Given the roles that bacteriophages have been shown to play in the microbial ecology of numerous ecosystems and the roles that human viruses play in causing oral disease, we are testing the hypothesis that there are a significant number of diverse viruses (both bacteriophage and human viruses) that inhabit the human oral cavity and that examination of their genome sequences will unveil new clues to the role of viruses in the microbial ecology of the mouth (e.g. biofilm formation and disease). The goal of this discovery-driven R21 project is to determine the genetic diversity of oral viruses using Sanger/pyrosequencing hybrid sampling as a precursor to a more comprehensive evaluation of the genomic capacity of the virus component of the human mouth. Sequencing the viral gene pool within the mouth will give insight into the biological role that phage and human viruses play in this environment. There is one specific aim: To use a metagenomic approach to understand viral (both human viruses and bacteriophages) diversity from Mitomycin C-induced and non-induced human saliva and supra- gingival plaque samples. This approach differs significantly from previous attempts to identify oral bacteriophages through traditional culture-based procedures. There are a number of standard, robust methodologies for the estimation of the total numbers of viruses in environmental samples as well as new tools for the analysis of virus metagenomics data that will enable estimation of diversity and community structure. In our proposal, we will use TEM, epifluorescence quantitation techniques and a Sanger/pyrosequencing hybrid approach to obtain virus DNA sequences. Analysis of these data will provide the framework for future thorough examination of the genomic content of the viruses that exist in the human mouth and their role in human disease. Public Health: The goal of this discovery-based R21 proposal is to determine the genetic diversity of oral viruses as the first step to more comprehensive examination of the viral gene pool in the human oral cavity. Though this study is not meant to be comprehensive, we will still gain valuable insight into the gene content of oral viruses, leading to a better understanding of how these overlooked vectors influence homeostasis and disease. A future application of the data is the control of bacterial pathogens through the identification and production of bacteriophage-encoded products. ? ? ?
| Fouts, Derick E (2018) Amplification for Whole Genome Sequencing of Bacteriophages from Single Isolated Plaques Using SISPA. Methods Mol Biol 1681:165-178 |
| Depew, Jessica; Zhou, Bin; McCorrison, Jamison M et al. (2013) Sequencing viral genomes from a single isolated plaque. Virol J 10:181 |
