URLhttps://www.bco-dmo.org/dataset/660489
Download URLhttps://www.bco-dmo.org/dataset/660489/data/download
Media Typetext/tab-separated-values
CreatedOctober 4, 2016
ModifiedOctober 18, 2016
StateFinal no updates expected
Brief DescriptionSSU rRNA gene sequences from marine sediments, marine subseafloor, and deep seawater

Acquisition Description

Sampling Methodology:

Sediment coring was performed by the IODP (described in Expedition 327 Scientists, 2011; Integrated Ocean Drilling Program). SSU rRNA genes were obtained as described in Jungbluth et al., 2013; Proceedings of the Integrated Ocean Drilling Program (see Supplementary Information Document(.DOC) for this publication).

CORK borehole fluids were sampled using a custom-built water sampler (described in Cowen et al., 2012). For a detailed description of the pump system see Lin, et al. 2012.

Pore water dissolved organic carbon (from Lin et al., 2015 Materials and Methods ).

Sedimentary pore water DOC concentrations were measured by high-temperature combustion using a Shimadzu TOC-VCSH analyzer. The combustion temperature was set at 720C to ensure complete oxidation of organic matter. Samples were acidified to pH <2 by the addition of 45 uL of 2 M HCl to 3 mL samples. No acid contamination was observed based on monitoring the DOC value of low-carbon deionized water. Samples were purged with nitrogen gas within the autosampler syringe for 2 min in order to remove inorganic carbon. An injection volume of 150 uL was used, with five or six injections per sample. The reproducibility between replicate injections was <1 uM. Analytical reference materials (ARM) supplied by Dr. Dennis Hansell (RSMAS, University of Miami) were measured before, between, and after analysis of environmental samples (Sharp et al., 2002; Dickson et al., 2007). At least one ARM was measured every five samples. The average measured concentration of the ARM was 42 plus or minus 2 uM (n = 44); the reported value was 41–43 uM. Our detection limit for DOC concentrations was ~2 uM.

Sediment organic carbon and nitrogen  (relevant text extracted from Lin et al., 2015 Materials and Methods ).

Whole sediment samples were analyzed for concentration of total carbon, organic carbon, and total nitrogen using an elemental combustion system (Costech ECS 4010) connected inline to an isotope-ratio mass spectrometer (Thermo Finnigan Delta XP). The amount of powdered sediment used for the analyses was optimized to provide sufficient carbon and nitrogen for isotopic composition analysis and varied between 26 and 425 mg. A subset of samples was acidified by fuming with concentrated HCl (Hedges and Stern, 1984) in order to remove inorganic carbon and quantify the particulate organic carbon (POC) content. Acid fuming did not remove inorganic nitrogen, resulting in insignificant differences between whole and acid-fumed total particulate nitrogen (PN) concentrations.

Analytical methods for geochemistry

Text below extracted from Supplementary Information (PDF) Junbluth et al., 2016.  See reference for full description.

Major ions (Ca2+, Mg2+, K+ , Na+, Cl- , SO4 2- and Br- ) were analyzed by ion chromatography on a Dionex ICS-1100s (Sunnyvale, CA, USA). In addition, magnesium and calcium concentrations were also analyzed by EDTA (colorimetric) and EGTA (electrometric) titration (Grasshoff et al., 1999), or inductively coupled plasma optical emission spectroscopy (ICP-OES) (Lin et al., 2012). 

Silicate, nitrate, nitrite, phosphate, dissolved sulfide and dissolved manganese concentrations were measured by colorimetry (Brewer and Spencer, 1971; Phillips et al., 1997; Grasshoff et al., 1999). 

 Ammonium concentrations were measured by a flow injection-fluorometric method (Jones, 1991). The detection limit was ~2 µM for ammonium in basement fluids and the analytical uncertainty is 0.5 µM.

Ferrous iron was measured directly by a Ferrozine colorimetry method (Stookey, 1970; Gibbs, 1976).

For total iron analysis, samples were first reduced with ascorbic acid and analyzed as ferrous iron. The detection limit for both ferrous iron and total iron was 0.1 µM.

