Characterization of Pathogenic <i>Escherichia coli</i> in River Water by Simultaneous Detection and Sequencing of 14 Virulence Genes

The occurrence of pathogenic <i>Escherichia coli</i> in environmental waters increases the risk of waterborne disease. In this study, 14 virulence genes in 669 <i>E. coli</i> isolates (549 isolates from the Yamato River in Japan, and 30 isolates from each of the following hosts: humans, cows, pigs, and chickens) were simultaneously quantified by multiplex PCR and dual index sequencing to determine the prevalence of potentially pathogenic <i>E. coli</i>. Among the 549 environmental isolates, 64 (12%) were classified as extraintestinal pathogenic <i>E. coli</i> (ExPEC) while eight (1.5%) were classified as intestinal pathogenic <i>E. coli</i> (InPEC). Only ExPEC-associated genes were detected in human isolates and pig isolates, and 11 (37%) and five (17%) isolates were classified as ExPEC, respectively. A high proportion (63%) of cow isolates possessed Shiga-toxin genes (<i>stx1</i> or <i>stx2</i>) and they were classified as Shiga toxin-producing <i>E. coli</i> (STEC) or enterohemorrhagic <i>E. coli</i> (EHEC). Among the chicken isolates, 14 (47%) possessed <i>iutA</i>, which is an ExPEC-associated gene. This method can determine the sequences as well as the presence/absence of virulence genes. By comparing the sequences of virulence genes, we determined that sequences of <i>iutA</i> were different among sources and may be useful for discriminating isolates, although further studies including larger numbers of isolates are needed. Results indicate that humans are a likely source of ExPEC strains in the river