Infectious bovine keratoconjunctivitis (IBK), caused by the bacteria Moraxella bovis, will be used as a model system for human bacterial eye disease. IBK is a widespread, highly contagious ocular disease of cattle causing temporary and, occasionally, permanent blindness. IBK is an especially model system because; 1) its pili are closely related to the pili of human ophthalmic disease bacteria, and 2) well-defined animal model systems exists for testing the pathogenesis of altered bacteria in vivos as well as in vitro. The focus of these studies will be the role of the two different types of pili found within a given strain of M. bovis in the disease process, and particularly the function of the phase variation which occurs between the two pili types alpha, and beta in strain EPP63. The near term goals of these studies will be: 1) to determine the exact position of the inversion-recombination sites by sequence analysis of both alpha and beta expressing DNA's, 2) to produce phase-locked mutants of M. bovis strain EPP63 capable of stably expressing either only alpha or only beta-pili, 3) to test the M. bovis phase-locked mutants in vitro, in mice and in calves, and 4) to investigate the other genes involved in the pilin gene inversion event and potentially the other genes involved in pili assembly and expression. The sequence analysis will be done from lambda gt11 clones of beta-pilin producing DNA and either Charon 4A or lambda gt11 clones of alpha-pilin producing DNA. Phase-locked mutants will be produced by in vitro replacement of the inversion-recombination site distal to the expression site locus by a drug resistance gene marker. DNA from the drug resistant mutants will be used to transform alpha-piliated and beta-piliated M. bovis strains. Transformants unable to undergo phase variation, and containing either only alpha-pili or only beta-pili will be tested for their relative adherence ability in vitro using a corneal tissue assay. The phase-locked M. bovis strains will then be tested in eye inoculation studies in mice and calves. Beta- pilin-lacZ fusions will be constructed to study the control of the inversion event in either E. coli or M. bovis. Deletion, transposition and in vitro mutagenesis will be used to localize the positions of the gene(s) responsible for the inversion event, as well as potential genes involved in pili assembly and expression. The long range goal of this research is to obtain a better understanding of the role played by pili phase variation in ophthalmic bacterial diseases.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
1R01EY007125-01
Application #
3264057
Study Section
Bacteriology and Mycology Subcommittee 1 (BM)
Project Start
1987-08-01
Project End
1990-07-31
Budget Start
1987-08-01
Budget End
1988-07-31
Support Year
1
Fiscal Year
1987
Total Cost
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Type
Schools of Public Health
DUNS #
791277940
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Weir, S; Lee, L W; Marrs, C F (1997) Type-4 pili of Kingella denitrificans. Gene 192:171-6
Weir, S; Lee, L W; Marrs, C F (1996) Identification of four complete type 4 pilin genes in a single Kingella denitrificans genome. Infect Immun 64:4993-9
Ruehl, W W; Marrs, C; Beard, M K et al. (1993) Q pili enhance the attachment of Moraxella bovis to bovine corneas in vitro. Mol Microbiol 7:285-8
Weir, S; Marrs, C F (1992) Identification of type 4 pili in Kingella denitrificans. Infect Immun 60:3437-41
Tonjum, T; Marrs, C F; Rozsa, F et al. (1991) The type 4 pilin of Moraxella nonliquefaciens exhibits unique similarities with the pilins of Neisseria gonorrhoeae and Dichelobacter (Bacteroides) nodosus. J Gen Microbiol 137:2483-90
Patel, P; Marrs, C F; Mattick, J S et al. (1991) Shared antigenicity and immunogenicity of type 4 pilins expressed by Pseudomonas aeruginosa, Moraxella bovis, Neisseria gonorrhoaea, Dichelobacter nodosus, and Vibrio cholerae. Infect Immun 59:4674-6
Rozsa, F W; Marrs, C F (1991) Interesting sequence differences between the pilin gene inversion regions of Moraxella lacunata ATCC 17956 and Moraxella bovis Epp63. J Bacteriol 173:4000-6
Beard, M K; Mattick, J S; Moore, L J et al. (1990) Morphogenetic expression of Moraxella bovis fimbriae (pili) in Pseudomonas aeruginosa. J Bacteriol 172:2601-7
Fulks, K A; Marrs, C F; Stevens, S P et al. (1990) Sequence analysis of the inversion region containing the pilin genes of Moraxella bovis. J Bacteriol 172:310-6
Marrs, C F; Rozsa, F W; Hackel, M et al. (1990) Identification, cloning, and sequencing of piv, a new gene involved in inverting the pilin genes of Moraxella lacunata. J Bacteriol 172:4370-7

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