Increased Adherence and Expression of Virulence Genes in a Lineage of Escherichia coli O157:H7 Commonly Associated with Human Infections

Abstract

BackgroundEnterohemorrhagic Escherichia coli (EHEC) O157:H7, a food and waterborne pathogen, can be classified into nine phylogenetically distinct lineages, as determined by single nucleotide polymorphism genotyping. One lineage (clade 8) was found to be associated with hemolytic uremic syndrome (HUS), which can lead to kidney failure and death in some cases, particularly young children. Another lineage (clade 2) differs considerably in gene content and is phylogenetically distinct from clade 8, but caused significantly fewer cases of HUS in a prior study. Little is known, however, about how these two lineages vary with regard to phenotypic traits important for disease pathogenesis and in the expression of shared virulence genes.Methodology/Principal FindingsHere, we quantified the level of adherence to and invasion of MAC-T bovine epithelial cells, and examined the transcriptomes of 24 EHEC O157:H7 strains with varying Shiga toxin profiles from two common lineages. Adherence to epithelial cells was >2-fold higher for EHEC O157:H7 strains belonging to clade 8 versus clade 2, while no difference in invasiveness was observed between the two lineages. Whole-genome 70-mer oligo microarrays, which probe for 6088 genes from O157:H7 Sakai, O157:H7 EDL 933, pO157, and K12 MG1655, detected significant differential expression between clades in 604 genes following co-incubation with epithelial cells for 30 min; 186 of the 604 genes had a >1.5 fold change difference. Relative to clade 2, clade 8 strains showed upregulation of major virulence genes, including 29 of the 41 locus of enterocyte effacement (LEE) pathogenicity island genes, which are critical for adherence, as well as Shiga toxin genes and pO157 plasmid-encoded virulence genes. Differences in expression of 16 genes that encode colonization factors, toxins, and regulators were confirmed by qRT-PCR, which revealed a greater magnitude of change than microarrays.Conclusions/SignificanceThese findings demonstrate that the EHEC O157:H7 lineage associated with HUS expresses higher levels of virulence genes and has an enhanced ability to attach to epithelial cells relative to another common lineage.

The influence of the accessory genome on bacterial pathogen evolution

“…39 Moreover, the clade that is associated with more severe symptoms also had a higher ability to attach to epithelial cells and had higher virulence gene expression. 40 This suggests that bacteriophage repertoires are relatively stably associated with individual lineages within EHEC O55:O157.…”

“…It is also important to point out that EHEC O55:O157 is not the only EHEC lineage within E. coli as at least 24 other E. coli lineages also acquired an enterohaemorrhagic lifestyle: the comparison of O55:O157 with three of these other lineages revealed that they independently acquired different bacteriophages that carry similar virulence gene repertoires. 42 In conclusion, specific bacteriophage repertoires appear to be vertically inherited within specific bacterial lineages contributing to lineage-specific fitness and virulence 39,40 while horizontal transfer of bacteriophages between phylogenetically quite distant groups allows for emergence of new pathogenic lineages with very similar phenotypes.…”

Enteroaggregative E. coli O104 from an outbreak of HUS in Germany 2011, could it happen again?

“…Cases of EHEC infection normally present to health facilities as bloody diarrhoea although more severe complications can occur. The frequency of these complications is dependent on the toxin encoded; the presence of vtx2a has been shown to be associated with a more virulent infection [49] partly due to increased expression [50]. The ability to adhere to intestinal cells has also been shown to be associated with virulence and although EHEC, as with EPEC, normally adhere using the LEE [47], there are other mechanisms of attachment within E. coli and outbreaks have been caused by several different lineages of EHEC using non-LEE mediated attachment.…”

A Quantitative Enzyme-Linked Immunosorbent Assay for Shiga Toxin 2a Requiring Only Commercially Available Reagents

“…It was previously reported that E. coli O157:H7 isolates from lineage II produced less Stx2 than those from lineage I and I/II 28 ; however, our findings show that the Stx2 levels in strains from lineage II (PA40, 41) were not significantly different from strains belonging to lineage I (PA5, 31, 32, 49) or lineage I/II (PA22, 35,38,47). Second, although strains from clade 8 were suggested to produce higher Stx2 levels than isolates from other clades, 29,30 our data indicates this is not universal. In our study, clade 8 strain PA35 produced similar amounts of Stx2 as others clades, including clade 2 (PA31), clade 5 (22), clade 6 (PA38) and clade 7 (PA40, 41).…”

“…Several studies have concluded, for example, that lineage I and I/II isolates produce more Stx2 than lineage II isolates, 28 and that clade 8 isolates are high toxin producers compared to isolates from other clades. 29,30 Most studies measure Stx2 production by semi-quantitative commercial kits, 10 qPCR/microarray, 10,29,30 or by semi-quantitative western blots. 30 Concerning mRNA quantification, several studies have reported that Stx2 transcript levels do not always correlate with Stx2 toxin production.…”