Table of Contents

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

16S rRNA sequence data from venting fluids and microbial mats at Axial Seamount, 2011. Samples collected on the AT18-08 cruise.

Diffuse fluids were collected from newly discovered snowblower vents at Axial Seamount in late July 2011 with the ROV Jason II using the hydrothermal fluid and particle sampler (Butterfield et al., 2004). White and orange flocculent materials were collected on the subsequent University Of Washington Visions' 11 cruise, in support of the Regional Scale Nodes component of the Ocean Observatories Initiative in August 2011. White flocculent material was collected from the orifice of the Subway snowblower vent on dives R1467 (White Floc 1) and R1472 (White Floc 2) and orange flocculent material was collected on the seafloor distal to Marker 33 during dive R1472 where it coated freshly deposited basalt. All of the fluid and floc samples analyzed in this study are from a small area in the south rift zone at the southeastern edge of Axial Caldera, with the exception of background seawater which was collected outside of the caldera.

Total genomic DNA was extracted from Sterivex filters as previously described (Sogin et al., 2006) with the minor modifications described by Akerman et al. (2013). Total genomic DNA was extracted from 20 to 30mg of wet flocculent material using a MoBio UltraClean® Soil DNA Isolation Kit. The V6 region of 16S rRNA genes were amplified in triplicate for each sample with previously reported primers designed for archaea and bacteria (Huber et al., 2007) that were modified to include indices and barcodes compatible with the Illumina HiSeq1000 platform rather than 454 Life Sciences Adapters (Eren et al., 2013). Triplicate PCR amplifications were pooled for each sample, cleaned with a Qiagen MinElute kit, and quanitified by PicoGreen assay on a Turner Biosystems spectrophotometer. Fifty nanograms of each cleaned amplicon library was then size selected with a 2% agarose PippinPrep cassette to produce a narrow range of fragment sizes from 200 to 300 bp for sequencing and cleaned again to remove agarose. All of the amplicon libraries included in this study were sequenced in the same run and on the same paired end lane, along with 60 other libraries. Equimolar amounts of pooled amplicon libraries and a metagenomic library were run in the same lane to avoid known difficulties of sequencing low complexity amplicon libraries with Illumina (Caporaso et al., 2012).

Related references:
Meyer, J.L., Akerman, N.H., Proskurowski, G. and J.A. Huber. 2013. Microbiological characterization of post-eruption "snowblower" vents at Axial Seamount, Juan de Fuca Ridge. Frontiers in Microbiology. 4:153. doi:10.3389/fmicb.2013.00153

Paired Illumina sequencing reads were quality filtered to remove any reads containing ambiguous nucleotides and only pairs with perfectly overlapping reads were used for further analysis. Quality-filtered reads are publicly available through the VAMPS database, http://vamps.mbl.edu,under the project name JAH_AXV_Bv6 and JAH_AXV_Av6, where orange floc is listed as "eruption mat", white floc 1 is listed as "snow_R1467", and white floc 2 is listed as "snow_R1472". Sequences were clustered at 97% similarity with a minimum word length of 30, using usearch (Edgar, 2010). Taxonomy was assigned by global alignment for sequence taxonomy (GAST; Huse et al., 2008) with the SILVA 111 database (Quast et al., 2012). Operational taxonomic units (OTUs) were then analyzed with Qiime 1.5 (Caporaso et al., 2010). Even sequencing depth per sample was established by multiple rarefactions to roughly 75% of the smallest sequencing depth, using a total of 195,000 bacterial reads and 145,000 archaeal reads per sample. To compare bacterial communities in snowblower fluids and flocculent samples to background seawater by dendrogram, we retrieved bacterial V6 454 reads from background seawater collected outside the Axial Caldera from the VAMPS database under the project name KCK_SMT_Bv6, fluid sample FS501. To compensate for the fewer number of reads in the background seawater sample, a second set of multiple rarefactions was performed with 7112 reads per sample. Distance matrices were calculated for 10 rarefactions using the Morisita-Horn index (Horn, 1966) and the resulting tree topographies were clustered using UPGMA to create a final jackknifed tree.

BCO-DMO Processing:
- modified parameter names to conform with BCO-DMO naming conventions;
- removed "m" (meters) in depth column;
- changed format of date to YYYYmmdd;
- replaced commas with semi-colons;
- Added cruise_id field.

Collaborative Research: Function, activity, and adaptation of microbial communities in geochemically diverse subseafloor habitats

The integration of both laboratory and field-based chemical and microbiological measurements into a quantitative predictive framework is crucial to understanding the microbial ecology of marine systems. This project work will provide a quantitative assessment of the functional diversity, activity, and physiological adaptation of microbial communities in geochemically diverse subseafloor habitats. Results will guide development of models for linking biogeochemical processes with particular microbial communities at deep-sea hydrothermal vents, with implications for other marine habitats as well. The focus of the effort is at Axial Seamount, a well-studied, active, deep-sea hydrothermal seamount in the NE Pacific Ocean. Samples already collected from Axial, along with a field program in Year 2, will serve as the foundation for the three objectives, which are to:

1. Determine and quantify the functional diversity and activity (expression) of key subseafloor microbial lineages at Axial Seamount.

2. Determine physiological adaptations to the subseafloor habitat by quantifying the growth response of Axial Seamount isolates to in-situ geochemical parameters.

3. Develop a quantitative predictive framework for linking particular types of geochemical vent conditions with specific microbial functional groups and activities at Axial Seamount.

Specific outcomes of this project include the creation of a comprehensive quantitative microbiological and chemical dataset on diffuse and adjacent high-temperature vents within Axial Seamount. This database will include chemical measurements (gases, nutrients, metals, isotopes, and calculated Gibbs free energies) relevant to microbial metabolic processes that can be compared to microbiological data (abundance and activity of microbial lineages and functional genes, growth rates of subseafloor isolates at relevant environmental conditions) using statistical analysis to identify how specific microbial activity is linked to the geochemical environment. This project builds on previous studies of microbial population structure and geochemical measurements at Axial Seamount and addresses critical gaps in current knowledge and understanding that are impeding progress of modeling hydrothermal systems. Results will increase understanding of deep-sea hydrothermal ecosystems as well as provide new insights into controls on the distribution and activity of marine microbial communities throughout the world's oceans.

NeMO10 TN253 Cruise Report