Dissolved organic carbon (DOC) was measured by high-temperature combustion using a TOC-VCSH analyzer (Sharp et al., 2002a; Dickson et al., 2007) (Shimadzu Corp., Kyoto, Japan). 

Total dissolved nitrogen (TDN) was measured with a chemiluminescence detector in-line with a Shimadzu TOC-VCSH analyzer (Sharp et al., 2002b).

Alkalinity was determined by acid titration. Acid (0.1N HCl) was standardized with CO2 certified reference materials (CRMs) purchased from the office of Andrew Dickson at Scripps Institution of Oceanography. 

An Orion 911600 Semi-micro pH electrode (ThermoFisher Scientific, Waltham, MA, USA) was used to measure the pH and electrode potential during the titration process. The Gran function plot method was used to evaluate titration end-points and calculate sample alkalinity (Dickson et al., 2007). The analytical reproducibility for alkalinity measurements was <0.02 mM.

SSU rRNA gene cloning and sequencing (from Supplementary Information Document(DOC) for Junbluth et al., 2013).

Small subunit ribosomal RNA (SSU rRNA) gene fragments were amplified via the polymerase chain reaction (PCR) using the universal oligonucleotide forward and reverse primers 519F (5’-CAGCMGCCGCGGTAATWC-3’) and 1406R (5’-ACGGGCGGTGTGTRC-3’), respectively. Each 20 ul PCR reaction contained 0.25 U of PicoMaxx high fidelity DNA polymerase (Stratagene, La Jolla, CA), 1x PicoMaxx reaction buffer, 200 uM of each of the four deoxynucleoside triphosphates (dNTPs), 200 nM of both forward and reverse primer, and ~3-4 ng of environmental DNA template. PCR cycling conditions consisted of an initial denaturation step at 95C for 4 minutes, followed by 35 to 38 cycles of 95C denaturation for 30 sec, 55C annealing for 1 min, 72C extension for 2 min, and a final extension step at 72C for 20 min. For the 2008 borehole fluid sample, a 3-cycle reconditioning PCR was performed in order to help eliminate heteroduplexes (Thompson et al., 2002). Amplification products of the anticipated length were excised from an agarose gel and subsequently purified using the QIAquick gel extraction kit (Qiagen, Valencia, CA). Products were cloned using either the pGEM-T Easy kit (Promega, Madison, WI) or the TOPO TA Cloning kit (Invitrogen, Carlsbad, CA) following the manufacturer’s instructions. Clones were sequenced unidirectionally on an ABI 3730XL DNA Analyzer (Applied Biosystems, Carlsbad, CA). 

Fluorescence Microscopy: microbial cell counts  (from Supplementary Information Document(DOC) for Junbluth et al., 2016).

Sample preparation for microscopy and fluorescence microscopy Fluid samples for microscopy collected in 2011 were prepared in similar fashion to those collected in sampling years 2008-2010 and described previously (Jungbluth et al., 2013). Briefly, 40 to 120 ml sub-samples were fixed with a final concentration of 3% of 0.2 um-filtered formaldehyde for 2 to 4 hours at 4C, and subsequently filtered through 0.2 um pore-sized polycarbonate membranes (Whatman, Maidstone, United Kingdom). After air-drying, membranes were stored desiccated at -80ºC until microscopic analysis. Filter sections were prepared for fluorescence microscopy using a mix of Citifluor/VectaShield/PBS/DAPI as described previously (Jungbluth et al., 2013a). Stained filter sections were inspected with a Leica DM5000B epifluorescence microscope (Leica Microsystems, Wetzlar, Germany) (samples: SSF1-2, SSF4, MIX1-4, SW1-5, SW9-11, SW14-15) or an Eclipse 90i (Nikon Corp., Tokyo, Japan) epifluorescence microscope (all other samples). Both microscopes were equipped with 100x objectives and filter sets appropriate for DAPI fluorescence.

