Oxidation von atmosphärischem Methan in Böden: Ein neuer quantitativer Ansatz zur Erfassung der Struktur und Aktivität methanotropher Gilden
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Date
2004-01-22
Authors
Publisher
Philipps-Universität Marburg
item.page.supervisor-of-thesis
Abstract
Methane (CH4) is the most important greenhouse
gas after CO2. Its atmospheric mixing ratio is increasing by 1
% y-1 due to anthropogenic impact. The main sources of CH4 are
wetlands (e.g. peat bogs or rice paddies) where methanogenesis
takes place. Stratospheric destruction of CH4 by hydroxyl
radicals, stratospheric loss into space, and CH4 uptake by
upland soils are main sinks for atmospheric methane. In upland
soils methane-oxidizing bacteria (MOB) are responsible for this
acitivity. The aim of this work was to assess quantitatively
the composition of this functional group of bacteria, and the
acitivity of predominant taxa. Due to the lack of an applicable
culture-independent method for quantitation of methanotrophic
bacteria in soils, a novel molecular method was developed.
Functional marker genes (pmoA and mmoX) that reflect the
phenotype and phylogeny of methanotrophs were chosen as targets
for quantitation by real-time PCR. These genes encode subunits
of methane monooyxgenases. A comparison of 16S rRNA-based
phylogenies to phylogenies based on these functional marker
genes revealed distinct groups of methanotrophic genera (main
groups). Real-time PCR assays based on pmoA and mmoX detection
were developed to target these main groups. Amplicon detection
was achieved by using SybrGreen. The introduction of an
additional temperature step after elongation (4-step protocol)
used for data acquisition led to detection limits comparable to
probe-based assays (e.g. TaqMan or hybridization probes). Thus,
a quantitative detection of methanotrophs by targeting the
genes encoding key enzymes was possible. The cell number
estimates obtained by application of this technique to soil
were reproducable and comparable to estimates obtained by
cultivation-dependent methods in other studies.
Two upland
forest soils in Germany that oxidized atmospheric methane were
investigated. MF was an acidic (pH 4.3) sandy soil developed
from sandstone, and GF was a neutral (pH 7.7) clay-textured
soil developed from limestone. Both soils displayed high rates
of atmospheric methane uptake (MF: 95[±32] mg CH4 m-2 d-1; GF:
50[±7] mg CH4 m-2 d-1). The dominant methanotrophic guilds were
represented by two pmoA sequence groups from as-yet
uncultivated methanotrophs. USC a (6,4[±0,2]x106 target
molecules g-1 soil [dw], soil MF) and USC g (11,6[±3,3]x106
target molecules g-1 soil [dw], soil GF) contributed at least
85 % to the total detectable abundance of methanotrophic
bacteria. Cell-specific activity of the taxon USC a in MF
(17,1x10-5 fmol cell-1 h-1) was higher than the cell-specific
activity of USC g in GF (2,77x10-5 fmol cell-1 h-1) despite the
fact that the abundances of USC g were higher. Supporting this
difference in metabolic activity, only mRNA of USC a
(pmoA-Genes) was detectable above detection limit in soil MF,
while no mRNA was detectable in soil GF. In total, these
findings led to the hypothesis that these two main pmoA groups
represent ?high affinity methanotrophs? adapted to
atmospheric methane concentration.
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Metadata
Contributors
Supervisor:
Dates
Created: 2004Issued: 2004-01-22Updated: 2011-08-10
Faculty
Fachbereich Biologie
Language
ger
Data types
DoctoralThesis
Keywords
molecular ecology, phylogenetic markergeneEcologyReal-time PCRmolekulare Ökologiephylogenetisches MarkergenReal-time PCR
DFG-subjects
Methanoxidierende Bakterien , Polymerase-KettenreaktionMethylotropher MikroorganismusMethanBodenbiologieBiozönoseMikrobiologie
DDC-Numbers
570
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Kolb, Steffen (128701463): Oxidation von atmosphärischem Methan in Böden: Ein neuer quantitativer Ansatz zur Erfassung der Struktur und Aktivität methanotropher Gilden. : Philipps-Universität Marburg 2004-01-22. DOI: https://doi.org/10.17192/z2004.0070.
License
This item has been published with the following license: In Copyright