This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Supplemental material
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Pratscher, J.
Right arrow Articles by Braker, G.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Pratscher, J.
Right arrow Articles by Braker, G.
Agricola
Right arrow Articles by Pratscher, J.
Right arrow Articles by Braker, G.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, February 2009, p. 802-810, Vol. 75, No. 3
0099-2240/09/$08.00+0     doi:10.1128/AEM.01992-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Application of Recognition of Individual Genes-Fluorescence In Situ Hybridization (RING-FISH) To Detect Nitrite Reductase Genes (nirK) of Denitrifiers in Pure Cultures and Environmental Samples {triangledown} ,{dagger}

Jennifer Pratscher,1 Catrin Stichternoth,1,2 Katrin Fichtl,3 Karl-Heinz Schleifer,3 and Gesche Braker1*

Max Planck Institute for Terrestrial Microbiology, Marburg, Germany,1 Heinrich Heine University, Düsseldorf, Germany,2 Technical University of Munich, Munich, Germany3

Received 27 August 2008/ Accepted 22 November 2008

Denitrification is an alternative type of anaerobic respiration in which nitrate is reduced to gaseous products via nitrite. The key step in this process is the reduction of nitrite to nitric oxide, which is catalyzed by two structurally different but functionally equivalent forms of nitrite reductase encoded by the nirK and nirS genes. Cultivation-independent studies based on these functional marker genes showed that in the environment there was a dominance of organisms with nirK and nirS genes presumably derived from organisms that have not been cultured yet. However, the phylogenetic affiliation of these organisms has not been resolved since the ability to denitrify is widespread in phylogenetically unrelated organisms. To unravel the phylogeny of the organisms from which the nitrite reductase (nirK) genes originated, one option is to use a special variant of whole-cell hybridization termed recognition of individual genes-fluorescence in situ hybridization (RING-FISH). In RING-FISH a multiply labeled transcript polynucleotide probe is used to detect a single gene on the bacterial chromosome during FISH. Here, RING-FISH was used with laboratory cultures and environmental samples, such as activated sludge. Furthermore, probe-based cell sorting using magnetic beads could also be carried out with mixtures of pure cultures, which led to effective depletion of the nirK-negative organism but capture of the nirK-positive organism, which was demonstrated by terminal restriction fragment length polymorphism analysis based on 16S rRNA genes. The results indicate that RING-FISH coupled with probe-based cell sorting could be used with environmental samples, which could provide a means for phylogenetic classification of nirK-type denitrifiers. Thus, the results of RING-FISH could increase our understanding of the phylogeny and function of denitrifying microorganisms in the environment.

* Corresponding author. Mailing address: Max Planck Institute for Terrestrial Microbiology, Karl-von-Frisch-Strasse, D-35043 Marburg, Germany. Phone: 49 6421 178 733. Fax: 49 6421 178 999. E-mail: braker{at}mpi-marburg.mpg.de

{triangledown} Published ahead of print on 12 December 2008.

{dagger} Supplemental material for this article may be found at http://aem.asm.org/.

Applied and Environmental Microbiology, February 2009, p. 802-810, Vol. 75, No. 3
0099-2240/09/$08.00+0     doi:10.1128/AEM.01992-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Ishii, S., Yamamoto, M., Kikuchi, M., Oshima, K., Hattori, M., Otsuka, S., Senoo, K. (2009). Microbial Populations Responsive to Denitrification-Inducing Conditions in Rice Paddy Soil, as Revealed by Comparative 16S rRNA Gene Analysis. Appl. Environ. Microbiol. 75: 7070-7078 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»