MECHANISMS INVOLVED IN THE ADAPTATION OF Escherichia coli O157:H7 TO INTESTINAL MICROENVIRONMENT

A BRUBALLA; S MCATEER; GPINEDA; DAVID GALLY; R FERNÁNDEZ-BRANDOI; MV RAMOS; M PALERMO

Florencia

Congreso; 10th International Symposium on Shiga Toxin (Verocytotoxin) producing Escherichia coli infections; 2018

Societá Italiana di Microbiologia (SIM)

Although the production of Shiga toxin byenterohemorragic Escherichia coli(EHEC) determines Hemolytic Uremic Syndrome (HUS) onset, factors that modulateintestinal colonization are key components in pathogenesis and host mucosalimmune response. Type III secretion system (T3S) is essential for colonization andis encoded on the locus of enterocyte effacement island (LEE). The first operonencodes its own regulator Ler, which controls the transcription of the othersfour operons (LEE2-5). The aim of this work was to study whether Ler overexpression, with the subsequentoverexpression of LEE 2-5, determines an increase on EHEC pathogenesis. To dothis we transformed a human-isolated EHEC strain (125/99) with a plasmidcontaining the ler sequence under thecontrol of an IPTG-inducible promotor (125pLer). As a control we transformedthe same strain with the empty plasmid (125pWsk29). We tested the expression ofT3S proteins by SDS-PAGE and Western blot, and the adhesion to intestinal epithelialcells (HCT-8 and Caco-2). We also studied Shiga toxin (Stx2) production byELISA, as T3S proteins are crossregulated with the Stx prophage. We observed anincreased expression of EspB/D on 125pLer by Coomasie blue-staining and Westernblot. Besides, 125pLer strain showed an increased adhesion to HCT-8 and Caco-2cells (% adherence HCT-8= pLer: 43, pW: 16; Caco-2= pLer: 22, pW: 12). Stx2production was dependent on the time of pLer induction, ie more Stx2 productionwith more time of induction