Comprehensive Genomic Survey of Antimicrobial-Resistance Bacteria in the Sewage Tank Replacement with Hospital Relocation

Miwa Katagiri,1,2 Makoto Kuroda,2 Tsuyoshi Sekizuka,2 Norihide Nakada,3 Yukitaka Ito,4 Masanobu Otsuka,4 Manabu Watanabe,1 Shinya Kusachi1 1Department of Surgery, Toho University Ohashi Medical Center, Meguro-ku, Tokyo, Japan; 2Laboratory of Bacterial Genomics, Pathogen Genomics Center, National Institute of Infectious Diseases, Shinjuku-ku, Tokyo, Japan; 3Research Center for Environmental Quality Management, Graduate School of Engineering, Kyoto University, Otsu-shi, Shiga, Japan; 4Division of Clinical Microbiology Laboratory, Toho University Ohashi Medical Center, Meguro-ku, Tokyo, JapanCorrespondence: Miwa KatagiriDepartment of Surgery, Toho University Ohashi Medical Center, 2-22-36 Ohashi, Meguro-ku, Tokyo, 153-8515, JapanTel +81354333086Fax +81354333091Email girioyoshikazu0513@gmail.comMakoto KurodaLaboratory of Bacterial Genomics, Pathogen Genomics Center, National Institute of Infectious Diseases, 1-23-1 Toyama, Shinjuku-ku, Tokyo, 162-8640, JapanTel +81352851111Fax +81352851150Email makokuro@niid.go.jpBackground: Excrement containing antimicrobial-resistant bacteria (ARB) is discharged from the hospital sewage through wastewater treatment plants (WWTP) into rivers, increasing the antimicrobial resistance (AMR) burden on the environment.Purpose: We illustrate the contamination of hospital sewage tanks with ARB harboring antimicrobial resistance genes (ARGs) using comprehensive metagenomic sequencing. During the study period, we moved to a new hospital building constructed for renovation. Therefore, we investigated the difference in bacterial flora in the sewage tanks for each building with different departments, and the change in bacterial flora over time in new sewage tanks. Furthermore, we performed a comparative genome analysis of extended spectrum β-lactamase (ESBL)-producing organisms (EPOs) from hospital sewage and clinical samples. Residual antibiotics in the sewage tank were also measured.Methods: Metagenomic analysis was performed on the hospital sewage samples, followed by whole genome sequencing of EPOs.Results: The bacterial composition of new sewage tanks was comparable with that of old tanks within 1 month after relocation and was instantly affected by excrement. The bacterial composition of sewage tanks in the old and new buildings, containing rooms where seriously ill patients were treated, was similar. Selection on CHROMagar ESBL allowed detection of EPOs harboring blaCTX-M and carbapenemase genes in all sewage tanks. One of the sewage Escherichia coli strain comprising ST393 harboring blaCTX-M-27 corresponded to the clinical isolates based on core genome analysis. Moreover, the levels of levofloxacin and clarithromycin in the hospital sewage were 0.0325 and 0.0135 μg/mL, respectively.Conclusion: Hospital sewage was contaminated with many ARB species, ARGs and residual antibiotics, which can cause a burden on WWTP sewage treatment. The bacterial flora in the sewage tank was rapidly affected, especially by the ward with seriously ill patients. AMR monitoring of hospital sewage may help detect carriers prior to nosocomial ARB-associated outbreaks and control the outbreaks.Keywords: hospital sewage, antimicrobial resistance, antimicrobial resistance gene, metagenome analysis, whole genome sequencing, residual antimicrobial reagents