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Applied and Environmental Microbiology, February 1999, p. 396-403, Vol. 65, No. 2
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Molecular Analysis of Ammonia-Oxidizing Bacteria of the beta  Subdivision of the Class Proteobacteria in Compost and Composted Materials

George A. Kowalchuk,1,* Zinaida S. Naoumenko,2 Piet J. L. Derikx,3 Andreas Felske,4 John R. Stephen,5 and Irina A. Arkhipchenko2

Department of Plant-Microorganism Interactions, Netherlands Institute of Ecology, Center for Terrestrial Ecology, 6666 ZG, Heteren, The Netherlands1; Laboratory for Microbial Ecotechnology, Research Institute for Agricultural Microbiology, St. Petersburg-Pushkin 8, 189620, Russia2; Department of Manure Technology, Institute of Agricultural and Environmental Engineering (IMAG-DLO), NL-6700 AA, Wageningen, The Netherlands3; Department of Microbiology, Wageningen Agricultural University, 6703 CT, Wageningen, The Netherlands4; and Center for Environmental Biotechnology, University of Tennessee, Knoxville, Tennessee 37932-25755

Received 18 September 1998/Accepted 9 November 1998

Although the practice of composting animal wastes for use as biofertilizers has increased in recent years, little is known about the microorganisms responsible for the nitrogen transformations which occur in compost and during the composting process. Ammonia is the principle available nitrogenous compound in composting material, and the conversion of this compound to nitrite in the environment by chemolithotrophic ammonia-oxidizing bacteria is an essential step in nitrogen cycling. Therefore, the distribution of ammonia-oxidizing members of the beta  subdivision of the class Proteobacteria in a variety of composting materials was assessed by amplifying 16S ribosomal DNA (rDNA) and 16S rRNA by PCR and reverse transcriptase PCR (RT-PCR), respectively. The PCR and RT-PCR products were separated by denaturing gradient gel electrophoresis (DGGE) and were identified by hybridization with a hierarchical set of oligonucleotide probes designed to detect ammonia oxidizer-like sequence clusters in the genera Nitrosospira and Nitrosomonas. Ammonia oxidizer-like 16S rDNA was detected in almost all of the materials tested, including industrial and experimental composts, manure, and commercial biofertilizers. A comparison of the DGGE and hybridization results after specific PCR and RT-PCR suggested that not all of the different ammonia oxidizer groups detected in compost are equally active. amoA, the gene encoding the active-site-containing subunit of ammonia monooxygenase, was also targeted by PCR, and template concentrations were estimated by competitive PCR. Detection of ammonia-oxidizing bacteria in the composts tested suggested that such materials may not be biologically inert with respect to nitrification and that the fate of nitrogen during composting and compost storage may be affected by the presence of these organisms.

* Corresponding author. Mailing address: Department of Plant-Microorganism Interactions, Netherlands Institute of Ecology, Center for Terrestrial Ecology, Boterhoeksestraat 22, Postbox 40, 6666 ZG Heteren, The Netherlands. Phone: 31 (0) 2647 91314. Fax: 31 (0) 2647 23227. E-mail: gkowal{at}cto.nioo.knaw.nl.

Applied and Environmental Microbiology, February 1999, p. 396-403, Vol. 65, No. 2
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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Copyright © 1999 by the American Society for Microbiology. All rights reserved.