References:

Brewer P, Spencer D (1971). Colorimetric determination of manganese in anoxic waters. Limnol Oceanogr 16: 107-110. doi: 10.4319/lo.1971.16.1.0107

Cowen, James P., et al. "Advanced instrument system for real-time and time-series microbial geochemical sampling of the deep (basaltic) crustal biosphere." Deep Sea Research Part I: Oceanographic Research Papers 61 (2012): 43-56.  http://dx.doi.org/10.1016/j.dsr.2011.11.004

Dickson AG, Sabine CL, Christian JR (eds) (2007). Guide to best practices for ocean CO2 measurements, 191pp. (URL: http://cdiac.ornl.gov/oceans/Handbook_2007.html)

Expedition 327 Scientists, 2011. Methods. In Fisher, A.T., Tsuji, T., Petronotis, K., and the Expedition 327 Scientists, Proc. IODP, 327: Tokyo (Integrated Ocean Drilling Program Management International, Inc.).

Gibbs C (1976). Characterization and application of ferrozine iron reagent as a ferrous iron indicator. Anal Chem 48: 1197-1201. doi: 10.1021/ac50002a034

Grasshoff K, Kremling K, Ehrhardt M (eds) (1999) Methods of seawater analysis. Wiley: Weinheim. doi: 10.1002/9783527613984

Hedges, J.I., and Stern, J.H., 1984. Carbon and nitrogen determinations of carbonate-containing solids. Limnol. Oceanogr., 29(3):657–663.doi:10.4319/​lo.1984.29.3.0657

Jones RD (1991). An improved fluorescence method for the determination of nanomolar concentrations of ammonium in natural-waters. Limnol Oceanogr 36: 814-819. doi: 10.4319/lo.1991.36.4.0814

Jungbluth, Sean P., et al. "Data report: microbial diversity in sediment near Grizzly Bare Seamount in Holes U1363B and U1363G." Proc. IODP Volume. Vol. 327. 2013. http://dx.doi.org/10.2204/iodp.proc.327.201.2013

Jungbluth, Sean P., et al. "Novel microbial assemblages inhabiting crustal fluids within mid-ocean ridge flank subsurface basalt." The ISME journal (2016). dx.doi.org/10.1038/ismej.2015.248

Lin, H-T, Hsieh, C-C, Cowen, JP, Rappe, MS (2015). Data report: dissolved and particulate organic carbon in the deep sediments of IODP Site U1363 near Grizzly Bare seamount. Proceedings of the Integrated Ocean Drilling Program 327: 1-16.  dx.doi.org/10.2204/iodp.proc.327.202.2015

Lin, Huei-Ting, et al. "Inorganic chemistry, gas compositions and dissolved organic carbon in fluids from sedimented young basaltic crust on the Juan de Fuca Ridge flanks." Geochimica et Cosmochimica Acta 85 (2012): 213-227. http://dx.doi.org/10.1016/j.gca.2012.02.017

Phillips BM, Anderson BS, Hunt JW (1997). Measurement and distribution of interstitial and overlying water ammonia and hydrogen sulfide in sediment toxicity tests. Mar Environ Res 44: 117-126. doi: 10.1016/S0141-1136(96)00087-6

Sharp JH, Carlson CA, Peltzer ET, Castle-Ward DM, Savidge KB, Rinker KR (2002a). Final dissolved organic carbon broad community intercalibration and preliminary use of DOC reference materials. Mar Chem 77: 239-253. doi: 10.1016/S0304- 4203(02)00002-6

Sharp JH, Rinker KR, Savidge KB, Abell J, Yves Benaim J, Bronk DA et al. (2002b). A preliminary methods comparison for measurement of dissolved organic nitrogen in seawater. Mar Chem 78: 171-184. doi: 10.1016/S0304-4203(02)00020-8

Stookey LL (1970). Ferrozine- a new spectrophotometric reagent for iron. Anal Chem 42: 779-781. doi: 10.1021/ac60289a016

Thompson JR, Marcelino LA, Polz MF. (2002). Heteroduplexes in mixed-template amplifications: formation, consequence and elimination by 'reconditioning PCR'. Nucleic Acids Res 30: 2083-2088.

Processing Description

These data have been quality controlled as described in Jungbluth et al., 2013, and Jungbluth et al., 2016.

For seawater samples, elevation was set equal to sea-level (value: 0) and all sample depths are reported as positive values.
For sediment and borehole samples, elevation was set equal to the depth of the seafloor (all values negative) and depths into the seafloor are reported as positive values.

