Intervention strategies of hemorrhagic colitis and HUS

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DESCRIPTION (provided by applicant): The broad aim of this proposal is to develop and utilize new and established animal model of enterohemorrhagic E. coli (EHEC) infection, in rabbits and dogs, to develop therapeutic regimens to prevent and treat EHEC disease. It is well recognized that shiga-toxin- (Stx)-producing strains of E. coli, acquired by ingestion of inadequately cooked meat, or other contaminated foods, cause hemorrhagic colitis, and may induce fatal hemolytic uremic syndrome (HUS). EHEC strains produce potent protein toxins named Shiga-like toxins (Stxs) because of their relatedness to Shiga toxin of Shigella dysenteriae. In addition, most EHEC share the ability to adhere intimately to intestinal epithelial cells by "attaching and effacing" (A/E)(7) mechanisms (Fig.2). Although EHEC attachment mechanisms may directly contribute to diarrheal disease, and may influence toxin delivery, the most severe intestinal and renal manifestations of EHEC infection result from toxin-mediated damage to vascular endothelium, with tissue edema, inflammatory infiltrates, cytokine production and vascular thrombi. At present, only supportive care is available to prevent the development of the severe, and frequently fatal, complications of EHEC infection. Strategies aimed at decreasing the toxin burden and preventing the interaction of Stxs with their endothelial receptors should prevent or ameliorate disease and damage in target organs (gut, CNS and kidney). Interventions developed in animal models can subsequently be applied to the prevention and management of EHEC disease. E. coli strain RDEC-HI9A infection of rabbits serves as the established animal model of EHEC disease for the initial intervention studies (Aims 1-4). RDEC-H19A, produced by the transfer of the toxin-converting phage H19A of an O26:H11 EHEC to the rabbit entero-pathogenic E. coli RDEC-1, is an attaching and effacing rabbit pathogen which produces high levels of Shiga-like toxin I (Stx-I), colonizes cecum and colon, and induces intestinal disease in rabbits with pathologic changes resembling human EHEC disease. Specific aims (1-4) of the proposal are to use animal models of EHEC infection to: 1). Test the ability of new toxin-receptor analogs, administered paretenteraly or enterically to prevent EHEC disease. 2). Further test the ability of passively administered immunoglobulin with anti-toxic activity to prevent EHEC disease. 3). Further examine whether antibiotic therapy has beneficial or harmful effects on the course of disease. 4). Further develop strategies for active immunization against EHEC using the Stx toxins of EHEC. 5). Specific aim 5 is to utilize canine specific A/E strains to produce new STECcapable of infecting dogs, which are susceptible to renal vascular lesions. We will transfer our labeled Stx-1 encoding phage to dog-specific A/E strains of E. coli and test their ability to produce intestinal and renal disease. The clinical studies in dogs will be performed at Kansas State University by Dr. Brad Fenwick who has described the Cutaneous and Renal Glomerular Vasculopathy (CRGV) in greyhound dogs exposed to Stx. 6). Specific Aim 6 is to extend our rabbit and dog models to be able to test similar strategies against EBEC strains expressing Stx-2. We will label and transfer toxin converting phage encoding Stx-2 to rabbit and dog specific A/E strains.