Environmental Genome Shotgun Sequencing of Marine Virioplankton
Marine viruses are the most numerous, most diverse and the least genetically characterized biological entities on Earth. Bacteriophages, viruses that specifically infect prokaryotes (Bacteria and Archaea) comprise the great majority of viruses in marine ecosystems. Through cycles of infection, replication and host cell lysis, phages impact multiple pathways and processes involved in the population biology of host organisms and the functioning of marine environments as a whole. This research project seeks to: 1) extensively sequence, annotate and analyze virioplankton metagenomes within two oxic surface water environments (Dry Tortugas and Gulf of Maine) and one hypoxic marine environment (Chesapeake Bay O2 minimum zone); 2) compare the diversity of virioplankton with host populations and temperate viruses existing as prophage; and 3) investigate whether phages contribute to some of the apparent genomic differences between Prochlorococcus marinus (MIT9312) and Candidatus pelagibacter ubique HTCC1062 (SAR11) and their respective close environmental relatives. Owing to their ubiquity and abundance in the pelagic ocean, together Prochlorococcus and SAR11 are perhaps the most important bacterial genera within the global carbon cycle. The Dry Tortugas sample was collected as part of the Sorcerer II Global Ocean Sampling Expedition and the Gulf of Maine sample was collected aboard the NOAA R/V Delaware II as part of an NSF-funded bacterioplankton Microbial Observatory. As such, each of these samples can be paired with synoptic microbial metagenomic data. Lastly, we have collected virioplankton samples from a hypoxic basin within the Chesapeake Bay. It is here that the experimental objective of assaying temperate phage diversity will be addressed. At all sites, 16S rDNA and 18S rDNA libraries are under analysis to compare the relative levels of diversity between viruses and hosts. Noting the extraordinary abundance of viruses in the biosphere (1031 individuals), representation of viral genetic data is woefully inadequate. The viral metagenomic data generated from this project will be invaluable to our understanding of marine viral diversity, the fundamental nature of virus-host interactions and their influence on bacterial evolution in the marine environment.
About EGSSMV:
Science Crew during the NOAA sponsored survey cruise ECOMON
Marine virioplankton: Sampling locations
Top: Dr. Shannon Williamson pre-filtering the (freezing cold) water through a pool filter into a 200L carboy on the ship’s deck. Bottom: the de-watering pump that was submerged to collect water. This pump was used since the team needed to collect the water rapidly and the peristaltic pump was too slow to fill the giant 200L carboy.
Grant Number: CB-0626826
Presentations:
2008 Fifth Aquatic Viral Workshop, Vancouver, BC, Canada
Making sense of the chaff: What will metagenomic approaches tell us about viral ecology?
K. Eric Wommack
High-throughput sequencing of viral genomic nucleic acids from environmental samples, i.e., viral metagenomics, is adding exquisite genetic detail to our view of natural viral assemblages. Indeed, as compared to microbial consortia, viral assemblages are ideally suited to metageomic approaches owing to the small coding-dense nature of viral genomes. For example, we now appreciate that in aquatic environments the phenomena of ‘viral photosynthesis’ is common and widespread; and that the extraordinary genotypic diversity seen in local viral assemblages may rival that of the entire global marine virome. It is quite possible that dsDNA viruses alone are among the greatest reservoirs of genetic diversity in the biosphere. Despite these and other notable advances, one consistent message we’ve obtained from the analysis of viral sequence data (metagenome or genome) is that the majority of viral genes are completely novel or show homology to only genes of unknown function. Two decades of aquatic viral ecology research have established that marine microbial communities support large viral populations with rapid turnover rates. As a consequence we can assume that most genes within aquatic viral metagenomes are actively replicated and expressed within aquatic microbial communities. Thus, one challenge for the coming third decade of the ‘third age of phage’ is to leverage metagenomic resources for an increasingly sophisticated view of the role of viral processes within microbial communities. This talk will review the current status of viral metagenomics and propose paths forward to meeting this challenge.
Widening the Camera Lens: Metagenomic and Biogeochemical Characterization of Marine Virioplankton
Danielle M. Winget 1, Douglas W. Fadrosh 2, K. Eric Wommack 1, Shannon J. Williamson 3
Viruses are widely recognized as abundant and active members of marine microbial communities. Numerous recent studies of viral processes and genomics in temperate surface marine ecosystems have revealed stunning viral diversity and infection patterns. However, research has generally focused on lytic dsDNA bacteriophages and rarely considered viral processes and genetics concurrently. Do all marine environments share the same viruses, or does viral diversity and their ecosystem impacts vary with environmental characteristics? To address this lack of information, shotgun metagenome sequence libraries of dsDNA viral assemblages in surface water samples from the Dry Tortugas (DT) and Gulf of Maine (GOM) and anoxic water samples from the Chesapeake Bay (CB), were collected while simultaneously measuring variations in viral and bacterial abundance and production. To date, ~130,000 sequence reads have been collected for the DT and GOM clone libraries. At the CB site, the diversity of inducible temperate phages was also compared to that of the total viral assemblage, and 3,800 quality control reads for two induced phage and four total virioplankton libraries have been gathered. Samples for the total viral assemblage libraries were collected over a diel cycle from anoxic CB waters for correlation with environmental parameters. Initial analysis revealed about 60% of viral metagenome sequences had no homologs within GenBank nr and the induced viral libraries contained a lower frequency of homologs to known phages. Interestingly, all the anoxic libraries showed a high frequency of hits to the anaerobic proteobacteria Anaeromyxobacter sp. with highest occurrences in the induced libraries. Significant diel cycles were observed in viral and bacterial production in CB surface water, but in viral and bacterial abundance. Likewise, significant diel cycles in bacterial production and viral abundance occurred in CB anoxic waters, without a corresponding variation in bacterial abundance. Thus, virioplankton within anoxic Chesapeake Bay waters appear to be both genetically and functionally distinct from previously examined viral assemblages.
1. College of Marine and Earth Studies, University of Delaware, Newark, DE, USA
2. J. Craig Venter Institute, Microbial and Environmental Genomics, Rockville, MD, USA
3. J. Craig Venter Institute, Microbial and Environmental Genomics, La Jolla, CA, USA