Project Number: 5030-32000-100-00-D
Project Type: In-House Appropriated
Start Date: Dec 28, 2010
End Date: Dec 27, 2015
1) Determine and characterize molecular mechanisms promoting colonization, effective adherence, and persistence of Escherichia coli (E. coli) O157:H7 and other Shiga toxin-producing E. coli (STEC) in cattle. Evaluate the role of specific virulence factors in adherence of important non-O157:H7 STEC to tissue cultured cells and bovine intestinal tissues. 2) Understand the impact of bovine intestinal environment and immune responses on growth, adherence, and persistence of E. coli O157:H7 and other STECs in cattle. Determine the effects of signaling molecules produced in the bovine intestinal environment on selected virulence attributes of non-O157 STEC. 3) Conduct comparative analysis of bovine E. coli O157:H7 and STEC isolates of public health significance to identify components for use in developing rapid diagnostic tools and effective interventions; and 4) Develop and test efficacy of chemical, biological, subunit proteins, and whole cell vaccines to prevent or reduce colonization of cattle intestines by E. coli O157:H7 and STECs.
Experimental animal models, tissue cultures, and specific mutants will be used to describe molecular mechanism(s) enabling Escherichia coli (E. coli) O157:H7 bacteria to grow, adhere, and colonize the cattle intestine. Mutant strains lacking one or more of the virulence factors, such as type III secretion, motility, and bacterial cell surface fimbriae, will be constructed and tested in adherence assays using cultured cells and/or bovine intestinal tissues to delineate the importance of these factors in adherence of non-O157 Shiga toxin-producing E. coli (STEC) to these tissues. Reporter gene fusions and global gene analysis technologies will be used to determine effects of host gastrointestinal environment and innate immune system on the expression of specific bacterial genetic systems and metabolic pathways that promote E. coli O157:H7 persistence in cattle intestine. Select group of non-O157 STEC strains will also be examined for the effects of signaling molecules, especially stress hormones norepinephrine or epinephrine, and fermentation products produced in the bovine intestinal tract on the expression of virulence factors and adherence of these STEC strains to bovine intestinal tissues. The focus would be to examine virulence factors that are secreted through the type III secretion system or those expressed on bacterial cell surface for promoting bacterial motility, adherence, and aggregation on bovine intestinal tissues. Emerging non-O157 STEC serotypes will be compared with E. coli O157:H7 to identify genetic and molecular features unique to these serotypes. Bacterial genes or gene products identified in these studies will be used, based on their importance in colonization, for developing whole-cell or protein/subunit protein vaccines for reducing or eliminating E. coli O157:H7 and non-O157 STEC colonization and shedding in cattle.