Instruments

pump [Pump]
Details
Instance Description (pump)

A custom pump system was used to sample basement fluid. It connected to the CORK observatorys' fluid delivery lines.  The design of this system changed over the course of the study.  It included the pump components Pelagic Electronics 5010 series deep sea pump, and Sea-Bird SBE-5T submersible titanium pump.  For a detailed description see:

Lin, Huei-Ting, et al. "Inorganic chemistry, gas compositions and dissolved organic carbon in fluids from sedimented young basaltic crust on the Juan de Fuca Ridge flanks." Geochimica et Cosmochimica Acta 85 (2012): 213-227. http://dx.doi.org/10.1016/j.gca.2012.02.017

A pump is a device that moves fluids (liquids or gases), or sometimes slurries, by mechanical action. Pumps can be classified into three major groups according to the method they use to move the fluid: direct lift, displacement, and gravity pumps
NanoDrop ND-1000 spectrophotometer [Spectrophotometer]
Details
Instance Description (NanoDrop ND-1000 spectrophotometer)

NanoDrop ND-1000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA) was used to quantify resulting genomic DNA.

An instrument used to measure the relative absorption of electromagnetic radiation of different wavelengths in the near infra-red, visible and ultraviolet wavebands by samples.

ABI 3730XL DNA Analyzer [Automated DNA Sequencer]
Details
Instance Description (ABI 3730XL DNA Analyzer)

Clones were sequenced unidirectionally on an ABI 3730XL DNA Analyzer (Applied Biosystems, Carlsbad, CA).

General term for a laboratory instrument used for deciphering the order of bases in a strand of DNA. Sanger sequencers detect fluorescence from different dyes that are used to identify the A, C, G, and T extension reactions. Contemporary or Pyrosequencer methods are based on detecting the activity of DNA polymerase (a DNA synthesizing enzyme) with another chemoluminescent enzyme. Essentially, the method allows sequencing of a single strand of DNA by synthesizing the complementary strand along it, one base pair at a time, and detecting which base was actually added at each step.

Thermo Finnigan Delta XP [Mass Spectrometer]
Details
Instance Description (Thermo Finnigan Delta XP)

Whole sediment samples were analyzed for concentration and isotopic composition of total carbon, organic carbon, and total nitrogen using an elemental combustion system (Costech ECS 4010) connected inline to an isotope-ratio mass spectrometer (Thermo Finnigan Delta XP).

General term for instruments used to measure the mass-to-charge ratio of ions; generally used to find the composition of a sample by generating a mass spectrum representing the masses of sample components.
Dionex ICS-1100s [Ion Chromatograph]
Details

Ion chromatography is a form of liquid chromatography that measures concentrations of ionic species by separating them based on their interaction with a resin. Ionic species separate differently depending on species type and size. Ion chromatographs are able to measure concentrations of major anions, such as fluoride, chloride, nitrate, nitrite, and sulfate, as well as major cations such as lithium, sodium, ammonium, potassium, calcium, and magnesium in the parts-per-billion (ppb) range. (from http://serc.carleton.edu/microbelife/research_methods/biogeochemical/ic.html)

TOC-VCSH analyzer [Shimadzu TOC-V Analyzer]
Details
A Shimadzu TOC-V Analyzer measures DOC by high temperature combustion method.
Orion 911600 Semi-micro pH electrode [pH Sensor]
Details
Instance Description (Orion 911600 Semi-micro pH electrode)

 Orion 911600 Semi-micro pH electrode (ThermoFisher Scientific, Waltham, MA, USA) 

General term for an instrument that measures the pH or how acidic or basic a solution is.

