The Role of the AggR Regulon in the Virulence of the Shiga Toxin-Producing Enteroaggregative Escherichia coli Epidemic O104:H4 Strain in Mice

Abstract

An O104:H4 Shiga toxin (Stx)-producing enteroaggregative <i>Escherichia coli</i> (EAEC) strain caused a large outbreak of bloody diarrhea and the hemolytic uremic syndrome in 2011. We previously developed an ampicillin (Amp)-treated C57BL/6 mouse model to measure morbidity (weight loss) and mortality of mice orally infected with the prototype Stx-EAEC strain C227-11. Here, we hypothesized that mice fed C227-11 cured of the pAA plasmid or deleted for individual genes on that plasmid would display reduced virulence compared to animals given the wild-type (wt) strain. C227-11 cured of the pAA plasmid or deleted for the known pAA-encoded virulence genes <i>aggR</i>, <i>aggA</i>, <i>sepA</i>, or <i>aar</i> were fed to Amp-treated C57BL/6 mice at doses of 10¹⁰-10¹¹CFU. Infected animals were then either monitored for morbidity and lethality for 28 days or euthanized to determine intestinal pathology and colonization levels at selected times. The pAA-cured, <i>aggR</i>, and <i>aggA</i> mutants of strain C227-11 all showed reduced colonization at various intestinal sites. However, the <i>aggR</i> mutant was the only mutant attenuated for virulence as it showed both reduced morbidity and mortality. The <i>aar</i> mutant showed increased expression of the aggregative adherence fimbriae (AAF) and caused greater systemic effects in infected mice when compared to the C227-11 wt strain. However, unexpectedly, both the <i>aggA</i> and <i>aar</i> mutants displayed increased weight loss compared to wt. The <i>sepA</i> mutant did not exhibit altered morbidity or mortality in the Amp-treated mouse model compared to wt. Our data suggest that the increased morbidity due to the <i>aar</i> mutant could possibly be via an effect on expression of an as yet unknown virulence-associated factor under AggR control.