Water linkingthe surface withthe subsurface biogeosphere

Kirsten KüselAquatic GeomicrobiologyFriedrich Schiller University JenaGermany

Research Centre AquaDiva will focus onW

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the important role of water (Aqua) and biodiversity (Diva)for shaping the structure, properties and functions of the subsurface

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• carbon storage• energy storage

and recovery• water resources(people and ecosystems)• filtration and

transformation

Functions of the subsurface

Human impact on the subsurface

http://esd.lbl.gov/research/programs/erwr/research_areas/erss.html

Human impact on the subsurface

http://esd.lbl.gov/research/programs/erwr/research_areas/erss.html

Terrestrial and marine subsurface contains half of the Earth´s biomass.Amend and Teske 2005

Input of carbon to the subsurface

Akob and Küsel 2011 Biogeosciences

e.g.,CH4, N2O

Carbon fixation pathways in subsurface

1. Calvin-Benson-Bassham cycle Aerobic and anaerobic Bacteria and ArchaeaRubisCO I (cbbL) and RubisCO II gene (cbbM) (Alfreider et al. 2003, 2009)

2. Reductive tricarboxylic acid cycleAnaerobic or microaerophilic Bacteria (Berg, 2011)ATP citrate lyase α-subunit (aclA) (Hügler et al. 2005)

3. Reductive acetyl-CoA pathway Anaerobic microbes (Berg, 2011)acetyl-CoA carboxylase subunit (accC) (Auget et al. 2008)

4. 3-hydroxypropionate cycleArchaeaacetyl-CoA carboxylase subunit (accC) (Auget et al. 2008)

5. 3-hydroxypropionate/4-hydroxybutyrate cycleacetyl-CoA carboxylase subunit (accC) (Auget et al. 2008)

Calvin-cycle

Berg 2011 AEM

RubisCO

I (cbbL gene)Form IA: proteobacteriaForm IC: alpha- and beta-proteobacteria

II (cbbM gene)

different forms:

• form I: low CO2 availability, oxic conditions

• form II: requires higher CO2 concentrations, anoxic/microoxic conditions

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Is input affected by surface changes?

• Decline in surface biodiversity• Land use change• Disturbances

(e.g. management practices)• Weather extremes(heavy rain falls, snow melt events)

Coupling theme

Follow the surface inputinto the groundwater• Identification of

surface signals• Transformation of

signals DOM patterns metabolomics colloidal transport

Event themeHow important aresingular and extreme events for carboninput into thesubsurface?• Effect of heavy rain

falls or snow meltson substrate supply

Kammerforst

Wutha-Farnroda Jena

Mühlhausen

KammerforstBad Langensalza

Groundwater observation transect

Mapsource: Google inc.

Drilling and Core Sample AnalysisThree drilling campaigns (2009-2014)

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Drilling and Core Sample AnalysisAnalyses of rock cores

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Part 1: Review 15

Hydrogeological Characterization and Stratigraphy

Aquifer-Situation (actual state) Spatial Correlation of 2 Aquifer Systems

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Aquifer-Situation (actual state) Groundwater Isotopic Signatures

δ13C DIC -11‰

δ13C DIC -9‰20% depleted in modern carbon

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CMT tubegas-phasesampling

Second wellp-T loggers

Hydrogeochemistry + Gas analyses

Main well

Investigate gases simultaneously with high time resolution and specificity

Keiner et al. 2015 Anal. CA Frosch, et al. 2013 Analytical Chemistry

Look into the aquifer

Part 1: Review 20

Following extreme events: 8 in 3 years

> 20 mm/day

H1H2

H3H4

H5

H2

H5

H3

H4

Part 1: Review 21

Following extreme events

> 20 mm/day

Do bacterial abundances follow groundwatertable fluctuations?

