Porphyromonas gingivalis is recognized as a prime etiological agent in periodontitis, a destructive disease of the supporting structures of the teeth which is prevalent in adults. Molecular genetic research on P. gingivalis has begun to build a foundation of understanding concerning how this organism colonizes the oral cavity and causes pathology. A dramatic increase in this understanding is likely now that the nucleotide sequence of the genome of P. gingivalis W83 has been determined. To date, genetic analyses of this organism have been possible using a number of tools adapted from other bacterial systems. A few genetic tools have been specifically created for use in Porphyromonas sp., but the systematic construction of plasmid vectors is critically needed if we are to fully exploit the genomic nucleotide sequence of this pathogen. We propose to accomplish this by developing, testing, and distributing plasmid constructs needed for the in-depth postgenomic analyses of Porphyromonas gingivalis. Specifically, we shall: 1. Characterize and native and recombinant plasmids from oral black pigmented Gram negative anaerobes for their ability to stably support extrachromosomal replication in P. gingivalis; 2. Design, build, characterize and test several robust shuttle vectors for the analysis of P. gingivalis; 3. Using 4 different non-clonal P. gingivalis strains, evaluate cellular and molecular variables that may affect their suitability as hosts for molecular genetic analyses; 4. Distribute these new genetic tools and knowledge about their use to workers studying Porphyromonas gingivalis and other black pigmented Gram-negative anaerobes.
