Biospecimen Analysis Core: Generating high quality data to characterize the human virome

Project Summary- Biospecimen Analysis Core The primary objective of the Biospecimen Analysis Core is to leverage the resources and expertise of the Broad Institute team in genomics–and in viral metagenomics in particular–to characterize the viromes from our diverse cohort of over 4,000 individuals from all over the United States. Specifically, we will use both well-vetted and, as needed, bespoke metagenomic approaches to capture viral sequences from seven anatomical sites (stool, urine, blood, mouth, nose, skin, and where relevant, vagina) from each participant. Leveraging the research and technical expertise of the Sabeti lab and Microbial Omics Core (MOC), we will develop sequencing process pipelines for each sample type, optimized to address the wide variation in total biomass, and RNA and DNA quantity and quality anticipated from the sample collection, with the goal of maximizing viral (and phage) sequence recovery. For example, RNA-Seq of mixed sample types will require innovative protocol development to deplete abundant off target RNA molecules prior to sequencing, and low-biomass samples such as plasma will require an agnostic all-nucleic-acid sequencing protocol to discover RNA and DNA viruses. Other areas of proposed methods development include long read sequencing to improve genome assemblies and pooled multiplex library construction to increase throughput and decrease costs. We will leverage the Broad Institute’s large scale Genomics Platform to support sample management and tracking, and for standard sequencing at scale, whereas specimens requiring more bespoke, high touch sample handling will be sequenced in the Sabeti Lab and/or MOC. The sequence data will serve two purposes: assembling a comprehensive and richly annotated catalog of viral genomes found at each anatomical site and/or time point, and characterizing the composition of the virome in each single sample. We expect to find many known viruses in our metagenomic sequencing data, but we expect to find even more unknown viruses. In order to find out which human cells they infect, and what other cell-types are around them in human tissues, we will perform spatial RNA sequencing of tissue sections on a limited set of newly discovered viruses. To image and characterize the virus itself, we will use a novel class of non-bleaching probes to decorate viral particles in vitro and then image the particle as it docks to the cell, invades the cell, and then continues monitoring its intracellular trafficking pathway. Together, our data will reduce the “dark matter” in the virome, i.e. the large number of viruses yet to be discovered, generate a large data set of biospecimens with quantitative virome profiles, and molecular snapshots and movies of viruses in tissues and living cells. In addition, consistent with our track record, we will share protocols and data rapidly and openly, and would welcome close collaboration with other VCC BACs to share insights and solutions to collectively develop the most comprehensive catalog of viral sequences possible across the Human Virome Program.