CONICET | Buscador de Institutos y Recursos Humanos

Shiga toxins-producing Escherichia coli (STEC) strains are responsible for intestinal disease and hemolytic uremic syndrome (HUS). STEC O157:H7 is by far the most prevalent serotype associated with HUS although STEC non-O157 strains isolated from children with HUS drastically increased during the past ten years. In Argentina, the main virulence factor of STEC is the Shiga toxin type 2 (Stx2) present in strains O157 and non-O157. Recently, it has been reported that some STEC non-O157 and, among them the serotype O113:H21, produce other cytotoxin named Subtilase (SubAB) which may contribute to the pathogenesis of HUS. The purpose of the present study was to examine the physiological and morphological effects of STEC O113:H21 on human colonic mucosa in order to better understand the mechanisms by which STEC non-O157 induces diarrhea. We have used O113:H21 STEC strain 98NK2 isolated from a patient with HUS and derivatives of this STEC strain with mutations in either stx2 (98NK2∆stx2) or subAB genes (98NK2∆subAB) kindly provided by Dr James Paton, University of Adelaide, Adelaide, Australia. The 98NK2 wild type and its derivatives mutated strains were grown to exponential phase in mammalian cell media and added to the mucosal side of the colonic mucosa placed between two identical Ringer solutions in the Ussing chamber. Transepithelial net water flux (Jw) and short circuit current (Isc) were recorded automatically using a special electro-optical device connected to a computer and an automatic voltage clamp system, respectively. All bacteria strains showed inhibition of Jw that was not observed when the corresponding bacteria supernatant was used. The Jw inhibition was time-dependent and already significant after 60 min of incubation whereas Isc remained unchanged for 90 min. Human colon mucosa incubated with purified SubAB also caused a significant inhibition of water absorption although the response at 0.4µg/ml was less than that observed with the bacterial strains. Histopathological studies of colon mucosa treated 90 min with 98NK2 wild type and 98NK2∆subAB strains revealed destruction of the upper third of the colonic mucosa and presence of hemorrhagic areas. On the other hand, 98NK2∆stx2 strain destroyed the surface epithelium, decreased the goblet cells and produced a moderate infiltration of neutrophils. The present results indicate that SubAB causes a significant inhibition of water absorption and a selective damage to the absorptive epithelial cells. These damages are extended to the crypt cells when Stx2 is present in the lumen. This is the first study showing that Stx2 and SubAB produced by the O113:H21 STEC strain may participate in the mechanisms of diarrhea.

enviar mensaje