Persistence of E. coli 0157:H7 in Ruminants

Enterohemorrhagic E. coli (EHEC) are recognized as the leading cause of infectious bloody diarrhea in the United States as well as a major underlying cause of acute renal failure among children. The disease in humans can be fatal, especially foryoung children and the elderly, but chronic illness is also an important feature of the disease. It has been estimated that EHEC O157:H7 causes more than 110,000 cases ofillness and 91 deaths/year in the U.S. Outbreaks have been traced to contaminated beef,milk or manure-contaminated produce and fruit juices. While much has been learned about the epidemiology of naturally-occurring EHEC infections in ruminants (cows andsheep), we still do not understand why ruminants appear to be the principal reservoir for these organisms and what factors may be involved in their colonization of and persistence in ruminants. We will utilize a genetic technique called Signature Tagged Mutagenesis(STM) to identify the genes responsible for colonization and persistence of EHEC O157:H7 in sheep. This technique involves the use of genetic elements containing unique molecular bar codes that are inserted at random into the chromosome of virulent EHEC O157:H7 disrupting genes causing loss of functions. By inoculating sheep, it will bepossible to identify specific insertions that result in organisms incapable of colonizing orpersisting in the ruminant gastrointestinal tract. The genes in which they are inserted canbe identified by DNA sequence analysis. These genes can then become targets for new intervention strategies to prevent colonization of EHEC O157:H7 in ruminants.