AUTHOR=Heljanko Viivi , Johansson Venla , Räisänen Kati , Anttila Veli-Jukka , Lyytikäinen Outi , Jalava Jari , Weijo Irma , Lehtinen Jaana-Marija , Lehto Kirsi-Maarit , Lipponen Anssi , Oikarinen Sami , Pitkänen Tarja , Heikinheimo Annamari ,  WastPan Study Group , Al-Mustapha Ahmad , Kurittu Paula , Länsivaara Annika , Hyder Rafiqul , Tiwari Ananda , Hokajärvi Anna-Maria , Kolehmainen Aleksi , Möttönen Teemu , Luomala Oskari , Juutinen Aapo , Blomqvist Soile , Savolainen-Kopra Carita , Sarekoski Anniina 

TITLE=Genomic epidemiology of nosocomial carbapenemase-producing Citrobacter freundii in sewerage systems in the Helsinki metropolitan area, Finland

JOURNAL=Frontiers in Microbiology

VOLUME=14

YEAR=2023

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2023.1165751

DOI=10.3389/fmicb.2023.1165751

ISSN=1664-302X

ABSTRACT=<p>Multi-drug resistance is emerging in <italic>Citrobacter freundii</italic>, which is the third most common carbapenemase-producing (CP) <italic>Enterobacteriaceae</italic> in humans in Finland due to recent outbreaks. The objective of this study was to determine if wastewater surveillance (WWS) could detect CP <italic>C. freundii</italic> strains causing infections in humans. Selective culturing was used to isolate CP <italic>C. freundii</italic> from the hospital environment, hospital wastewater, and untreated municipal wastewater in Helsinki, Finland, between 2019 and 2022. Species were identified using MALDI-TOF, and presumptive CP <italic>C. freundii</italic> isolates were subjected to antimicrobial susceptibility testing and further characterized by whole genome sequencing. A genomic comparison was conducted to compare isolates collected from the hospital environment, untreated municipal wastewater, and a selection of isolates from human specimens from two hospitals in the same city. We also examined the persistence of CP <italic>C. freundii</italic> in the hospital environment and the impact of our attempts to eradicate it. Overall, 27 <italic>bla</italic><sub>KPC − 2</sub>-carrying <italic>C. freundii</italic> were detected in the hospital environment (ST18; <italic>n</italic> = 23 and ST8; <italic>n</italic> = 4), while 13 <italic>bla</italic><sub>KPC − 2</sub>-carrying <italic>C. freundii</italic> (ST8) and five <italic>bla</italic><sub>VIM − 1</sub>-carrying (ST421) <italic>C. freundii</italic> were identified in untreated municipal wastewater. CP <italic>C. freundii</italic> was not identified in hospital wastewater. We found three clusters (cluster distance threshold ≤ 10 allelic difference) after comparing the recovered isolates and a selection of isolates from human specimens. The first cluster consisted of ST18 isolates from the hospital environment (<italic>n</italic> = 23) and human specimens (<italic>n</italic> = 4), the second consisted of ST8 isolates from the hospital environment (<italic>n</italic> = 4), untreated municipal wastewater (<italic>n</italic> = 6), and human specimens (<italic>n</italic> = 2), and the third consisted of ST421 isolates from the untreated municipal wastewater (<italic>n</italic> = 5). Our results support previous studies suggesting that the hospital environment could act as a source of transmission of CP <italic>C. freundii</italic> in clinical settings. Furthermore, the eradication of CP Enterobacteriaceae from the hospital environment is challenging. Our findings also showed that CP <italic>C. freundii</italic> is persistent throughout the sewerage system and demonstrate the potential of WWS for detecting CP <italic>C. freundii</italic>.</p>