Person: Sean P. Jungbluth

Carboxydotrophy potential of uncultivated Hydrothermarchaeota from the subseafloor crustal biosphere

The ISME Journal

Authors: Stephanie A. Carr, Sean P. Jungbluth, Emiley A. Eloe-Fadrosh, Ramunas Stepanauskas, Tanja Woyke, Michael S. Rappé, Beth N. Orcutt

Published: February 7, 2019

C-DEBI Contribution Number: 451

Abstract

The exploration of Earth’s terrestrial subsurface biosphere has led to the discovery of several new archaeal lineages of evolutionary significance. Similarly, the deep subseafloor crustal biosphere also harbors many unique, uncultured archaeal taxa, including those belonging to Candidatus Hydrothermarchaeota, formerly known as Marine Benthic Group-E. Recently, Hydrothermarchaeota was identified as an abundant lineage of Juan de Fuca Ridge flank crustal fluids, suggesting its adaptation to this extreme environment. Through the investigation of single-cell and metagenome-assembled genomes, we provide insight into the lineage’s evolutionary history and metabolic potential. Phylogenomic analysis reveals the Hydrothermarchaeota to be an early-branching archaeal phylum, branching between the superphylum DPANN, Euryarchaeota, and Asgard lineages. Hydrothermarchaeota genomes suggest a potential for dissimilative and assimilative carbon monoxide oxidation (carboxydotrophy), as well as sulfate and nitrate reduction. There is also a prevalence of chemotaxis and motility genes, indicating adaptive strategies for this nutrient-limited fluid-rock environment. These findings provide the first genomic interpretations of the Hydrothermarchaeota phylum and highlight the anoxic, hot, deep marine crustal biosphere as an important habitat for understanding the evolution of early life.

Source: http://dx.doi.org/10.1038/s41396-019-0352-9

Divergent methyl-coenzyme M reductase genes in a deep-subseafloor Archaeoglobi

The ISME Journal

Authors: Joel A. Boyd, Sean P. Jungbluth, Andy O. Leu, Paul N. Evans, Ben J. Woodcroft, Grayson L. Chadwick, Victoria J. Orphan, Jan P. Amend, Michael S. Rappé, Gene W. Tyson

Published: January 16, 2019

C-DEBI Contribution Number: 457

Abstract

The methyl-coenzyme M reductase (MCR) complex is a key enzyme in archaeal methane generation and has recently been proposed to also be involved in the oxidation of short-chain hydrocarbons including methane, butane, and potentially propane. The number of archaeal clades encoding the MCR continues to grow, suggesting that this complex was inherited from an ancient ancestor, or has undergone extensive horizontal gene transfer. Expanding the representation of MCR-encoding lineages through metagenomic approaches will help resolve the evolutionary history of this complex. Here, a near-complete Archaeoglobi metagenome-assembled genome (MAG; Ca. Polytropus marinifundus gen. nov. sp. nov.) was recovered from the deep subseafloor along the Juan de Fuca Ridge flank that encodes two divergent McrABG operons similar to those found in Ca. Bathyarchaeota and Ca. Syntrophoarchaeum MAGs. Ca. P. marinifundus is basal to members of the class Archaeoglobi, and encodes the genes for β-oxidation, potentially allowing an alkanotrophic metabolism similar to that proposed for Ca. Syntrophoarchaeum. Ca. P. marinifundus also encodes a respiratory electron transport chain that can potentially utilize nitrate, iron, and sulfur compounds as electron acceptors. Phylogenetic analysis suggests that the Ca. P. marinifundus MCR operons were horizontally transferred, changing our understanding of the evolution and distribution of this complex in the Archaea.

