Table of Contents

• Dataset Description
  • Methods & Sampling
  • Data Processing Description
• Parameters
• Deployments
• Project Information
• Program Information
• Funding

40 16S amplicon libraries collected from sediments of the Adelie Basin during the Integrated Ocean Drilling Program (IODP) Expedition 318, aboard R/V JOIDES Resolution, Site U1357.

Samples were collected at 66.413S, 140.425E from depths 0.05 - 97 meters below sea floor.

Samples for molecular biological analysis were collected shipboard from 10 cm sections of the intact core using sterile cut-end 5 mL syringes. Samples were immediately frozen at –80 degrees C and maintained at this temperature during transport and storage to the home laboratory. Genomic DNA was extracted from sediment samples using a bead beating/phenol chloroform protocol. Amplicons of the 16S rRNA gene were prepared for 454 pyrosequencing using a PCR-touchdown annealing temperature strategy from Don et al. (1991) with procedure modifications and modified universal primers 515f and 927r of Osburn et al. (2011). Forward primers included the 454 Life Science A adaptors and a sample specific 8 nt barcode. The reverse primer included the 454 Life Science B adaptors. Amplicon concentrations for each sample were quantified using a 2100 Bioanalyzer (Agilent Technologies, Colorado Springs, CO, USA), pooled (22 ng DNA/sample) and concentrated using a Savant DNA 12 Speed Vac Concentrator (Thermo Scientific, Waltham, MA, USA). The pooled DNA was gel purified using the Montage DNA Gel Extraction Kit (Millipore, Bellerica, MA, USA), and sequenced on a Roche 454 FLX titanium platform at Engencore, University of South Carolina (now Selah Genomics).

Pyrosequencing reads were analyzed using the Quantitative Insights Into Microbial Ecology (QIIME) Pipeline. Reads between 200 and 500 base pairs with a quality score of 27 or above were denoised using the QIIME denoiser for titanium runs, and clustered into operational taxonomic units (OTUs) at a 97% similarity threshold using UCLUST ( Edgar, 2010 ). Taxonomy of OTUs was assigned by BLAST  against the Silva SSU NR Reference database, release 102 within QIIME. Chimeras were identified and removed using QIIME’s ChimeraSlayer Wrapper.

Sequences were deposited into  the MG-RAST database under accession numbers 4624791.3–4634830.3.

Bacteria belonging to the newly classified candidate phylum “Atribacteria” (formerly referred to as “OP9” and “JS1”) are common in anoxic methane-rich sediments. However, the metabolic functions and biogeochemical role of these microorganisms in the subsurface remains unrealized due to the lack of pure culture representatives. In this study of deep sediment from Antarctica’s Adélie Basin, collected during Expedition 318 of the Integrated Ocean Drilling Program (IODP), Atribacteria-related sequences of the 16S rRNA gene were abundant (up to 51% of the sequences) and steadily increased in relative abundance with depth throughout the methane-rich zones. To better understand the metabolic potential of Atribacteria within this environment, and to compare with phylogenetically distinct Atribacteria from non-deep-sea environments, individual cells were sorted for single cell genomics from sediment collected from 97.41 m below the seafloor from IODP Hole U1357C. As observed for non-marine Atribacteria, a partial single cell genome suggests a heterotrophic metabolism, with Atribacteria potentially producing fermentation products such as acetate, ethanol, and CO2. These products may in turn support methanogens within the sediment microbial community and explain the frequent occurrence of Atribacteria in anoxic methane-rich sediments. This first report of a single cell genome from deep sediment broadens the known diversity within the Atribacteria phylum and highlights the potential role of Atribacteria in carbon cycling in deep sediment.

This project was funded by a C-DEBI Postdoctoral Fellowship

The mission of the Center for Dark Energy Biosphere Investigations (C-DEBI) is to explore life beneath the seafloor and make transformative discoveries that advance science, benefit society, and inspire people of all ages and origins.

C-DEBI provides a framework for a large, multi-disciplinary group of scientists to pursue fundamental questions about life deep in the sub-surface environment of Earth. The fundamental science questions of C-DEBI involve exploration and discovery, uncovering the processes that constrain the sub-surface biosphere below the oceans, and implications to the Earth system. What type of life exists in this deep biosphere, how much, and how is it distributed and dispersed? What are the physical-chemical conditions that promote or limit life? What are the important oxidation-reduction processes and are they unique or important to humankind? How does this biosphere influence global energy and material cycles, particularly the carbon cycle? Finally, can we discern how such life evolved in geological settings beneath the ocean floor, and how this might relate to ideas about the origin of life on our planet?

C-DEBI's scientific goals are pursued with a combination of approaches:
(1) coordinate, integrate, support, and extend the research associated with four major programs—Juan de Fuca Ridge flank (JdF), South Pacific Gyre (SPG), North Pond (NP), and Dorado Outcrop (DO)—and other field sites;
(2) make substantial investments of resources to support field, laboratory, analytical, and modeling studies of the deep subseafloor ecosystems;
(3) facilitate and encourage synthesis and thematic understanding of submarine microbiological processes, through funding of scientific and technical activities, coordination and hosting of meetings and workshops, and support of (mostly junior) researchers and graduate students; and
(4) entrain, educate, inspire, and mentor an interdisciplinary community of researchers and educators, with an emphasis on undergraduate and graduate students and early-career scientists.

Note: Katrina Edwards was a former PI of C-DEBI; James Cowen is a former co-PI.

Data Management:
C-DEBI is committed to ensuring all the data generated are publically available and deposited in a data repository for long-term storage as stated in their Data Management Plan (PDF) and in compliance with the NSF Ocean Sciences Sample and Data Policy. The data types and products resulting from C-DEBI-supported research include a wide variety of geophysical, geological, geochemical, and biological information, in addition to education and outreach materials, technical documents, and samples. All data and information generated by C-DEBI-supported research projects are required to be made publically available either following publication of research results or within two (2) years of data generation.

To ensure preservation and dissemination of the diverse data-types generated, C-DEBI researchers are working with BCO-DMO Data Managers make data publicly available online. The partnership with BCO-DMO helps ensure that the C-DEBI data are discoverable and available for reuse. Some C-DEBI data is better served by specialized repositories (NCBI's GenBank for sequence data, for example) and, in those cases, BCO-DMO provides dataset documentation (metadata) that includes links to those external repositories.