seq analysis and the isolation of potential extended-spectrum <i>β</i>-lactamase (ESBL)-producing bacteria suggested that ozone exposure for at least 20 min is required for the adequate inactivation of DNA and ESBL-producing bacteria. <i>Escherichia coli</i> and <i>Klebsiella</i> species were markedly susceptible to 20-min ozone exposure, whereas <i>Raoultella ornithinolytica</i> and <i>Pseudomonas putida</i> were isolated even after an 80-min exposure. These ozone-resistant bacteria might play a pivotal role as AMR reservoirs in the environment. Nine antimicrobials (ampicillin, cefdinir, cefpodoxime, ciprofloxacin, levofloxacin, clarithromycin, chlortetracycline, minocycline, and vancomycin) were detected at 373 ng/L to 27 μg/L in the hospital wastewater, and these were removed (96–100% removal) after a 40-min treatment. These results facilitate a comprehensive understanding of the AMR risk posed by hospital wastewater and provides insights for devising strategies to eliminate or mitigate the burden of antimicrobial-resistant bacteria and the flow of antimicrobials into the environment. To the best of our knowledge, this is the first report on the implementation of a batch-type, plant-scale ozone treatment system in a hospital facility to execute and evaluate the inactivation of drug-resistant bacteria and antimicrobials.