Source: http://dx.doi.org/10.1038/s41396-018-0343-2

Genomic comparisons of a bacterial lineage that inhabits both marine and terrestrial deep subsurface systems

PeerJ

Authors: Sean P. Jungbluth, Tijana Glavina del Rio, Susannah G. Tringe, Ramunas Stepanauskas, Michael S. Rappé

Published: April 6, 2017

C-DEBI Contribution Number: 356

Abstract

It is generally accepted that diverse, poorly characterized microorganisms reside deep within Earth’s crust. One such lineage of deep subsurface-dwelling bacteria is an uncultivated member of the Firmicutes phylum that can dominate molecular surveys from both marine and continental rock fracture fluids, sometimes forming the sole member of a single-species microbiome. Here, we reconstructed a genome from basalt-hosted fluids of the deep subseafloor along the eastern Juan de Fuca Ridge flank and used a phylogenomic analysis to show that, despite vast differences in geographic origin and habitat, it forms a monophyletic clade with the terrestrial deep subsurface genome of “Candidatus Desulforudis audaxviator” MP104C. While a limited number of differences were observed between the marine genome of “Candidatus Desulfopertinax cowenii” modA32 and its terrestrial relative that may be of potential adaptive importance, here it is revealed that the two are remarkably similar thermophiles possessing the genetic capacity for motility, sporulation, hydrogenotrophy, chemoorganotrophy, dissimilatory sulfate reduction, and the ability to fix inorganic carbon via the Wood-Ljungdahl pathway for chemoautotrophic growth. Our results provide insights into the genetic repertoire within marine and terrestrial members of a bacterial lineage that is widespread in the global deep subsurface biosphere, and provides a natural means to investigate adaptations specific to these two environments.

Source: http://dx.doi.org/10.7717/peerj.3134

Metagenome sequencing and 98 microbial genomes from Juan de Fuca Ridge flank subsurface fluids

Scientific Data

Authors: Sean P. Jungbluth, Jan P. Amend, Michael S. Rappé

Published: March 28, 2017

C-DEBI Contribution Number: 357

Abstract

The global deep subsurface biosphere is one of the largest reservoirs for microbial life on our planet. This study takes advantage of new sampling technologies and couples them with improvements to DNA sequencing and associated informatics tools to reconstruct the genomes of uncultivated Bacteria and Archaea from fluids collected deep within the Juan de Fuca Ridge subseafloor. Here, we generated two metagenomes from borehole observatories located 311 meters apart and, using binning tools, retrieved 98 genomes from metagenomes (GFMs). Of the GFMs, 31 were estimated to be >90% complete, while an additional 17 were >70% complete. Phylogenomic analysis revealed 53 bacterial and 45 archaeal GFMs, of which nearly all were distantly related to known cultivated isolates. In the GFMs, abundant Bacteria included Chloroflexi, Nitrospirae, Acetothermia (OP1), EM3, Aminicenantes (OP8), Gammaproteobacteria, and Deltaproteobacteria, while abundant Archaea included Archaeoglobi, Bathyarchaeota (MCG), and Marine Benthic Group E (MBG-E). These data are the first GFMs reconstructed from the deep basaltic subseafloor biosphere, and provide a dataset available for further interrogation.

Source: http://dx.doi.org/10.1038/sdata.2017.37

Viruses in the Oceanic Basement

mBio

Authors: Olivia Nigro, Sean P. Jungbluth, Huei-Ting Lin, Chih-Chiang Hsieh, Jaclyn A. Miranda, Christopher R. Schvarcz, Michael S. Rappé, Grieg Steward

