Lability of the pAA Virulence Plasmid in Escherichia coli O104:H4: Implications for Virulence in Humans

Abstract

Background Escherichia coli O104:H4 that caused the large German outbreak in 2011 is a highly virulent hybrid of enterohemorrhagic (EHEC) and enteroaggregative (EAEC) E. coli. The strain displays “stacked-brick” aggregative adherence to human intestinal epithelial cells mediated by aggregative adherence fimbriae I (AAF/I) encoded on the pAA plasmid. The AAF/I-mediated augmented intestinal adherence might facilitate systemic absorption of Shiga toxin, the major virulence factor of EHEC, presumably enhancing virulence of the outbreak strain. However, the stability of pAA in the outbreak strain is unknown. We therefore tested outbreak isolates for pAA, monitored pAA loss during infection, and determined the impact of pAA loss on adherence and clinical outcome of infection.Methodology/Principal Findings E. coli O104:H4 outbreak isolates from 170 patients (128 with hemolytic uremic syndrome [HUS] and 42 with diarrhea without HUS) were tested for pAA using polymerase chain reaction and plasmid profiling. pAA-harboring bacteria in stool samples were quantified using colony blot hybridization, and adherence to HCT-8 cells was determined. Isolates from 12 (7.1%) patients lacked pAA. Analyses of sequential stool samples demonstrated that the percentages of pAA-positive populations in the initial stools were significantly higher than those in the follow-up stools collected two to eight days later in disease (P≤0.01). This indicates a rapid loss of pAA during infections of humans. The pAA loss was associated with loss of the aggregative adherence phenotype and significantly reduced correlation with HUS (P = 0.001).Conclusions/SignificanceThe pAA plasmid can be lost by E. coli O104:H4 outbreak strain in the human gut in the course of disease. pAA loss might attenuate virulence and diminish the ability to cause HUS. The pAA instability has clinical, diagnostic, epidemiologic, and evolutionary implications.

QseC Signaling in the Outbreak O104:H4 Escherichia coli Strain Combines Multiple Factors during Infection

“…In contrast, EAEC tends to be more efficient when transporting Shiga toxin from intestinal epithelium to the bloodstream due to thick biofilm formation mediated by AAFs and TIF, factors that complicate EAEC pathogenesis. These data are consistent with a study by Zhang et al (89), who showed that the outbreak strain of patients with hemorrhagic colitis had lost the pAA plasmid during the course of the disease in the European outbreak, which in turn caused a decrease in the ability of the EAEC O104:H4 (Stx ϩ ) strain to cause HUS, since these strains become less adherent, showing the genetic plasticity of these bacteria during the course of the outbreak. Motility has been well described in EHEC, as well as its regulation by QseC (16,34,90), similarly to S. Typhimurium mechanisms (19,22).…”

Selection of horizontal gene transfer through public good production

“…For instance, Agrobacterium plasmids can be lost at high rates (25). Many secreted toxins (likely to behave as public goods) are both transferable and unstable, like virulence toxins in Escherichia coli (26) (27) or effector genes from the plant pathogen Pseudomonas syringae (28).…”

Sorbitol-Fermenting Enterohemorrhagic Escherichia coli O157:H − Isolates from Czech Patients with Novel Plasmid Composition Not Previously Seen in German Isolates

“…Plasmid profiles and Southern blot hybridization. Plasmid profiles were determined as described earlier (58,59). For Southern blot hybridization, plasmid-extracted DNAs (Plasmid Midi kit; Qiagen, Hilden, Germany), either undigested or digested with BamHI or SmaI (New England BioLabs, Frankfurt, Germany), were separated in a 0.6% agarose gel, blotted to a nylon membrane (Roche Molecular Biochemicals, Mannheim, Germany), and hybridized with digoxigenin-labeled (DIG High Prime kit; Roche Molecular Biochemicals) sfpA, EHEC-hlyA, katP, espP, and etpD probes prepared as described earlier (60).…”