Parameters

database [brief_desc]
Details
database

Database to which the accession_id belongs to

brief description, open ended, specific to the data set in which it appears

Accession_id [accession number]
Details
Accession_id
Identification number for GenBank; SRA; or IMG databases
Database identifier assigned by repository and linked to GenBank or other repository.
Accession_Link [accession number]
Details
Accession_Link
URL link to the accession for GenBank; SRA; or IMG
Database identifier assigned by repository and linked to GenBank or other repository.
BioSample_ID [accession number]
Details
BioSample_ID
Identifier for BioSample at NCBI. A BioSample corresponds to descriptions of biological source materials used in experimental assays.
Database identifier assigned by repository and linked to GenBank or other repository.
BioProjectID [accession number]
Details
BioProjectID
Identifier for BioProject at NCBI. A BioProject is a collection of biological data related to a single initiative originating from a single organization or from a consortium.
Database identifier assigned by repository and linked to GenBank or other repository.
description [sample_descrip]
Details
description
Description of sample and source material origin
text description of sample collected
sample_title [sample_descrip]
Details
sample_title
Project-specific sample title
text description of sample collected
sample_name [sample_descrip]
Details
sample_name
Descriptive sample title
text description of sample collected
organism [sample_descrip]
Details
organism
Type of organism(s) sampled
text description of sample collected
collection_date [date]
Details
collection_date

Date of sample collection in format dd-mmm-yy.

date; generally reported in GMT as YYYYMMDD (year; month; day); also as MMDD (month; day); EqPac dates are local Hawaii time. ISO_Date format is YYYY-MM-DD (http://www.iso.org/iso/home/standards/iso8601.htm)

depth [depth]
Details
depth

Depth of sample. Sea-water sample depths are reported as positive values. For sediment and borehole samples, elevation was set equal to the depth of the seafloor (all values negative) and depths into the seafloor are reported as positive values.

Observation/sample depth below the sea surface. Units often reported as: meters, feet.


When used in a JGOFS/GLOBEC dataset the depth is a best estimate; usually but not always calculated from pressure; calculated either from CTD pressure using Fofonoff and Millard (1982; UNESCO Tech Paper #44) algorithm adjusted for 1980 equation of state for seawater (EOS80) or simply equivalent to nominal depth as recorded during sampling if CTD pressure was unavailable.

elev [altitude]
Details
elev

Elevation of sample.  Sea-water samples are set to sea-level (0) . For sediment and borehole samples, elevation was set equal to the depth of the seafloor (all values negative).

Altitude of sampler above the bottom

env_biome [site_descrip]
Details
env_biome
Biome of sample site

Description or characteristics of a sampling site

env_feature [site_descrip]
Details
env_feature
Environmental features of sample site

Description or characteristics of a sampling site

env_material [site_descrip]
Details
env_material
Environmental material of sample site

Description or characteristics of a sampling site

geo_loc_name [site]
Details
geo_loc_name
Geolocation name of sample site
Sampling site identification.
lat [latitude]
Details
lat
latitutde

latitude, in decimal degrees, North is positive, negative denotes South; Reported in some datasets as degrees, minutes

lon [longitude]
Details
lon
longitude; west is negative

longitude, in decimal degrees, East is positive, negative denotes West; Reported in some datsets as degrees, minutes

ph [pH]
Details
ph
pH

pH: The measure of the acidity or basicity of an aqueous solution

Details
oxygen

oxygen (O2)

Concentration of a trace element in seawater. Does not include trace metals and metaloids.  Refer to dataset for element name and units.

Periodic Table

Details
calcium

calcium (Ca)

Concentration of a trace metal in seawater. Refer to dataset for element name and units.
magnesium [trace_metal_conc]
Details
magnesium

magnesium (Mg)

Concentration of a trace metal in seawater. Refer to dataset for element name and units.
potassium [trace_metal_conc]
Details
potassium

potassium (K)

Concentration of a trace metal in seawater. Refer to dataset for element name and units.
Details
sodium

sodium (Na)

Concentration of a trace metal in seawater. Refer to dataset for element name and units.
chloride [chloride]
Details
chloride

chloride (Cl-)

Chloride ion Cl-

bromide [bromides]
Details
bromide

bromide (Br-)

Bromides including Br-

silicate [silica]
Details
silicate

silicon dioxide (SiO2)

Silicon Dioxide SiO2 (silica)

ammonium [Ammonium]
Details
ammonium

ammonium (NH4)

Ammonium and ammonia concentration parameters in any body of fresh or salt water.

phosphate [PO4]
Details
phosphate

phosphate (PO4)