Editors: Stephen J. Giovannoni

Published: March 7, 2017

C-DEBI Contribution Number: 354

Abstract

Microbial life has been detected well into the igneous crust of the seafloor (i.e., the oceanic basement), but there have been no reports confirming the presence of viruses in this habitat. To detect and characterize an ocean basement virome, geothermally heated fluid samples (ca. 60 to 65°C) were collected from 117 to 292 m deep into the ocean basement using seafloor observatories installed in two boreholes (Integrated Ocean Drilling Program [IODP] U1362A and U1362B) drilled in the eastern sediment-covered flank of the Juan de Fuca Ridge. Concentrations of virus-like particles in the fluid samples were on the order of 0.2 × 10⁵ to 2 × 10⁵ ml⁻¹ (n = 8), higher than prokaryote-like cells in the same samples by a factor of 9 on average (range, 1.5 to 27). Electron microscopy revealed diverse viral morphotypes similar to those of viruses known to infect bacteria and thermophilic archaea. An analysis of virus-like sequences in basement microbial metagenomes suggests that those from archaeon-infecting viruses were the most common (63 to 80%). Complete genomes of a putative archaeon-infecting virus and a prophage within an archaeal scaffold were identified among the assembled sequences, and sequence analysis suggests that they represent lineages divergent from known thermophilic viruses. Of the clustered regularly interspaced short palindromic repeat (CRISPR)-containing scaffolds in the metagenomes for which a taxonomy could be inferred (163 out of 737), 51 to 55% appeared to be archaeal and 45 to 49% appeared to be bacterial. These results imply that the warmed, highly altered fluids in deeply buried ocean basement harbor a distinct assemblage of novel viruses, including many that infect archaea, and that these viruses are active participants in the ecology of the basement microbiome.

IMPORTANCE The hydrothermally active ocean basement is voluminous and likely provided conditions critical to the origins of life, but the microbiology of this vast habitat is not well understood. Viruses in particular, although integral to the origins, evolution, and ecology of all life on earth, have never been documented in basement fluids. This report provides the first estimate of free virus particles (virions) within fluids circulating through the extrusive basalt of the seafloor and describes the morphological and genetic signatures of basement viruses. These data push the known geographical limits of the virosphere deep into the ocean basement and point to a wealth of novel viral diversity, exploration of which could shed light on the early evolution of viruses.

This paper is dedicated to the late James P. Cowen, whose collegiality, infectious enthusiasm for science, and pioneering studies of the ocean basement microbiome inspired the work.

Source: http://dx.doi.org/10.1128/mbio.02129-16

Datasets

Juan de Fuca Ridge Flank SSU rRNA gene sequences

Data Project: Microbiology and biogeochemistry of Juan de Fuca Ridge flank borehole fluids

Dataset Maintainers: Michael S. Rappé, Michael S. Rappé, Sean P. Jungbluth, Amber York, Michael S. Rappé, Michael S. Rappé

Data Project Maintainers: Michael S. Rappé

Last Modified: October 18, 2016

URL	https://www.bco-dmo.org/dataset/660489
Download URL	https://www.bco-dmo.org/dataset/660489/data/download
Media Type	text/tab-separated-values
Created	October 4, 2016
Modified	October 18, 2016
State	Final no updates expected
Brief Description	SSU 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 Res30: 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

Pump

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.

Spectrophotometer

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).

Automated DNA Sequencer

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).

Mass Spectrometer

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 Chromatograph

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

Shimadzu TOC-V Analyzer

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)

pH Sensor

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_desc

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

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.

depth

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

Altitude of sampler above the bottom

env_biome [site_descrip]

Details

env_biome

Biome of sample site

site_descrip

Description or characteristics of a sampling site

env_feature [site_descrip]

Details

env_feature

Environmental features of sample site

site_descrip

Description or characteristics of a sampling site

env_material [site_descrip]

Details

env_material

Environmental material of sample site

site_descrip

Description or characteristics of a sampling site

geo_loc_name [site]

Details

geo_loc_name

Geolocation name of sample site

site

Sampling site identification.

lat [latitude]

Details

lat

latitutde

latitude

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

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

oxygen [trace_element_conc]

Details

oxygen

oxygen (O2)

trace_element_conc

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

Periodic Table

calcium [trace_metal_conc]

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.

sodium [trace_metal_conc]

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

Chloride ion Cl-

bromide [bromides]