Upper aquifer (H4-3)

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bac-16S rRNA genesgroundwater table

Abundance:qPCR

(bacterial and archaeal 16S rRNA genes,

selected functional genes: CO2 fixation, nitrogen cycle)

September 2013 Diversity:16S bacteria, archaea, 18S rRNA , ITS

(DNA + RNA-based)cbbM, cbbL 1A, cbbL 1C (DNA-based)

454 pyrosequencing19.000 – 30.000 reads

Summer 2014 Diversity (on-going):Illumina sequencing

filtration

ground-water

chemistry0.2 µm

Molecular analysis pipelineDNA (+ RNA)

Monthly Sampling Campaigns

160 isolates

-> The Archaea represented ca. 9 % of the total prokaryotes in the lower aquifer

ca. 2 % of the total prokaryotes in the upper aquifer

1,00E+00

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H4-3

H4-3 arch

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H4-1

H4-1 arch

Upper aquifer(0-25% O2)

Lower aquifer(40-80% O2)

Fluctuations of prokaryotesge

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L-1

Archaeal diversity along transectUpper Aquifer (0-25% O2) Lower Aquifer (40-80% O2)

-> largely dominated by the Marine Group I (MG-I) Thaumarchaeota=> ammonia oxidizers (NH4

+ NO2- ….. NO3

-) -> important for geochemistry data

0%

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H3-2 H4-2 H5-2 H4-3 H5-3 H3-1 H4-1 H5-1

South African Gold Mine Euryarchaeotal Group 1

Marine Benthic Group E

Unclassified Thermoplasmatales

Valkea Kotinen group II

Valkea Kotinen group III

Marine Benthic Group A (+pSL12)

Forest Soil Crenarchaeotal Group

Soil Crenarchaeotal Group

Miscellaneous Crenarchaeotal Group

Unclassified Thaumarchaeota

South African Gold Mine Crenarchaeotal Group 1

Marine Group I (MG-I)Marine Group I

Which other Archaea are detected?

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H3-2 H4-2 H4-3 H5-2 H5-3 H3-1 H4-1 H5-1

South African Gold Mine Euryarchaeotal Group 1

Marine Benthic Group E

Unclassified Thermoplasmatales

Valkea Kotinen group II

Valkea Kotinen group III

Marine Benthic Group A (+pSL12)

Forest Soil Crenarchaeotal Group

Soil Crenarchaeotal Group

Miscellaneous Crenarchaeotal Group

Unclassified Thaumarchaeota

South African Gold Mine Crenarchaeotal Group 1

Marine Group I

Upper Aquifer (0-25% O2) Lower Aquifer (40-80% O2)

-> soil associated archaeal groups

(VAL-III)

Possible archaeal metabolism

50 m

H5-3 NO3-:

10 µmol L-1

O2: <1% sat.

VAL-III- Boreal forest lake- unknown function

Miscellaneous Crenarchaeotal Group:- protein degradation (Lloyd et al, 2013)- anaerobic CO2 fixation (reductive acetyl-coA pathway; Lazar et al. in press)

H5-190 m

NO3-:

120 µmol L-1

O2: 25% sat.

Marine Group I:- ammonia oxidizing (Könneke, 2005)- CO2 fixation (3HP/4HB pathway)

Can we detect key bacterial taxa involvedin CO2 fixation in the aquifers?

Up to 17% harbor RubisCO encoding genes

lower aquiferupper aquiferH3-2 H4-2 H4-3 H5-2 H5-3 H3-1 H4-1 H5-1

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bacterial 16S rRNA cbbM

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Herrmann et al., AEM 2015

CO2-fixing communities: RubisCO form II (CbbM)

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H3-2 H4-2 H4-3 H5-2 H5-3 H3-1 H4-1

lower aquiferupper aquifer

H5-1

100oOOOOOOOOOOOOOOOOOOOOOOSideroxydans lithotrophicus

Acidithiobacillus ferrooxidans

Cand. Thiodictyon syntrophicumAcidithiobacillus caldusSulfuritalea hydrogenivoransThiobacillus thioparusSulfuritalea hydrogenivoransSulfuritalea hydrogenivoransThiohalomonas denitrificansThiobacillus thiophilusThiothrix lacustrisThiobacillus thioparusCand. Accumulibacter sp.Halothiobacillus sp.Leptothrix ochraceaeThiobacillus thioparusLeptothrix colodniiDechloromonas aromaticaAlbidiferax ferrireducensThiobacillus thioparusMagnetospirillum gryphiswaldense

Phaeospirillum fulvumothers

Constantly high fraction of groups oxidizingreduced sulfur and iron compounds

RubisCO form IA (CbbL): most abundant OTUs100

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H3-2 H4-2 H4-3 H5-2 H5-3 H3-1 H4-1

lower aquiferupper aquifer

H5-1

% OOOOOOOOOOOOOOOOOOOO

others

Sulfuricella denitrificans

Thiobacter subterraneus

Acidithiobacillus ferrooxidans

unclassified

unclassifiedunclassifiedunclassifiedNitrosomonas sp.

