INFLUENCE OF Ler OVEREXPRESSION ON THE PATHOGENICITY OF A SHIGA TOXIN-PRODUCING Escherichia coli O157:H7 (EHEC) STRAIN

RFERNÁNDEZ-BRANDO; G PINEDA; D GALLY,; A BRUBALLA; S MCATEER; M GÓMEZ; M V RAMOS; M PALERMO

Buenos Aires

Congreso; Reunión Conjunta de Sociedades de Biociencia, 13-17 de Noviembre,; 2017

SAIC-SAIB-SAI-SAA-SAB-SAB-SAFE-SAFI-SAH-SAP

Although the production ofShiga toxin by enterohemorragic Escherichiacoli (EHEC) determines Hemolytic Uremic Syndrome (HUS) onset, factors thatmodulate intestinal colonization are key components in pathogenesis and hostmucosal immune response. Type III secretion system (T3S) is essential forcolonization and is encoded on the locus of enterocyte effacement island (LEE).The first operon encodes its own regulator Ler, which controls thetranscription of the others four operons (LEE2-5). The aim of this work was tostudy whether Ler overexpression,with the subsequent overexpression of LEE 2-5, determines an increase on EHECpathogenesis. To do this we transformed a human-isolated EHEC strain (125/99) witha plasmid containing the ler sequenceunder the control of an IPTG-inducible promotor (125pLer). As a control wetransformed the same strain with the empty plasmid (125pWsk29). We tested theexpression of T3S proteins by SDS-PAGE and Western blot, and the adhesion to intestinalepithelial cells (HCT-8 and Caco-2). We also studied Shiga toxin (Stx2)production by ELISA, as T3S proteins are crossregulated with the Stx prophage. Weobserved an increased expression of EspB/D on 125pLer by Coomasie blue-stainingand Western blot. Besides, 125pLer strain showed an increased adhesion to HCT-8and Caco-2 cells (% adherence HCT-8= pLer: 43, pW: 16; Caco-2= pLer: 22, pW: 12).Stx2 production was dependent on the time of pLer induction, ie more Stx2production with more time of induction