Details

bromide

bromide (Br-)

bromides

Bromides including Br-

silicate [silica]

Details

silicate

silicon dioxide (SiO2)

silica

Silicon Dioxide SiO2 (silica)

ammonium [Ammonium]

Details

ammonium

ammonium (NH4)

Ammonium

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

phosphate [PO4]

Details

phosphate

phosphate (PO4)

PO4

Orthophosphate (phosphate, reactive phosphorus), Various units.

nitrite [NO2]

Details

nitrite

nitrite (NO2)

NO2

Nitrite

nitrate [NO3]

Details

nitrate

nitrate (NO3)

NO3

Nitrate concentration in the water column

nitrate_and_nitrate [NO3_NO2]

Details

nitrate_and_nitrate

combined nitrate and nitrate (NO3 and NO2)

NO3_NO2

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

sulfate [SO4]

Details

sulfate

sulfate (SO4)

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.

Mn2plus [trace_metal_conc]

Details

Mn2plus

Manganese ion (Mn2+)

Total Dissolved Nitrogren

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)

sulfide

concentration of sulfide

dissolved_organic_carbon [DOC]

Details

dissolved_organic_carbon

dissolved organic carbon (DOC)

DOC

dissolved organic Carbon

TDN [Total Dissolved Nitrogren]

Details

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

TALK

Total alkalinity.

methane [CH4]

Details

methane

methane (CH4)

CH4

methane

hydrogen [hydrogen]

Details

hydrogen

hydrogen (H)

hydrogen

Hydrogen (H). Includes concentration and total weight.

microbial_cell_abundance [abundance]

Details

microbial_cell_abundance

microbial cell abundance

abundance

number of the specific taxa/group counted per unit area or volume

Dataset Maintainers

Name	Affiliation	Contact
Michael S. Rappé	University of Hawaii at Manoa (SOEST)
Michael S. Rappé	University of Hawaii at Manoa (SOEST)
Sean P. Jungbluth	University of Southern California (USC)	✓
Amber York	University of Southern California (USC)	✓
Michael S. Rappé	University of Hawaii at Manoa (SOEST)
Michael S. Rappé	University of Hawaii at Manoa (SOEST)

BCO-DMO Project Info

Project Title	Microbiology and biogeochemistry of Juan de Fuca Ridge flank borehole fluids
Acronym	microJdFR
URL	https://www.bco-dmo.org/project/635868
Created	January 21, 2016
Modified	December 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.

Data Project Maintainers

Name	Affiliation	Role
Michael S. Rappé	University of Hawaii at Manoa (HIMB)	Principal Investigator

Nanocalorimetric Characterization of Microbial Activity in Deep Subsurface Oceanic Crustal Fluids

Frontiers in Microbiology

Authors: Alberto Robador, Douglas E. LaRowe, Sean P. Jungbluth, Huei-Ting Lin, Michael S. Rappé, Kenneth H. Nealson, Jan P. Amend

Published: April 5, 2016

C-DEBI Contribution Number: 327

Abstract

Although fluids within the upper oceanic basaltic crust harbor a substantial fraction of the total prokaryotic cells on Earth, the energy needs of this microbial population are unknown. In this study, a nanocalorimeter (sensitivity down to 1.2 nW ml^-1) was used to measure the enthalpy of microbially catalyzed reactions as a function of temperature in samples from two distinct crustal fluid aquifers. Microorganisms in unamended, warm (63°C) and geochemically altered anoxic fluids taken from 292 meters sub-basement (msb) near the Juan de Fuca Ridge produced 267.3 mJ of heat over the course of 97 h during a step-wise isothermal scan from 35.5 to 85.0°C. Most of this heat signal likely stems from the germination of thermophilic endospores (6.66 × 10⁴ cells ml^-1_FLUID) and their subsequent metabolic activity at temperatures greater than 50°C. The average cellular energy consumption (5.68 pW cell^-1) reveals the high metabolic potential of a dormant community transported by fluids circulating through the ocean crust. By contrast, samples taken from 293 msb from cooler (3.8°C), relatively unaltered oxic fluids, produced 12.8 mJ of heat over the course of 14 h as temperature ramped from 34.8 to 43.0°C. Corresponding cell-specific energy turnover rates (0.18 pW cell^-1) were converted to oxygen uptake rates of 24.5 nmol O₂ ml^-1_FLUID d^-1, validating previous model predictions of microbial activity in this environment. Given that the investigated fluids are characteristic of expansive areas of the upper oceanic crust, the measured metabolic heat rates can be used to constrain boundaries of habitability and microbial activity in the oceanic crust.

