Genetic Analysis of Virulence Potential of Escherichia coli O104 Serotypes Isolated From Cattle Feces Using Whole Genome Sequencing

Abstract

Escherichia coli O104:H4, a Shiga toxin-producing hybrid pathotype that was implicated in a major foodborne outbreak in Germany in 2011, has not been detected in cattle. However, serotypes of O104, other than O104:H4, have been isolated from cattle feces, with O104:H7 being the most predominant. In this study, we investigated, based on whole genome sequence analyses, the virulence potential of E. coli O104 strains isolated from cattle feces, since cattle are asymptomatic carriers of E. coli O104. The genomes of ten bovine E. coli O104 strains (six O104:H7, one O104:H8, one O104:H12, and two O104:H23) and five O104:H7 isolated from human clinical cases were sequenced. Of all the bovine O104 serotypes (H7, H8, H12, and H23) that were included in the study, only E. coli O104:H7 serotype possessed Shiga toxins. Four of the six bovine O104:H7 strains and one of the five human strains carried stx1c. Three human O104 strains carried stx2, two were of subtype 2a, and one was 2d. Genomes of stx carrying bovine O104:H7 strains were larger than the stx-negative strains of O104:H7 or other serotypes. The genome sizes were proportional to the number of genes carried on the mobile genetic elements (phages, prophages, transposable elements and plasmids). Both bovine and human strains were negative for intimin and other genes associated with the type III secretory system and non-LEE encoded effectors. Plasmid-encoded virulence genes (ehxA, epeA, espP, katP) were also present in bovine and human strains. All O104 strains were negative for antimicrobial resistance genes, except one human strain. Phylogenetic analysis indicated that bovine E. coli O104 strains carrying the same flagellar antigen clustered together and STEC strains clustered separately from non-STEC strains. One of the human O104:H7 strains was phylogenetically closely related to and belonged to the same sequence type (ST-1817) as the bovine O104:H7 STEC strains. This suggests that the bovine feces could be a source of human illness caused by E. coli O104:H7 serotype. Because bovine O104:H7 strains carried virulence genes similar to human clinical strains and one of the human clinical strains was phylogenetically related to bovine strains, the serotype has the potential to be a diarrheagenic pathogen in humans.

Keywords: E. coli O104; cattle feces; serotypes; virulence genes; whole genome sequencing.

Figure 1

Phylogenetic tree and sequence types (ST) of stx-positive or –negative E. coli O104 strains isolated from cattle feces and human clinical strains constructed using Parsnp v1.2 and visualized using FigTree 1.4.3 (bootstrap values are provided for each branch).