Campylobacter rectus (formerly Wolinella recta, Cr) is an important contributor to the progression of periodontitis. It is found in high numbers in diseased sites of HIV-associated periodontitis, in sites in refractory periodontitis patients, and in patients with both insulin- dependent and non-insulin-dependent diabetes mellitus. Our results indicate that fresh clinical isolates of a large number of Cr have a proteinaceous layer external to their outer membrane, which we refer to as the surface, or S layer. We have shown that the S layer is a 150 kDa protein; it has been purified to homogeneity and its amino terminal amino acid sequence determined. The S layer is lost from clinical isolates as they are passaged in vitro on solid media, or when cells are passaged in hemin limiting conditions. Importantly, S layer containing strains are more virulent in a murine lesion model than are S layer negative strains. The S layer also appears to interfere with the ability of Cr to interact with certain host cells in vitro. Biochemical, molecular genetic, and cell biology approaches will be combined in this grant to test the general hypothesis that: the function of the Cr S layer is as a surface component which protects the pathogen from host cellular and humoral defenses. To address this hypothesis, we will: 1) Determine the characteristics of the S layer by isolating and characterizing the gene (crs) encoding the S layer. The cloned gene will be used to construct isogenic crs+ and crs- mutants; 2) Determine the effect of the selected environmental stresses of carbon source limitation, hemin-stress, redox potential, and pH on S layer and omp expression using controlled chemostat conditions; 3) Using the isogenic crs+ and crs- strains, test the hole of the S layer on growth of Cr in a murine peritoneal growth chamber model and lesion formation in a murine abscess model; and 4) Determine the in vitro functional characteristics of wild-type (crs+) and mutant (crs-)Cr strains (1 above) with selected host cells of epithelial, fibroblast, and B cell lineage. The results of this grant will critically evaluate the role of the Cr S layer protein in both in vivo virulence models and in vitro functional assays. In addition, these experiments will begin to define the role of selected environmental stresses on the regulation and expression of the C. rectus S layer. Accomplishment of these goals could eventually permit the rational design of protocols which will reduce, alter or eliminate those environmental cues that enhance C. rectus colonization.
| Wang, B; Kraig, E; Kolodrubetz, D (2000) Use of defined mutants to assess the role of the Campylobacter rectus S-layer in bacterium-epithelial cell interactions. Infect Immun 68:1465-73 |
| Wang, B; Kraig, E; Kolodrubetz, D (1998) A new member of the S-layer protein family: characterization of the crs gene from Campylobacter rectus. Infect Immun 66:1521-6 |
| Nitta, H; Holt, S C; Ebersole, J L (1997) Purification and characterization of Campylobacter rectus surface layer proteins. Infect Immun 65:478-83 |
| Gilles, J A; Carnes, D L; Dallas, M R et al. (1997) Oral bone loss is increased in ovariectomized rats. J Endod 23:419-22 |