Source: http://dx.doi.org/10.3389/fmicb.2016.00454

Novel microbial assemblages inhabiting crustal fluids within mid-ocean ridge flank subsurface basalt

The ISME Journal

Authors: Sean P. Jungbluth, Robert M. Bowers, Huei-Ting Lin, James P. Cowen, Michael S. Rappé

Published: February 12, 2016

C-DEBI Contribution Number: 289

Abstract

Although little is known regarding microbial life within our planet’s rock-hosted deep subseafloor biosphere, boreholes drilled through deep ocean sediment and into the underlying basaltic crust provide invaluable windows of access that have been used previously to document the presence of microorganisms within fluids percolating through the deep ocean crust. In this study, the analysis of 1.7 million small subunit ribosomal RNA genes amplified and sequenced from marine sediment, bottom seawater and basalt-hosted deep subseafloor fluids that span multiple years and locations on the Juan de Fuca Ridge flank was used to quantitatively delineate a subseafloor microbiome comprised of distinct bacteria and archaea. Hot, anoxic crustal fluids tapped by newly installed seafloor sampling observatories at boreholes U1362A and U1362B contained abundant bacterial lineages of phylogenetically unique Nitrospirae, Aminicenantes, Calescamantes and Chloroflexi. Although less abundant, the domain Archaea was dominated by unique, uncultivated lineages of marine benthic group E, the Terrestrial Hot Spring Crenarchaeotic Group, the Bathyarchaeota and relatives of cultivated, sulfate-reducing Archaeoglobi. Consistent with recent geochemical measurements and bioenergetic predictions, the potential importance of methane cycling and sulfate reduction were imprinted within the basalt-hosted deep subseafloor crustal fluid microbial community. This unique window of access to the deep ocean subsurface basement reveals a microbial landscape that exhibits previously undetected spatial heterogeneity.

Source: http://dx.doi.org/10.1038/ismej.2015.248

Activity and phylogenetic diversity of sulfate-reducing microorganisms in low-temperature subsurface fluids within the upper oceanic crust

Frontiers in Microbiology

Authors: Alberto Robador, Sean P. Jungbluth, Douglas E. LaRowe, Robert M. Bowers, Michael S. Rappé, Jan P. Amend, James P. Cowen

Published: January 14, 2015

C-DEBI Contribution Number: 249

Abstract

The basaltic ocean crust is the largest aquifer system on Earth, yet the rates of biological activity in this environment are unknown. Low-temperature (<100°C) fluid samples were investigated from two borehole observatories in the Juan de Fuca Ridge (JFR) flank, representing a range of upper oceanic basement thermal and geochemical properties. Microbial sulfate reduction rates (SRR) were measured in laboratory incubations with ³⁵S-sulfate over a range of temperatures and the identity of the corresponding sulfate-reducing microorganisms (SRM) was studied by analyzing the sequence diversity of the functional marker dissimilatory (bi)sulfite reductase (dsrAB) gene. We found that microbial sulfate reduction was limited by the decreasing availability of organic electron donors in higher temperature, more altered fluids. Thermodynamic calculations indicate energetic constraints for metabolism, which together with relatively higher cell-specific SRR reveal increased maintenance requirements, consistent with novel species-level dsrAB phylotypes of thermophilic SRM. Our estimates suggest that microbially-mediated sulfate reduction may account for the removal of organic matter in fluids within the upper oceanic crust and underscore the potential quantitative impact of microbial processes in deep subsurface marine crustal fluids on marine and global biogeochemical carbon cycling.

