Biogeochemical Exhalations: Microbial Methane in Marine Sediments

1
Biogeochemical Exhalations Microbial Methane in
Marine Sediments

• Johnson State College, Johnson, VT
• November 11, 2009
• Rick Colwell
• College of Oceanic and Atmospheric
  Sciences, Oregon State University
• Collaborators
• B. Briggs (Oregon State Univ.)
• M. Delwiche, D. Reed, D. Newby (Idaho National
  Lab)
• S. Boyd (University of Idaho)
• W. Ussler (Monterey Bay Aquarium Research Inst.)
• G. Dickens (Rice University)
• F. Inagaki, Y. Morono (JAMSTEC)
• Acknowledgements U.S. Department of Energy,
  Office of Fossil Energy Integrated Ocean
  Drilling Program, Science Parties from Leg 204
  Ocean Leadership

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Outline

• The methane issue - How much? Where is it coming
  from? Do we care?
• Deep biosphere and subseafloor microbiology
• In situ rates of methane production and a
  strategy for deriving these rates
• Experimentally derived rates connected to models
• Opportunities

3
Methane emission to the atmosphere
500 Tg methane/yr 60x less than the
mass of CO2 added to the
atmosphere annually due to human
activities
(Schrag, 2009) 84 from living
biological sources (yellow and orange) 87 of
which is from microbes
(yellow)
80
69 is a consequence of human activities (red) 1
of the leakage comes from methane hydrates
Data compiled by Reeburgh, 2007 Redrawn by
Colwell and Ussler, 2009
4
Methane hydrates

• Stable by virtue of temperature and pressure
• Milkov et al. 2004 estimated 500-2500 Gt of CH4-C
  in hydrates
• (ca. 6.7-33.3 x 105 Tg CH4)
• Most hydrate methane is biogenic (Kvenvolden and
  Lorenson, 2001)

Collett
/DOE
T.
T.
Collett
/DOE
5
Modified from Judd and Hovland, 2007
6
Methane release and the carbon cycle
Figure from Weissert, 2000
7
Basic steady-state view of the global carbon cycle

• Amount, distribution, and behavior of gas
  hydrates should be studied as dynamic processes
• Accurate estimates to come from temporal modeling
  of CH4 inputs and outputs in appropriate hydrate
  environments
• CH4 inputs are poorly understood

Dickens, 2003
8
Some basic questions about subseafloor
microbiologyHow many microbes survive at
depth?What types of microbes are
present?Where are they?How active are they?
DHondt et al. 2009
9
How many subsurface microbes are there?

• Biomass (Whitman et al, 1998)
• Total prokaryotes on earth 4-6 x 1030 cells
• up to 94 of all prokaryotes in the subsurface
• 0.25-2.5 x 1030 cells in terrestrial subsurface
• 3.5 x 1030 cells in subseafloor
• Extreme uncertainty based on poor sampling
  techniques, low sample density, poor assay methods

10
Inagaki et al. 2006
11
Microbial activity in the subsurface biosphere
Surface world

• Low activities! Many estimates by geochemical
  modeling
• Subsurface rates range from 100 to 10-13 moles of
  CO2 /L/yr
• Doubling times as low as 1013 sec
• We know little about how they survive
• Figure modified from Onstott et al. 1999

organics reduced inorganics C1 cycling
radiolysis of water serpentinization
12
Metabolic activities expected from water
chemistry analyses versus metabolic activities
estimated from radiotracer experiments for
sediments from deep in the SE Coastal Plain
Data from Phelps et al. 1994
13
Modeling of hydrates requires biological data and
better biological data than we currently have

• key parameters in this model are the rate of
  sedimentation, the quantity and quality of the
  organic material, and a rate constant that
  characterizes the vigor of biological
  productivity (Davie and Buffett 2001)
• Models that predict the occurrence, distribution,
  and quantity of methane hydrates require better
  parameters for the biological contribution (Xu
  and Ruppel, 1999 Davie and Buffett, 2001
  Gering, 2003 Dickens, 2003)

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Objective Determine the realistic rates of
methanogenesis for sediments that contain hydrates

• Measure microbial methane production at
  maintenance levels of activity
• Determine methanogen numbers in sediments
  containing hydrates
• Estimate the amount of methane made per unit
  volume in the sediments for hydrate models

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Biomass recycle reactor (BRR) with Methanoculleus
submarinus(Mikucki et al. 2003)

• M. submarinus
• From 247 mbsf in Nankai Trough
• Sustained by H2/CO2 or formate at 25oC
• Doubling time ca. 100 h at 25oC
• BRR
• Post-exponential, chronic starvation, constant
  biomass at low activity
• Derive methane produced at maintenance level
  activity

