AUTHOR=Kvesić Marija , Šamanić Ivica , Novak Anita , Fredotović Željana , Dželalija Mia , Kamenjarin Juraj , Goić Barišić Ivana , Tonkić Marija , Maravić Ana 

TITLE=Submarine Outfalls of Treated Wastewater Effluents are Sources of Extensively- and Multidrug-Resistant KPC- and OXA-48-Producing Enterobacteriaceae in Coastal Marine Environment

JOURNAL=Frontiers in Microbiology

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2022.858821

DOI=10.3389/fmicb.2022.858821

ISSN=1664-302X

ABSTRACT=<p>The rapid and ongoing spread of carbapenemase-producing <italic>Enterobacteriaceae</italic> has led to a global health threat. However, a limited number of studies have addressed this problem in the marine environment. We investigated their emergence in the coastal waters of the central Adriatic Sea (Croatia), which are recipients of submarine effluents from two wastewater treatment plants. Fifteen KPC-producing <italic>Enterobacteriaceae</italic> (nine <italic>Escherichia coli</italic>, four <italic>Klebsiella pneumoniae</italic> and two <italic>Citrobacter freundii</italic>) were recovered, and susceptibility testing to 14 antimicrobials from 10 classes showed that four isolates were extensively drug resistant (XDR) and two were resistant to colistin. After ERIC and BOX-PCR typing, eight isolates were selected for whole genome sequencing. The <italic>E. coli</italic> isolates belonged to serotype O21:H27 and sequence type (ST) 2795, while <italic>K. pneumoniae</italic> isolates were assigned to STs 37 and 534. Large-scale genome analysis revealed an arsenal of 137 genes conferring resistance to 19 antimicrobial drug classes, 35 genes associated with virulence, and 20 plasmid replicons. The isolates simultaneously carried 43–90 genes encoding for antibiotic resistance, while four isolates co-harbored carbapenemase genes <italic>bla</italic><sub>KPC-2</sub> and <italic>bla</italic><sub>OXA-48</sub>. The <italic>bla</italic><sub>OXA-48</sub> was associated with IncL-type plasmids in <italic>E. coli</italic> and <italic>K. pneumoniae</italic>. Importantly, the <italic>bla</italic><sub>KPC-2</sub> in four <italic>E. coli</italic> isolates was located on ~40 kb IncP6 broad-host-range plasmids which recently emerged as <italic>bla</italic><sub>KPC-2</sub> vesicles, providing first report of these <italic>bla</italic><sub>KPC-2</sub>-bearing resistance plasmids circulating in <italic>E. coli</italic> in Europe. This study also represents the first evidence of XDR and potentially virulent strains of KPC-producing <italic>E. coli</italic> in coastal waters and the co-occurrence of <italic>bla</italic><sub>KPC-2</sub> and <italic>bla</italic><sub>OXA-48</sub> carbapenemase genes in this species. The leakage of these strains through submarine effluents into coastal waters is of concern, indicating a reservoir of this infectious threat in the marine environment.</p>