Source: http://dx.doi.org/10.3389/fmicb.2014.00748

Phylogenetic diversity of microorganisms in subseafloor crustal fluids from Holes 1025C and 1026B along the Juan de Fuca Ridge flank

Frontiers in Microbiology

Authors: Sean P. Jungbluth, Huei-Ting Lin, James P. Cowen, Brian T. Glazer, Michael S. Rappé

Published: March 25, 2014

C-DEBI Contribution Number: 244

Abstract

To expand investigations into the phylogenetic diversity of microorganisms inhabiting the subseafloor biosphere, basalt-hosted crustal fluids were sampled from Circulation Obviation Retrofit Kits (CORKs) affixed to Holes 1025C and 1026B along the Juan de Fuca Ridge (JdFR) flank using a clean fluid pumping system. These boreholes penetrate the crustal aquifer of young ocean crust (1.24 and 3.51 million years old, respectively), but differ with respect to borehole depth and temperature at the sediment-basement interface (147 m and 39°C vs. 295 m and 64°C, respectively). Cloning and sequencing of PCR-amplified small subunit ribosomal RNA genes revealed that fluids retrieved from Hole 1025C were dominated by relatives of the genus Desulfobulbus of the Deltaproteobacteria (56% of clones) and Candidatus Desulforudis of the Firmicutes (17%). Fluids sampled from Hole 1026B also contained plausible deep subseafloor inhabitants amongst the most abundant clone lineages; however, both geochemical analysis and microbial community structure reveal the borehole to be compromised by bottom seawater intrusion. Regardless, this study provides independent support for previous observations seeking to identify phylogenetic groups of microorganisms common to the deep ocean crustal biosphere, and extends previous observations by identifying additional lineages that may be prevalent in this unique environment.

Source: http://dx.doi.org/10.3389/fmicb.2014.00119

Dissolved hydrogen and methane in the oceanic basaltic biosphere

Earth and Planetary Science Letters

Authors: Huei-Ting Lin, James P. Cowen, Eric J. Olson, Marvin D. Lilley, Sean P. Jungbluth, Samuel T. Wilson, Michael S. Rappé

Published: November 1, 2014

C-DEBI Contribution Number: 226

Abstract

The oceanic basaltic crust is the largest aquifer on Earth and has the potential to harbor substantial subsurface microbial ecosystems, which hitherto remains largely uncharacterized and is analogous to extraterrestrial subsurface habitats. Within the sediment-buried 3.5 Myr old basaltic crust of the eastern Juan de Fuca Ridge flank, the circulating basement fluids have moderate temperature (∼65 °C) and low to undetectable dissolved oxygen and nitrate concentrations. Sulfate, present in high concentrations, is therefore expected to serve as the major electron acceptor in this subsurface environment. This study focused on the availability and potential sources of two important electron donors, methane (CH₄) and hydrogen (H₂), for the subseafloor biosphere. High integrity basement fluids were collected via fluid delivery lines associated with Integrated Ocean Drilling Program (IODP) Circulation Obviation Retrofit Kits (CORKs) that extend from basement depths to outlet ports at the seafloor. Two new CORKs installed during IODP 327 in 2010, 1362A and 1362B, were sampled in 2011 and 2013. The two CORKs are superior than earlier style CORKs in that they are equipped with coated casing and polytetrafluoroethylene fluid delivery lines, reducing the interaction between casing materials with the environment. Additional samples were collected from an earlier style CORK at Borehole 1301A.

