The influence of geographic distribution and type of habitat on the molecular epidemiology of ciprofloxacin resistant Escherichia coli was investigated. Ciprofloxacin resistant E. coli from wastewater, urban water with faecal contamination and faeces of gulls, pigeons and birds of prey, from Portugal, Spain and Sweden were compared based on multi‐locus sequence typing (MLST) and quinolone resistance genetic determinants. Multi‐locus sequence typing allowed the differentiation of E. coli lineages associated with birds of prey from those inhabiting gulls and waters. E. coli lineages of clinical relevance, such as the complex ST131, were detected in wastewater, streams and gulls in Portugal, Spain and Sweden.

Quinolone resistance was due to gyrA and parC mutations, although distinct mutations were detected in birds of prey and in wastewater, streams and gulls isolates. These differences were correlated with specific MLST lineages, suggesting resistance inheritance. Among the plasmid‐mediated quinolone resistance genes, only aac(6′)‐ib‐cr and qnrS were detected in wastewater, streams and gulls isolates, but not in birds of prey. The horizontal transfer of the gene aac(6′)‐ib‐cr could be inferred from its occurrence in different MLST lineages.