Cyclic‐di‐ GMP signalling and biofilm‐related properties of the Shiga toxin‐producing 2011 German outbreak E scherichia coli O104:H4

Abstract

In 2011, nearly 4,000 people in Germany were infected by Shiga toxin (Stx)‐producing E scherichia coli O104:H4 with > 22% of patients developing haemolytic uraemic syndrome ( HUS ). Genome sequencing showed the outbreak strain to be related to enteroaggregative E. coli ( EAEC ), suggesting its high virulence results from EAEC ‐typical strong adherence and biofilm formation combined to Stx production. Here, we report that the outbreak strain contains a novel diguanylate cyclase (DgcX)—producing the biofilm‐promoting second messenger c‐di‐ GMP —that shows higher expression than any other known E. coli diguanylate cyclase. Unlike closely related E. coli , the outbreak strain expresses the c‐di‐ GMP ‐controlled biofilm regulator CsgD and amyloid curli fibres at 37°C, but is cellulose‐negative. Moreover, it constantly generates derivatives with further increased and deregulated production of CsgD and curli. Since curli fibres are strongly proinflammatory, with cellulose counteracting this effect, high c‐di‐ GMP and curli production by the outbreak O104:H4 strain may enhance not only adherence but may also contribute to inflammation, thereby facilitating entry of Stx into the bloodstream and to the kidneys where Stx causes HUS . image The detailed genomic and experimental characterization of cyclic‐di‐ GMP signaling and production of biofilm and curli fibres, but not cellulose, reveals a unique combination of features that may provide clues to the high virulence of the 2011 German outbreak E scherichia coli O104:H4 strain. The outbreak strain has a novel c‐di‐ GMP ‐producing diguanylate cyclase (DgcX) with the highest expression observed to date for such an enzyme in E. coli . Several other c‐di‐ GMP ‐related enzymes also show altered expression that can contribute to high c‐di‐ GMP accumulation potential. High levels of the c‐di‐ GMP ‐regulated biofilm regulator CsgD and amyloid curli fibres are produced at human body temperature, combined with an inability to generate cellulose. Since curli fibres were previously shown to be highly inflammatory with cellulose counteracting this effect, this high production of ‘naked’, i.e. non‐cellulose‐associated curli fibres by the outbreak strain may enhance inflammation, thereby facilitating efficient transition of Stx into the blood stream and progression to HUS .