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Applied and Environmental Microbiology, November 2003, p. 6703-6714, Vol. 69, No. 11
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.11.6703-6714.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Diversity and Activity of Methanotrophic Bacteria in Different Upland Soils

Claudia Knief,1 André Lipski,2 and Peter F. Dunfield1*

Max-Planck-Institut für terrestrische Mikrobiologie, 35043 Marburg,1 Abteilung Mikrobiologie, Fachbereich Biologie/Chemie, Universität Osnabrück, 49069 Osnabrück, Germany2

Received 2 June 2003/ Accepted 19 August 2003

Samples from diverse upland soils that oxidize atmospheric methane were characterized with regard to methane oxidation activity and the community composition of methanotrophic bacteria (MB). MB were identified on the basis of the detection and comparative sequence analysis of the pmoA gene, which encodes a subunit of particulate methane monooxygenase. MB commonly detected in soils were closely related to Methylocaldum spp., Methylosinus spp., Methylocystis spp., or the forest sequence cluster (USC {alpha}), which has previously been detected in upland soils and is related to pmoA sequences of type II MB (Alphaproteobacteria). As well, a novel group of sequences distantly related (<75% derived amino acid identity) to those of known type I MB (Gammaproteobacteria) was often detected. This novel upland soil cluster {gamma} (USC {gamma}) was significantly more likely to be detected in soils with pH values of greater than 6.0 than in more acidic soils. To identify active MB, four selected soils were incubated with 13CH4 at low mixing ratios (<50 ppm of volume), and extracted methylated phospholipid fatty acids (PLFAs) were analyzed by gas chromatography-online combustion isotope ratio mass spectrometry. Incorporation of 13C into PLFAs characteristic for methanotrophic Gammaproteobacteria was observed in all soils in which USC {gamma} sequences were detected, suggesting that the bacteria possessing these sequences were active methanotrophs. A pattern of labeled PLFAs typical for methanotrophic Alphaproteobacteria was obtained for a sample in which only USC {alpha} sequences were detected. The data indicate that different MB are present and active in different soils that oxidize atmospheric methane.

* Corresponding author. Mailing address: Max-Planck-Institut für terrestrische Mikrobiologie, Karl-von-Frisch-Str., 35043 Marburg, Germany. Phone: 49-6421-178-733. Fax: 49-6421-178-809. E-mail: dunfield{at}staff.uni-marburg.de.

Applied and Environmental Microbiology, November 2003, p. 6703-6714, Vol. 69, No. 11
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.11.6703-6714.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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