Bacteriodes gingivalis is a Gram-negative, anaerobic bacterium which is strongly implicated as an etiological agent of periodontal disease. Because surface structures of pathogenic organisms are among the most important factos in initiating disease, especially those of mucosal surfaces, it is important to identify surface structures of B. gingivalis which play a significant role in the pathogenic capabilities of this organism. At the same time, it is also desirable to identify and characterize surface antigens of B. gingivalis in order to study the host's response to this organism and ultimately produce an effective vaccine for periodontal disease. The cloning of B. gingivalis surface structures would provide a new and much improved approach to defining the surface of this periodontopathogen. It is proposed here to accomplish this task by """"""""shot-gunning"""""""" B. gingivalis chromosomal DNA via an expression vector (plasmid) into Escherichia coli in order to produce a genomic library. Those E. coli transformants of the library which encode B. gingivalis antigens will be identified by a filter binding assay using antisera to B. gingivalis outer membrane and/or whole cells. Polyclonal, monospecific antisera to cloned antigens will be used in ELISA and immunoelectronmicroscopy to determine if the cloned protein is surface-associated in B. gingivalis antigen will be screened for the expression of the B. gingivalis adhesin for saliva-coated hydroxyapatite (SHA) and for expression of the B. gingivalis surface antigen active in hemagglutination (HA). The cloned B. gingivalis antigens found to be involved in adherence to SHA and/or in HA will be purified using immunoaffinity chromatography. Specific physico-chemical characteristics of the purified cloned proteins will be determined and the involvement of the proteins in adherence to SHA and in HA will be verified by testing them in the in vitro assays. HA and adherence to SHA are two of the very few functions assayable in vitro and ascribed thus far to surface components of B. gingivalis. The in vivo role of these """"""""adhesins"""""""" is unclear. The cloning of the B. gingivalis adhesins in addition to other surface-associated antigens of B. gingivalis should facilitate the identification of the roles of the adhesins in vivo, genetically isolate major antigens, and provide a more thorough understanding the B. gingivalis surface.
| Bélanger, Myriam; Kozarov, Emil; Song, Hong et al. (2012) Both the unique and repeat regions of the Porphyromonas gingivalis hemagglutin A are involved in adhesion and invasion of host cells. Anaerobe 18:128-34 |
