Table of Contents

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

soxB gene sequence data from microbial mats at Axial Seamount, 2011. Sample collected on cruise AT18-08.

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.

We targeted the amplification of the soxB gene in Epsilonproteobacteria to assess the diversity of sulfur oxidizers in white floc vs. orange floc samples. The soxB gene was amplified in one white flocculent sample (White Floc 2) and in the orange flocculent sample, using the newly designed primers and conditions described by Akerman et al. (2013). The PCR reaction mixture consisted of 1X buffer (Promega), 4 mM MgCl2, 0.2 mM of each deoxynucleoside triphosphate (dNTP), 0.6 uM of each primer, 1U GoTaq polymerase (Promega), 1 ul DNA template, and DEPC H2O to 25 ul. Thermocycling conditions on an Eppendorf thermal cycler consisted of an initial denaturation step at 94 degrees C for 3 min, followed by 35 cycles of 94 degrees C for 30s, 46 degrees C for 45 s, and 72 degrees C for 1 min, followed by a final extension at 72 degrees C for 5 min. The primers used in this study were sox527F (5'-TGGTWGGWCAYTGGGAATTTA-3') and sox1198R (5'-AGAANGTATCTCKYTTATAAAG-3'). These primers target the genera Sulfurovum, Sulfurimonas, and Nitratiruptor. The soxB gene in members of the Epsilonproteobacterial genera Arcobacter and Nitratifractor are more like sequences in Gammaproteobacteria and are not expected to amplify with this primer set. Successfully amplified soxB PCR products were cleaned with a Qiagen MinElute PCR purification kit and run on a 0.8% agarose gel. Bands in the expected size range were gel excised, purified with the MinElute kit, cloned, and sequenced as previously described (Huber et al., 2009).

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

Nucleotide sequences were translated into amino acids using EMBOSS Transeq (Rice et al., 2000) and phylogenetic relationships were analyzed using MEGA5 (Tamura et al., 2011). Sequences are deposited in GenBank under Accession numbers KC793341-KC793425.

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