Plans for subglacial lake research

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Plans for subglacial lake research

• Biology
• Russian
• (collaborating French in frame of GDRE Vostok)

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Plans for subglacial lake research
Vostok Accretion ice 1 and 2 (and lake
water) Biology print in unusual extreme
environment
Type1
Type2
3607m
3612m
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Plans for subglacial lake research
Vostok Accretion ice 1 and 2 (and lake
water) Biology print in unusual extreme
environment

• Investigate the accretion ice type1 at a scale of
  small ice slices containing mineral/chemical/gas
  impurities for the presence of indigenous life
  trapped by lake sediments

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Search for life signatures in Lake Vostok

• 16S rRNA gene sequencing
• Meltwater
• As it is or concentrating (up to 20 x 103 fold)
  using 3 Kd filter
• Direct PCR
• gDNA extraction (FastPrep-DNA extraction kits)
• PCR with broad-range universal degenerate primers
  targeting 16S rRNA gene (bacteria and archaea)
• Common
• Nested

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Vostok ice for Biology
GIVEN

• Microscopy and preliminary DNA trials of strictly
  decontaminated ice samples
• Very low biomass
• Microscopy - lt7 cells/ml (deduced)
• SYBR-Green flow cytometry
• 9 cells/ml (lake ice)
• 24 cells/ml (deep glacial ice)
• Traces of DNA
• Bacteria and Archaea - lt10 cells/ml (deduced from
  PCR)
• Fungi - lt0.1 cells/ml (deduced from PCR)

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Vostok ice for Biology
PREMISE

• High chance of
• Forward Contamination
• by reason of the Very Low Biomass

An accidental contamination of other worlds
with microbes brought from Earth
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Vostok ice for Biology
REQUIREMENTS

• Strict ice (and bio) decontamination protocols
  and clean (dust-free) room facilities
• Ancient DNA and Trace DNA analyses regulations
• Chemistry controls (DOC, ions-indicators)

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Vostok ice for Biology
REQUIREMENTS

• Comprehensive Biological controls

• Sham/mock DNA extraction
• Negative PCR
• Ice core wash water
• Lab Environment (dust)
• Vostok drill fluid
• Outer-core (optionally)

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Overall Contaminant Database 2160 entries

• Negative PCR
• Sham DNA extraction
• Ultra-pure water
• JPL clean room facilities (gt393 entries!)

• Contaminants in Forensic Science
• Human pathogens and saprophytes
• Deep drilling contaminants

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Indexing contaminant criteriafor Vostok lake ice
findings
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Lake Vostok BIO settings

• Giant subglacial lake
• 250 x 60 km in size more than 14 000 km2 in
  square
• Two basins with the ridge 150-200 m deep
• Max depth 1100-1200 m (south basin)
• Water renewal 80-100 Kyr
• Buried beneath 3750-4200 m thick ice sheet (3.5
  km asl)
• Pressure 337-377 bar
• Water temperature -2.65C
• DOC 17 7 ppbC
• No light
• High oxygen tension - 700-1300 mg/L (14 mg/L
  reference) (accumulation due to glacial ice
  melting)
• Isolated from surface biota for gt15 Ma
• gt0.8 Ma - provided melting feeds the lake
• Low salinity - lt0.10

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Bacterial phylotypes recovered in Vostok
accretion ice - SUMMARY
No nPCR and shDNA extraction clones included
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Vostok lake ice for Biology

• Microscopy trials of strictly decontaminated ice
  samples - Revised
• Very low biomass
• Microscopy - lt7 cells/ml (deduced)
• SYBR-Green flow cytometry
• 9 cells/ml (lake ice)

1 cells/ml
1 cells/ml
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Vostok lake ice for Biology

• Is Lake Vostok sterile?
• According to DAB 10 or USP XXII standards
  sterility means 0.1-1 cfu/1 ml

provided the lake ice represents whole lake
water column, i.e. lake water is not stratified
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Bacterial phylotypes recovered in Vostok
accretion ice - SUMMARY
Thermophile
50 - 52C
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Thermophiles in sediment containing accretion ice
15 Kyr-old
Laboratories location
Taiwan
Mid-Atlantic Ridge 2270-2320 m depth
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Geothermal environment in Lake Vostok
environment?

• Biology evidence
• Thermophiles ( mesophiles?)
• Geology and Geophysics evidence
• Rift structure - geological settings
• 4He degassing - long-term seismotectonic evidence
  from
• 18O shift - enrichment with the 18O isotope
  indication for hydrothermal spring waters

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Geothermal environment in deep crustal faults
within the lake bedrock
Seeping solutions from a crust encouraged by
seismotectonic events may boost hydrothermal
plume which flush out crustal thermophilic
bacteria and minerals up to their vents.
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(No Transcript)
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Bacterial phylotypes recovered in Vostok
accretion ice - SUMMARY
Thermophile
50 - 52C
Celine Lavire/Philippe Normand
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Geothermal environment in deep crustal faults
within the lake bedrock
West
East
Crust
Crustal faults revealed beneath the Lake Vostok
by earthquake exchange waves approach
Mantle
Polar Marine Geological Survey (report 2004)
km
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Lake ice BIO features
as compared to glacial ice
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Bacterial metabolic processes in accretion ice
type 1
(sediment inclusions, no oxygen)
?
H2 oxidation
?
Hydrogenophilus thermoluteolus Methanogenic
archaea Sulfate-reducing prokaryotes (SRPs)
Our 16S rDNA primers match well but were enable
to recover
No indication for CH4
No indication for H2S
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Vostok Accretion ice 1 CONCLUSION

• Contains a tiny amount of DNA (cells?)
  originating from within the lake bedrock deep
  faults(?)

