Periodontal diseases affect 35% of US adults and lead to bone and tooth loss. Treatment costs reach U$ 4.4 billion per year to American taxpayers. Studies have linked periodontal diseases and systemic conditions. Over half of the oral taxa have not been cultivated in vitro. There is a gap in knowledge regarding which of those taxa are common and numerous-and therefore relevant - in periodontal health and disease. Our long term goal is to unveil novel periodontal pathogens and determine the mechanisms they use for periodontal breakdown. The overall objective of this proposal is to identify prominent subgingival taxa associated with periodontal health or disease that are currently uncultivated or cultivable but currently unrecognized. Reports of uncultivated/ unrecognized taxa present in periodontitis have recently emerged. Our central hypothesis is that a subset of the uncultivated/unrecognized taxa in subgingival biofilms is pathogenic. Many oral uncultivated/unrecognized taxa are transients and brought in by food or liquids and have little, if any, relevance to periodontal health or disease. Thus, we do not aim at recovering just any """"""""domesticateable"""""""" uncultivated subgingival isolate;neither at growing uncultivated taxa in multispecies complex.
We aim at identifying numerically prominent uncultivated/unrecognized taxa in periodontal health or disease and recover them in pure culture as a first step in determining their role in periodontal health or disease. The rationale of this proposal is to eliminate the """"""""distraction"""""""" represented by transient taxa and focus the search for novel prominent subgingival bacteria. We will use innovative technologies to quantify a wide range of uncultivable/unrecognized taxa in large numbers of biofilm samples. We will test the central hypothesis and accomplish the objective of this application by pursuing the following Specific Aims. 1) We will identify prominent uncultivated/unrecognized bacteria in periodontal health and disease. This will be achieved by screening 320 individual subgingival biofilm samples from 16 periodontally healthy and 16 periodontitis subjects. We will employ an innovative technique developed by us, the RNA-Oligonucleotide Quantification Technique (ROQT). It is based on 16s rRNA targeted oligonucleotide probes and it will determine which of 140 uncultivated/unrecognized species/taxa/ clones are common and numerous in subgingival biofilms. Once prominent taxa are defined, 2) we will use the appropriate probes to isolate in pure culture, cultivable but as yet unrecognized prominent taxa. We will use 15 conditions for growth of those taxa.
This aim will distinguish taxa that can be cultivated under normal laboratory conditions. This research is significant because biologically meaningful taxa currently considered to be uncultivated/unrecognized will be isolated, characterized and properly classified. This is crucial for virulence, antigenicity and functional genomics studies and investigation of growth factors and signaling molecules. Results from this study will support NIDCR's mission by leading to the identification of pathogenic mechanisms and novel targets for disease prevention, diagnosis and therapy.
The proposed research is relevant of public health because, despite increasing knowledge of the development of periodontal diseases, these infections affect more than 30% of US adults and are more prevalent among minorities. These diseases have been linked to systemic conditions and their treatment costs billions annually. This project is relevant to NIDCR's mission because understanding of the role of uncultivated/unrecognized subgingival bacteria can provide new targets for prevention, diagnosis and treatment of oral infections.
