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Methods for Enrichment and Sequencing of Oral Viral Assemblages: Saliva, Oral Mucosa, and Dental Plaque Viromes

Protocol
First Online:
Part of the Methods in Molecular Biology book series (MIMB, volume 1838)

Abstract

The oral cavity is a major portal of entry for human pathogens including viruses. However, metagenomics has revealed that highly personalized and time-persistent bacteriophage assemblages dominate this habitat. Most oral bacteriophages follow lysogenic life cycles, deploying complex strategies to manage bacterial homeostasis. Although bacterial dysbiosis underlies common oral pathologies such as caries and periodontitis, the cause of these bacteria replacements remains obscure, and it is theorized that bacteriophages play an important role. The enormous sensitivity of metagenomics coupled with next-generation sequencing has made technically feasible to address the putative role of bacteriophages in oral dysbiosis and represents a valuable tool to discover new human viruses.

This chapter proposes a workflow that consists of a simple viral enrichment protocol, two alternative random amplification methods, and next-generation sequencing to access virome composition in three oral environments: supragingival plaque, saliva, and mucosa. These protocols circumvent some well-known sources of bias, providing genomic information about DNA and RNA viral communities with minimal contamination from human and bacterial sources.

Key words

Oral viromes Saliva Oral mucosa Supragingival plaque Metagenomics 
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Notes

Acknowledgments

This work was supported by the Spanish Ministry of Economy and Competitiveness through grant SAF2012-38421 and a “Formación de Personal Investigador” Ph.D. studentship to M. P-M.

We acknowledge Patricia Suárez Rodríguez and Ana Rodríguez Galet for technical support and Áurea Soro-Simón, Alex Mira, Asier Eguía and José Manuel Aguirre-Urizar for providing valuable samples.

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© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Centro de Biología Molecular Severo Ochoa (Universidad Autónoma de Madrid-Consejo Superior de Investigaciones Científicas)MadridSpain

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