Summary of the current status of the work of TUM-BO

Andreas Gattinger, Matthias Weiss, Michael Schloter

GSF-Research Center for Environment and HealthTechnical University of MunichInstitute of Soil EcologyD- 85764 Neuherberg

1. Adaptation of protocols for lipid biomarker analyses to peat

(bog) samples

2. Adaptation of protocols for DNA analyses to peat (bog)

samples

3. Growing of keystone plant species in the greenhouse for plant

litter labelling

4. Installation of the new analytical device GC/MS-C-IRMS for

compound specific isotope analysis

Summary of the current status of the work of TUM-BO

1. Adaptation of protocols for lipid biomarker analyses to peat (bog) samples

Determination of microbial communites

•Problem: less than 5% of the estimated total soil microflora can be cultivated

•Solution: cultivation-independent approaches enable the characterization of microbial communiteis in situ:

0

1

2

3

4 7

5

6

8

910

11

12

1314 15

PLEL derived hydrocarbons

genotypic approach:(example archaeal 16S rDNA,

separation with SSCP )

phenotypic approach:(side chain analysis of polar membrane lipids)

methanogens

The biosphere of soil

(mod. Version, M. Schmid, GSF)

Archaeal kingdoms (Reiche):Archaeal kingdoms (Reiche):- Euryarchaeota- Euryarchaeota- Crenarchaeota- Crenarchaeota- - (Korarchaeota)(Korarchaeota) - - (Nanoarchaeota)(Nanoarchaeota)

HO

O

CH2

C

O

C

O

O P O

O

O

NH3+

H O

CH2

O

O P O

O

O

NH3+ ArchaeaArchaea

etherlinked isoprenoids (PLEL):- saturated short chain: i20:0- saturated long chain: i40:0- cyclic long chain: i40:0-cy- unsaturated short chain: i20:1

Side chain analysis of polar membrane lipids of BacteriaEucarya and Archaea in soil

Bacteria & Bacteria & EucaryaEucaryaesterlinked fatty acids (PLFA):- saturated (SATFA)- monounsaturated (MUFA)- polyunsaturated (PUFA)- hydroxysubstituted (PLOH)

O

OH OH

NH

Bacteria & Bacteria & EucaryaEucaryanon-esterlinked fatty acids (NEL-PLFA):- hydroxysubstituted (UNOH)- unsubstituted fatty acids (UNSFA)

Extended phospholipid analysis

xBodenökologie

Sam ple

G C-M S

Neutral lipids Glyco lipids

B ligh-D yer-E xtrac tion

L ipids

G C-M S

NH -column2

G C-M S

Ether lipids (PLEL)

Hydroxy--

(EL-PLFA)substituted

SATFA MUFA+DMDS

PUFA PLOH+TMSI

UNSFA UNOH+TMSI

Reduktive dehalogenation

A lkyl iodides

Isoprenoid hydrocarbons

SCX-column NH -column2

SI-column

Phospholipids

Acid ic hydrolysis

Non-substituted (EL-PLFA)

Unsaponifiable lipids

Mild alkaline methanolysis

(NEL-PLFA)

Fatty acids (PLFA)Acid ic hydrolysis

ArchaeaBacteria & Eucarya

Lipid biomarker [nMol g-1 dry matter]

SATFA (mainly bacteria)MUFA (mainly aerobic bacteria)PUFA (eukaryotes: fungi, protozoa)PLOH (mainly Gram-negative bacteria)UNOH (anaerobic bacteria and some fungi)UNSFA (mainly anaerobic bacteria)PLEL (archaea)

100 200 300 400 5000

0-6

6-12

12-18

18-24

24-30

So

il d

ep

th [

cm

]

63

63

69

50

10

8

48

13

10

14

13

8

2

1

1

1

1

2

3

5

156

191

179

175

183

Numberpercentage of biomarker fraction

Microbial biomarker composition in a Scottish “test” peat core

63

69

13

10

14

2

1

1

1

2

3

5

Biomarker ratio [mol mol-1]

0 1 2 3 20 25 30 35

Bacteria : FungiMOB : Archaea

So

il d

ep

th [

cm

]

0-6

6-12

12-18

18-24

24-30

Lipid biomarker ratios in a Scottish “test” peat core

MOB IMOB IIArchaea

2 4 6 8 10 12 140

So

il d

epth

[cm

]

0-6

6-12

12-18

18-24

24-30

Lipid biomarker [nmol g-1dry matter]

MOB and archaeal biomarker in a Scottish “test” peat core

2. Adaptation of protocols for DNA analyses to peat (bog) samples

together with Rebekka Artz, Steve Chapman/MLURI

3. Growing of keystone plant species in the greenhouse for plant litter labelling

Plants were taken by A. Siegenthaler et al. at the French/Suisse

site:

4 plant species: 2 mosses (Sphagnum, Polytrichum), 2 grasses

(Eriophorum vaginatum, Eriophorum angustifolium) were

grown at the GSF greenhouse for estimating growth conditions

and biomass production

climatic conditions:

4. Installation of the new analytical device GC/MS-C-IRMS for compound specific isotope analysis

Simultaneous identification and quantification of PLFA/PLEL from environmental samples and their corresponding 12C/13C ratios by

GC/MS-C-IRMS

MS(DSQ)

IRMS(DeltaPlusAdvantage)

20% of the analyte

80% of the analyte

delta 13C values in PLFA/PLEL from test core samples varied

between -26 and -32 %0 (natural abundance):

due to dilution effects with the peat soil and 1-2%

measurement inaccuracy, an enrichment with 13C in the plant

litter of at least 70 %0 PDB is needed:

Which quality of CO2 gas for plant labelling?