growth medium
reactor
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How much methane is produced per cell under
chronic starvation?
A
1000
1000
100
100
cells/mL (x 106)
Methane conc (ug//L x 10,000)
10
10
1
1
Dissolved H2 and CH4 were removed from
suspensions of starved cells and then CH4
accumulation measured over time
Colwell et al. 2008
Methanogenic Methanogenic rate system (fmol
CH4/cell/day) Reference M. submarinus in BRR
0.017 Lake sediments 31.5 Lay et al.
1996 Marine sediments 45.0 Williams
Malcolm, 1980 Anaerobic reactors 108.8-135.0
Li Noike, 1992
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0.017 fmol CH4/cell/day 0.000085 g CH4-C/g
cell-C/h
Figure from Price and Sowers, 2004
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Use quantitative polymerase chain reaction
(Q-PCR) to determine the number of methanogens in
sediments
Involves precise control of the PCR reaction to
allow enumeration
19
Methyl Co-M reductase (mcr) gene primersTarget
the a subunit of the C component of the methyl
Co-M reductase gene test on DNA from five orders
of methanogens
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Methanocaldococcus jannaschii DNA extracted from
spiked sediment samplesUsing the primers
conduct step-wise, quantitative polymerase chain
reaction (Q-PCR) in the presence of sample DNA
and SYBR Green 1 The threshold cycle number
is proportional to the log of the initial
concentration of the target (mcr) DNA
Q-PCR targeting the mcr gene
Colwell et al. 2008
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0 mbsf
Vertical core section showing target samples
MF
MR
G/S
MF
MR
G/S
Other candidate core units
100 mbsf
Images from IODP
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ODP Leg 204 Figure from R. Collier
Leg 204, Hydrate Ridge
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Colwell et al. 2008

• Methanogen numbers for all Leg 204 samples
• 25 of the samples show evidence of methanogens
• When gt1000 methanogens per g detected samples
  usually from lt50 mbsf
• Numbers of methanogens typically well below (ca.
  lt1) total cell numbers
• Considerable variability evident

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Highest methanogen biomass above the sulfate
methane interface High methanogen biomass at
depth associated with geological
anomalies? Shallow sediments dominated by
microbial methane (Claypool et al. 2003)
Consistent with archaeal clone libraries from
site 1251 (Nunoura et al. 2008) Most rates lt1.7
x 10-6 nmol CH4/g/day
Sulfate methane interface
26,000
4.5
8
8
Bottom simulating reflector
125
134
42,000
HA (177-180)
196
BSR
Horizon A
2,800,000
1245
1244
1251
Colwell et al. 2008
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If buried organic carbon (OC) is the source of
the methane, are the rates consistent with the
amount of OC available and the time over which it
can be metabolized?
modeled values
most of our rates
Sediments are 1.5 wt OC or 15 mg/g 10 of C in
OC is available for CH4 (Clayton 1992), so, 1.5
mg CH4/g possible For a sample 375 meters deep
(Pliocene-Pleistocene) that is 1.6 million years
old, use a continuous (low) rate of
methanogenesis 1.7 x 10-6 nmole CH4/g/d for
1.6 x 106 yr 1000 nmole CH4/g 1 x 10-3 mmole
CH4/g 0.016 mg CH4/g
Data from Wellsbury et al. 1997 Cragg et al.
1996 Cragg et al. 1992 Reed et al. 2002
On a volumetric basis at South Hydrate Ridge our
rates would equal ca. 239 kg methane/yr which is
ca. 70 of estimated methane that leaks per yr
(Boetius and Suess, 2004)
Figure modified from Colwell et al. 2004
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• Limitations of the current study
• What about factors like pressure, thermodynamics,
  H2?
• DNA extraction efficiencies? Q-PCR detection
  limits? Validity of mcr gene signature?
• The line between viable vs. dead microorganisms?
• Still, newly derived rates (mostly lt1.7 x 10-6
  nmol methane/g/day) seem reasonable
• Lower than previous rates and theres enough
  organic carbon available
• Compare well to steady state geochemistry
  modeling rates of 1.3 x 10-5 nmol/g/day for Site
  1244 (Claypool et al. 2006) and 2.2 x 10-4
  nmol/g/day for N. Cascadia (Malinverno, 2008)
• May be typical of closed systems with little
  input of outside fluids
• Likely lower than rates in open systems with
  high fluid flux

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Production of gas from hydrates?
Modified from Judd and Hovland, 2007
Rapid gas release?
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An unusual microbial community in methane-rich
sediments?

• peachy-orange gelatinous material
• Shallow sediments associated with fractures?
• Large (10 um) coccoid cells, often clumped in
  tetrads

Offshore India
Hydrate Ridge
Offshore Vancouver Island (Riedel et al. 2006)
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Scientific Opportunities in the Deep Subseafloor
Biosphere
Are micro-eukarya present? Are viruses abundant
or rare? How do microbes respond to sediment
composition on cm to m scales? Which
intracellular activities are essential when a
cell is barely alive? What are the genomic
features of cells in the deep subseafloor
biosphere? What is the effect of metabolic
activity on genome evolution?
U.S. Working Group on IODP and the Deep
Biosphere, April 2006
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http//www.iodp.org/weblinks/Tasks-Scientists/Requ
est-Access-to-Samples/
http//www.iodp.org/
31
Hypothetical geologic cross-section biosphere
defined by temperature
0
5
Depth (km below land or sea surface)
10
15
Geothermal gradient 33 55
37 excessive 40 excessive
25 (C/km) Depth (km) to 6 2
3 land 3 rock-water
5 120C surface
interface
Other factors to consider pressure moisture
potential nutrients confinement flux combined
effects
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PhyloChip - process
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PhyloChip

• PhyloChip 500,000 probes
• (300k target 16S)

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Scientific Ocean Drilling History

• Project Mohole 1958-1966
• Deep Sea Drilling Program 1968-1983
• 1968-1973 US funded
• 1973-1983 International
• Ocean Drilling Program 1985-2003
• Integrated Ocean Drilling Program 2003-present

Glomar Challenger (Deep Sea Drilling Program)
JOIDES Resolution
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IODP - How to Participate

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http//www.oceanleadership.org/usssp