Y. enterocolitica causes a range of clinical syndromes. In young children and infants the disease manifests as watery diarrhea. In children and young adults infections result in mesenteric lymphadenitis and terminal ileitis. In adults more serious complications can occur such as erythema nodosum and reactive arthritis. Septicemia, with a 50% mortality rate, is a rare but serious complication, occurring most often in individuals whose underlying illnesses result in compromised host defenses. How Y. enterocolitica causes these diseases is as yet unknown. However, epithelial cell invasion is thought to be an important aspect of Y. enterocolitica pathogenesis. In preliminary experiments we identified two genetic loci, inv and ail, that confer an invasive phenotype on E. coli strain HB101. The experiments outlined in this proposal are aimed at characterizing the bacterial genes and their products involved in invasion, and at assessing the role these """"""""invasion"""""""" genes (and products) play in the virulence and pathogenesis of this organism. The long term goals are to understand the bacteria-host interaction at the molecular level, and to determine how the invasion process is coordinated with other aspects of Yersinia pathogenesis. Specifically, we propose the following experiments: 1) Further characterization of the inv and ail loci and identification of their gene products by transposon insertion mutagenesis, maxicell analysis, and DNA sequencing. 2) Determine if inv and/or ail are required for invasion of tissue culture cells and virulence in a mouse model. To do this, defined mutations constructed in E. coli will be recombined onto the Y. enterocolitica chromosome, replacing the wild type genes. The invasion phenotype of the mutants will be compared with that of the wild type strain. 3) Do the products of the inv and ail genes act directly by interacting with a receptor on the surface of the eukaryotic cell, or indirectly by modifying a bacterial cell surface structure. We will begin to address this question by a) determining if antibody to Inv or Ail blocks invasion, and b) determining if purified Inv or Ail can competitively inhibit invasion. 4) Study how the expression of the invasion of the genes is regulated by assessing the effect of environmental factors on fusions of lacZ or phoA to inv and ail.
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