Antimicrobial Resistance, Conjugative Plasmids and Pathogenicity in Escherichia spp. isolated from Natural water and Wastewater Environments in Stockholm, Sweden

To investigate the occurrence of antibiotic resistance genes (ARGs), conjugative plasmids and virulence genes in Swedish waterborne Escherichia spp. isolates, water samples were collected from urban freshwater swimming areas, beaches in the Baltic Sea, and the primary wastewater treatment plant (WWTP) in Stockholm, Sweden. During the summer of 2022, a total of 69 isolates were isolated from filters grown on ESBL-selective and non-selective agar. All isolates were analyzed by phenotypic antibiotic susceptibility tests, conjugation assays and whole genome sequencing. In addition, antibiotic residues were also quantified in the water samples. Of the 69 isolates, 44 (64%) were multidrug-resistant (MDR), exhibiting resistance to three or more antibiotic classes determined by disc diffusion. WWTP-derived E. coli isolates demonstrated a significantly higher prevalence of extended-spectrum β-lactamase (ESBL) genes compared to freshwater isolates (p < 0.0001). The isolates belonged to a diverse range of multi-locus sequence types (MLST) and most isolates harbored at least one plasmid. A total of 16 strains transferred conjugative plasmids carrying various combinations of ARGs conferring resistance to cefotaxime (CTX), tetracycline, streptomycin and trimethoprim. CTX resistance genes, including bla _CTX-M-15 , were found on IncF, IncN, IncBOKZ, and IncI plasmids as well as on the chromosome in a subset of ST38 isolates. IncF plasmids exhibited low (< 10 ^− 7 ) or undetectable transfer frequencies, while IncN and IncI1 plasmids displayed high transfer frequencies (10 ^− 2 − 10 ^− 4 ). This study demonstrates that while antimicrobial resistance in E. coli from natural water environments is still low in Stockholm, Sweden, WWTPs are a significant source of MDR and pathogenic E. coli . These strains frequently carry highly conjugative IncN and IncI1 plasmids containing multiple ARGs, posing a potential risk of increased environmental dissemination of MDR and/or pathogenic E. coli via treated effluent discharge.