Nitrosospira sp.

Thiothrix lacustris

Nitrosospira sp.Ferrovum mycofaciensNitrosomonas sp.Nitrosococcus halophilus

unclassifiedBeggiatoa sp.

unclassifiedNitrobacter winogradskyi

Nitrosospira sp.unclassified

OTU cutoff: 0.05 on protein level„unclassified“: less than 90 % similarityto cultured representatives

RubisCO form IC (CbbL): most abundant OTUs100

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H3-2 H4-2 H4-3 H5-2 H5-3 H3-1 H4-1

lower aquiferupper aquifer

H5-1

others

Burkholderiales bacterium

unclassifiedVariovorax sp.

unclassified

Nitrosomonas sp. Is79A3

unclassifiedRhodopseudomonas palustrisBradyrhizobium sp.Nitrosomonas sp. IsA73

Bradyrhizobium elkaniiNitrosospira sp.Nitrosospira sp.

Variovorax sp.Bradyrhizobium sp.

Nitrosospira sp.Rubrivivax gelatinosus

Rubrivivax benzoatilyticus

Acidithiomicrobium sp.

Rhizobium selenitireducensNevskia ramosa

Ammonia-oxidizing bacteria

DNA- and RNA-based results: species-level OTUs others

Gallionella capsiformisBdellovibrio exovorusNitrospira calidaSulfuritalea hydrogenivoransElusimicrobiaceaeNitrospira moscoviensisMagnetospirillum sp.Rhodobacter sp.BacillaceaeActinobacterium sp.Nitrospira sp.RhodocyclaceaeSediminibacterium sp.Geopsychrobacter electrodiphilus

Pseudomonas baeticaCoxiellaceaeLevilinea saccharolyticaCand. Nitrotoga arcticaNitrospira moscoviensisNitrospiraceae

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DNAH3-2

RNA DNA RNA DNA RNA DNAH5-3 H4-1 H5-1

lower aquiferupper aquifer

RNA

20 most abundant OTUs account for 33 % of total sequence reads.

Evidence of active nitrificationin the lower aquifer (well H4-1)

48 m

Aerobic ammonia oxidation:0.64 nmol NH4

+ oxidized L-1 h-1

(Opitz et al. FEMS Microb. Ecol 2014)

potential ammonia oxidizing archaea

Ammonia-oxidizingbacteria and archaea: mostly related toNitrosomonas ureae, Nitrosoarchaeumkoreensis

H4-1

NO3-:

120 µmol L-1

NH4+:

9 µmol L-1

O2: 40% sat.

totalprokaryotic community

totalarchaeal

community

Potential for anammox in the upper aquifer(well H5-3)

Anaerobic oxidation of ammonia(anammox)NH4

+ + NO2- N2 + H2O

Detection of ladderane lipids([5]-ladderane FAME) at 2.6 ng L-1

anammox-bacteria mostly related to Jettenia asiaticaBrocadia sp.50 m

H5-3 NO3-:

29 µmol L-1

O2: <1% sat.

NH4+:

36 µmol L-1

totalprokaryotic community

Carbon fixation

CO2 organic carbonFe (II)

Fe (III)

Sideroxydans lithotrophicus

S,thiosulfate

SO42-

NO3-

N2

Sulfuricelladenitrificans

NH4+

Nitrosomonassp.Is79A3

Iron cycling

NO2-

Sulfur cycling

Calvin cycle+ others

Nitrogen cycling

Nitrospira sp.

O2

H2OThiobacter subterraneus

N2

Water linkingthe surface tothe subsurfacebiogeosphere• Importance for

autotrophy • Input of microbes

from the surface• Input of electron

donors like NH4+

• High dynamics!!!!

Institute of GeosciencesRobert LehmannHeiko MinkmarValerie Schwab-LavricKai Uwe Totsche

Institute of EcologyMartina HerrmannCassandre LazarDenise AkobSebastian OpitzPatricia LangeSwatantar KumarBernd RuppeAnna Späthe

Max-Planck-Institute for BiogeochemistrySusan TrumboreMartin Nowack

Institute for Photonic Technologies, JenaTorsten FroschRobert KeinerJürgen Popp

Part 1: Review

Part 1: Review