The object of the proposed research is to understand the genetic and molecular nature of chromosomal microbial determinants that contribute to the pathogenesis of extraintestinal Escherichia coli infection and disease. Cloned DNA sequences have been isolated encompassing the operon specifying E. coli hemolysin (Hly), the Pil operon specifying type I fimbriae, and the operon Pap which is associated with the synthesis of P pili mediating adherence to uroepithelial cells. The Pap and Hly operons will be examined by in vitro methods to determine their specific gene products and the DNA sequence of the gene specifying the functional pili and hemolysin will be determined. The distribution of Hly and Pap-specific DNA sequences among E. coli and other enteric bacteria will be examined as will the relative location of these genes on the bacterial chromosome. In addition, the functional basis for the attachment of P pili to a di-galactoside receptor and other eucaryotic cell receptors will be defined. An ascending pyelonephritis model has been established in mice. We propose to examine the contribution of Hly, Pap, type 1 fimbriae and the ColV-associated iron sequestration system to the pathogenesis of urinary tract infection. An avirulent strain of E. coli will be genetically manipulated sequentially to provide a series of derivatives which are homogenic except for one or a selected combination of virulence determinants. These strains will be examined in order to evaluate the relative contribution of each gene product to the infectious process.
