DNA Probes for Gram Positive Periodontal Pathogens

Most diagnostic tests for periodontal pathogens have focused on enumeration of the major gram-negatassociated with diseased sites, although both gram-negative and gram-positive bacteria have been impagents of periodontal disease. The long-term objective of this project is to develop a DNA probe-basand quantitation of two gram-positive bacteria which have been associated with sites of periodontalPeptostreptococcus micros and Streptococcus intermedius. The specific aims of Phase I of the projectgenomic DNA from P. micros and S. intermedius, (2) to develop a sample preparation protocol which ascomplete lysis of gram-positive bacteria before immobilization of DNA on nitrocellulose, (3) to deteand specificity of whole genomic probes for P. micros and S. intermedius, (4) to construct genomic land S. intermedius, and (5) to identify clones from the genomic libraries which can be used as highlspecies-specific DNA probes. Many genomic DNA fragments will be screened by Southern blot analysis twhich do not cross-react with genomic DNA probes from closely related species. Achievement of theseallow the validation (in Phase II) of DNA probe based assays for the detection of P. micros and S. isubgingival plaque samples. Inclusion of these assays in a diagnostic test panel for periodontal patclinicians with a more comprehensive assessment of the microbial status of diseased sites.