Created August 16, 2016
Modified August 10, 2017
State Preliminary and in progress
Brief Description

40 16S amplicon libraries collected from sediments of the Adelie Basin

Acquisition Description

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).

Processing Description

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.



Dataset Maintainers

Stephanie A. CarrColorado School of Mines (CSM)
Beth N. OrcuttColorado School of Mines (CSM)
John R. SpearBigelow Laboratory for Ocean Sciences
Shannon RauchColorado School of Mines (CSM)
Shannon RauchColorado School of Mines (CSM)
Shannon RauchWoods Hole Oceanographic Institution (WHOI BCO-DMO)

BCO-DMO Project Info

Project Title Functional potential of the uncultivated Candidate Phylum OP9 from deep Antarctic marine sediments using single cell genome techniques
Acronym Adelie Basin Atribacteria
Created November 20, 2015
Modified November 20, 2015
Project Description

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

Data Project Maintainers
Stephanie A. CarrColorado School of Mines (CSM)Principal Investigator
Beth N. OrcuttBigelow Laboratory for Ocean SciencesCo-Principal Investigator
John R. SpearColorado School of Mines (CSM)Co-Principal Investigator