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Applied and Environmental Microbiology, August 2002, p. 3818-3829, Vol. 68, No. 8
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.8.3818-3829.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Biodiversity of Denitrifying and Dinitrogen-Fixing Bacteria in an Acid Forest Soil

Christopher Rösch, Alexander Mergel, and Hermann Bothe*

Botanisches Institut, Universität zu Köln, D-50923 Cologne, Germany

Received 30 January 2002/ Accepted 29 April 2002

Isolated soil DNA from an oak-hornbeam forest close to Cologne, Germany, was suitable for PCR amplification of gene segments coding for the 16S rRNA and nitrogenase reductase (NifH), nitrous oxide reductase (NosZ), cytochrome cd1-containing nitrite reductase (NirS), and Cu-containing nitrite reductase (NirK) of denitrification. For each gene segment, diverse PCR products were characterized by cloning and sequencing. None of the 16S rRNA gene sequences was identical to any deposited in the data banks, and therefore each of them belonged to a noncharacterized bacterium. In contrast, the analyzed clones of nifH gave only a few different sequences, which occurred many times, indicating a low level of species richness in the N2-fixing bacterial population in this soil. Identical nifH sequences were also detected in PCR amplification products of DNA of a soil approximately 600 km distant from the Cologne area. Whereas biodiversity was high in the case of nosZ, only a few different sequences were obtained with nirK. With respect to nirS, cloning and sequencing of the PCR products revealed that many false gene segments had been amplified with DNA from soil but not from cultured bacteria. With the 16S rRNA gene data, many sequences of uncultured bacteria belonging to the Acidobacterium phylum and actinomycetes showed up in the PCR products when isolated DNA was used as the template, whereas sequences obtained for nifH and for the denitrification genes were closely related to those of the proteobacteria. Although in such an experimental approach one has to cope with the enormous biodiversity in soils and only a few PCR products can be selected at random, the data suggest that denitrification and N2 fixation are not genetic traits of most of the uncultured bacteria.

* Corresponding author. Mailing address: Botanisches Institut, Universität zu Köln, Gyrhofstr. 15, D-50923 Cologne, Germany. Phone: 49 221 470 2760. Fax: 49 221 470 5039. E-mail: Hermann.Bothe{at}uni-koeln.de.

Applied and Environmental Microbiology, August 2002, p. 3818-3829, Vol. 68, No. 8
0099-2240/02/$04.00+0     DOI: 10.1128/AEM.68.8.3818-3829.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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