Orthophosphate (phosphate, reactive phosphorus), Various units. 

nitrite [NO2]
Details
nitrite

nitrite (NO2)

Nitrite

nitrate [NO3]
Details
nitrate

nitrate (NO3)

Nitrate concentration in the water column

nitrate_and_nitrate [NO3_NO2]
Details
nitrate_and_nitrate

combined nitrate and nitrate (NO3 and NO2)

Nitrate and Nitrite, µM (micromolar) or µg-at NO3-N and NO2-N/l

sulfate [SO4]
Details
sulfate

sulfate (SO4)

Concentration of sulfate (SO4) per unit volume

dissolved_iron [trace_metal_conc]
Details
dissolved_iron

dissolved iron (dFe)

Concentration of a trace metal in seawater. Refer to dataset for element name and units.
total_iron [trace_metal_conc]
Details
total_iron

total iron (Fe)

Concentration of a trace metal in seawater. Refer to dataset for element name and units.
Details
Mn2plus

Manganese ion (Mn2+)

Concentration of a trace metal in seawater. Refer to dataset for element name and units.
dissolved_hydrogen_sulfide [sulfide]
Details
dissolved_hydrogen_sulfide

dissolved hydrogen sulfide (dissolved H2S)

concentration of sulfide

dissolved_organic_carbon [DOC]
Details
dissolved_organic_carbon

dissolved organic carbon (DOC)

dissolved organic Carbon
TDN

total dissolved nitrogen (TDN)

The concentration of nitrogen remaining in a seawater sample after all particulate nitrogen has been removed by filtration. See dataset for units (may be micromoles/kilogram or micromolar).

alkalinity [TALK]
Details
alkalinity
alkalinity

Total alkalinity per unit mass of the water body.

methane [CH4]
Details
methane

methane (CH4)

methane

hydrogen [hydrogen]
Details
hydrogen

hydrogen (H)

Hydrogen (H). Includes concentration and total weight.

microbial_cell_abundance [abundance]
Details
microbial_cell_abundance
microbial cell abundance
number of the specific taxa/group counted per unit area or volume

Dataset Maintainers

NameAffiliationContact
Michael S. RappéUniversity of Hawaii at Manoa (SOEST)
Michael S. RappéUniversity of Hawaii at Manoa (SOEST)
Sean P. JungbluthUniversity of Southern California (USC)
Amber YorkWoods Hole Oceanographic Institution (WHOI BCO-DMO)

BCO-DMO Project Info

Project TitleMicrobiology and biogeochemistry of Juan de Fuca Ridge flank borehole fluids
AcronymmicroJdFR
URLhttps://www.bco-dmo.org/project/635868
CreatedJanuary 21, 2016
ModifiedDecember 15, 2016
Project Description

Extracted from the NSF award abstract:

Hydrothermally heated fluids circulate everywhere within the permeable basement rock of the upper ocean crust, providing warm temperatures and chemical gradients that support a deep subsurface marine biosphere. The volume of oceanic lithosphere habitable by microbial life is thought to be a substantial portion of the Earth’s crust - extending thousands of meters below the seafloor. During expeditions from 2008 to 2014 we repeatedly sampled basalt-hosted, deep subseafloor crustal fluids from four different boreholes drilled along the Juan de Fuca Ridge flank in the Northeast Pacific Ocean using pumps and samplers capable of collecting whole water and filtered particulates in situ. The instrumented boreholes, sitting at 2600 m depth, penetrate ~100 to 260 m of bottom sediments and another ~48 to 300 m of igneous basement where they tap into hot (up to 65 degrees C), anoxic fluid within Earth’s largest deep subsurface aquifer. Nearby bottom seawater and sediments were sampled as controls. Associated data sets include small subunit ribosomal RNA and functional gene amplicon DNA sequences, metagenome sequences, single cell genome sequences, direct counts of microbial cells and viruses, and a wide range of associated biogeochemical measurements including dissolved gases, particulate and dissolved organic carbon, sulfate, nitrate, and others.

Project Maintainers
NameAffiliationRoleContact
Michael S. RappéUniversity of Hawaii at Manoa (HIMB)Principal Investigator
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