Water body
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Life potential within the lake bedrock
Studinger et al., 2003

• A Anaerobic chemoautotrophic thermophiles
  (mesophiles)

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Vostok Accretion ice 2CONCLUSION

• Seems to be DNA-free (cell-free?) by reason of
• Supposed oxygen burst in the lake which can
  easily destroys any cells/DNA in a time
• No surface cells exist which survive transit to
  the lake and be trapped into the ice

700-1300 mg/L O2
Water body
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OUTLOOK

• Lake Vostok
• Long-term isolated - gt15 Ma
• High pressure
• Hyper-oxigenic? 700-1300 mg/L
• Highly oxidized environment?
• superoxide, hydrogen peroxide, singlet O,
  hydroxyl radical etc. (vs. e.g. carotinoids,
  catalase, superoxide dismutase)
• No potential for redox?
• No organic carbon (ultra-oligotrophic)
• Constantly low but life supporting temp.
  (-2.65oC)

RADARSAT, CSA
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Life potential in lake water
RADARSAT, CSA

• A Oxyphilic chemoautotrophic psychrophilic
  unknown (by energy source) life forms?

Hydrogen?
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Plans for subglacial lake research
Vostok Accretion ice 1 and 2 (and lake
water) Biology print in unusual extreme
environment

• Investigate more young accretion ice type2
  (closer to lake water) in search of lake life
  signatures

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Plans for subglacial lake research
Vostok Accretion ice 1 and 2 (and lake
water) Biology print in unusual extreme
environment

• Cleanly sample and investigate lake water for
  indigenous life signatures

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Plans for subglacial lake research
NGRIP red ice core (SArcticLE?) Control for
Lake Vostok water sampling etc

• Prove approaches and methods in ice/water sterile
  DNA-free sampling, decontamination, recovering
  and proving indigenous microbial biodiversity

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Plans for subglacial lake research
NGRIP red ice core (SArcticLE?) Control for
Lake Vostok water sampling etc
5x103 cell/mL
No cells
Bag 4 3039m
Bag 15 3045m
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Plans for subglacial lake research
Freshwater rift-based Antarctic lakes Positive
control for Lake Vostok water/ice
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Plans for subglacial lake research
Freshwater rift-based Antarctic lakes Positive
control for Lake Vostok water/ice

• Prove approaches and methods in ice/water sterile
  DNA-free sampling, decontamination, recovering
  and proving indigenous microbial biodiversity

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Plans for subglacial lake research
Polar Glacier ice (Vostok EPICA NGRIP)
Biology print in deep past and bedrock life
3489m
121m
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Plans for subglacial lake research
Polar Glacier ice (Vostok EPICA NGRIP)
Biology print in deep past and bedrock life

• Study more young ice (surface snow - 100 Kyr-old)
  and that of interglacial-glacial periods to
  estimate the time limit for DNA survival in
  Antarctic ice

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Plans for subglacial lake research
Polar Glacier ice (Vostok EPICA NGRIP)
Biology print in deep past and bedrock life

• Investigate ice horizons with gas contents spikes
  and other layers to test e.g. bacterial metabolic
  in situ activity hypothesis

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Plans for subglacial lake research
Polar Glacier ice (Vostok EPICA NGRIP)
Biology print in deep past and bedrock life

• Investigate BASAL glacial ice for the presence of
  indigenous subglacial bedrock life

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Glacial ice with Aeolian dust and bedrock
particles Findings

• Two confident phylotypes (of 8)
• Bacillus sp (Environ clone) or B. arsenicus
  (arsenic ore)
• Psychrobacter glacincola (Polar regions only?)

3483 m (704 Kyr -9oC)
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Plans for subglacial lake research
Temperate Glaciers (Alps Andes) Positive
control for polar ice
Illimani (Bolivia) 58m
Illimani (Bolivia) 136m
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Plans for subglacial lake research
Temperate Glaciers (Alps Andes) Positive
control for polar ice

• Study young glacial ice to estimate the time
  limit for DNA survival in ice (positive control
  for polar ice)

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Plans for subglacial lake research
Temperate Glaciers (Alps Andes) Positive
control for polar ice

• Study BASAL glacial ice for the presence of
  indigenous subglacial bedrock life (positive
  control for polar ice)

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Plans for subglacial lake research

• Vostok Accretion ice I and II (and lake
  water) Biology print in unusual extreme
  environment
• NGRIP red ice core (SArcticLE?)
• Control for Lake Vostok water sampling etc
• Freshwater rift-based Antarctic lakes
  Positive control for Lake Vostok water/ice
• Polar Glacier ice (Vostok EPICA- NGRIP)
  Biology print in deep past and bedrock life
• Temperate Glaciers (Alpes Andes) Positive
  control for polar ice Biology print of human
  activity in recent past