The basement fluids are enriched in H₂ (0.05–1.8 μmol/kg), suggesting that the ocean basaltic aquifer can support H₂-driven metabolism. The basement fluids also contain significant amount of CH₄ (5–32 μmol/kg), revealing CH₄ as an available substrate for subseafloor basaltic habitats. The δ¹³C values of CH₄ from the three boreholes ranged from −22.5 to −58‰, while the δ²H values ranged from −316 to 57‰. The isotopic compositions of CH₄ and the molecular compositions of hydrocarbons suggest that CH₄ in the basement fluids is of both biogenic and abiotic origins, varying among sites and sampling times. The δ²H values of CH₄ in CORK 1301A fluid samples are much more positive than found in all other marine environments investigated to date and are best explained by the partial microbial oxidation of biogenic CH₄. In conclusion, our study shows that CH₄ and H₂ are persistently available to fuel the deep biosphere and that CH₄ is both produced and potentially consumed by microorganisms in the oceanic basement.

Source: http://dx.doi.org/10.1016/j.epsl.2014.07.037

Publications > Book Chapter

Microbial Life of the Deep Biosphere

2. Life in the Oceanic Crust

Authors: Jennifer F. Biddle, Sean P. Jungbluth, Mark A. Lever, Michael S. Rappé

Editors: Jens Kallmeyer

Published: March 31, 2014

C-DEBI Contribution Number: 196

Abstract

The detection of life in extreme environments and the amazing capacity of microbes to obtain energy from the environment has led to the hypothesis that heat, not water or energy sources, is the final barrier that constrains life on Earth. As such, life is expected to extend far into Earth’s crust, potentially inhabiting an environment as extensive as the world’s oceans [1]. This chapter aims to summarize what is currently known about life in the oceanic crust, which contains all three domains of life and is a vastly underexplored habitat. This is not the first summary of this topic, and additional content can be found in numerous review and primary research papers [2–16], book chapters [17, 18], and books [19–21].

Source: http://dx.doi.org/10.1515/9783110300130.29

Publications > Monograph

Proceedings of the IODP

Data report: microbial diversity in sediment near Grizzly Bare Seamount in Holes U1363B and U1363G

Authors: Sean P. Jungbluth, L. G. H. Johnson, James P. Cowen, Michael S. Rappé

Published: September 5, 2011

C-DEBI Contribution Number: 181

Abstract

Microorganisms inhabiting sediment in close proximity to recharging basement outcrops are of interest because of the enhanced advective fluid flow in these locations, which is expected to exert unique selective pressures on the resident microbial communities. Here, Integrated Ocean Drilling Program (IODP) boreholes were used to access sediment microbial communities near Grizzly Bare recharge seamount on the Juan de Fuca Ridge eastern flank. The two locations examined in this study, Holes U1363G and U1363B, are 50 and 177 m away from the center of the outcrop, respectively. In general, small subunit ribosomal RNA gene clones from all three domains of life were detected; these groups were predominantly related to microorganisms known to reside in marine sediment. A large fraction of environmental gene clones recovered from Hole U1363B and U1363G sediment were related to uncultivated, candidate phyla of Bacteria such as BHI80-139, BRC1, JS1, OPB41, and TA06. Hole U1363B and U1363G sediment clone libraries were generally dominated by the domain Bacteria and particularly the phylum Chloroflexi, which comprised approximately one-quarter of the total gene clones identified. However, borehole sediment also contained several archaeal lineages that were phylogenetically affiliated with the Miscellanenous Crenarchaeotal Group. Eukaryotic fungi were only detected within the interstitial water sample from Hole U1363B. Finally, a minor portion of clones recovered from sediment in this study were also recovered from basement fluid samples previously characterized from Baby Bare discharge seamount and Ocean Drilling Program and IODP borehole Circulation Obviation Retrofit Kit Observatories (CORKs) in Holes 1026B and U1301A, which are ~50 km to the north-northeast.

Source: http://dx.doi.org/10.2204/iodp.proc.327.201.2013