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Applied and Environmental Microbiology, April 2009, p. 2111-2121, Vol. 75, No. 7
0099-2240/09/$08.00+0     doi:10.1128/AEM.01540-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Dynamics of Polyphosphate-Accumulating Bacteria in Wastewater Treatment Plant Microbial Communities Detected via DAPI (4',6'-Diamidino-2-Phenylindole) and Tetracycline Labeling ^,

S. Günther,¹ M. Trutnau,² S. Kleinsteuber,¹ G. Hause,³ T. Bley,² I. Röske,⁴ H. Harms,¹ and S. Müller^1*

Department of Environmental Microbiology, UFZ-Helmholtz Centre for Environmental Research, Permoserstrasse 15, 04318 Leipzig, Germany,¹ Institute of Food Technology and Bioprocess Engineering, Dresden University of Technology, Bergstrasse 120, 01069 Dresden, Germany,² Microscopy Unit, Biocenter of the University Halle-Wittenberg, Weinbergweg 22, 06120 Halle/Saale, Germany,³ Institute of Microbiology, Dresden University of Technology, Zellescher Weg 20b, 01217 Dresden, Germany⁴

Received 8 July 2008/ Accepted 15 January 2009

Wastewater treatment plants with enhanced biological phosphorus removal represent a state-of-the-art technology. Nevertheless, the process of phosphate removal is prone to occasional failure. One reason is the lack of knowledge about the structure and function of the bacterial communities involved. Most of the bacteria are still not cultivable, and their functions during the wastewater treatment process are therefore unknown or subject of speculation. Here, flow cytometry was used to identify bacteria capable of polyphosphate accumulation within highly diverse communities. A novel fluorescent staining technique for the quantitative detection of polyphosphate granules on the cellular level was developed. It uses the bright green fluorescence of the antibiotic tetracycline when it complexes the divalent cations acting as a countercharge in polyphosphate granules. The dynamics of cellular DNA contents and cell sizes as growth indicators were determined in parallel to detect the most active polyphosphate-accumulating individuals/subcommunities and to determine their phylogenetic affiliation upon cell sorting. Phylotypes known as polyphosphate-accumulating organisms, such as a "Candidatus Accumulibacter"-like phylotype, were found, as well as members of the genera Pseudomonas and Tetrasphaera. The new method allows fast and convenient monitoring of the growth and polyphosphate accumulation dynamics of not-yet-cultivated bacteria in wastewater bacterial communities.

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* Corresponding author. Mailing address: UFZ-Helmholtz Centre for Environmental Research Leipzig-Halle, Department of Environmental Microbiology, Permoserstrasse 15, 04318 Leipzig, Germany. Phone: 49 341 235 1318. Fax: 49 341 235 1351. E-mail: susann.mueller{at}ufz.de

Published ahead of print on 30 January 2009.

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

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Applied and Environmental Microbiology, April 2009, p. 2111-2121, Vol. 75, No. 7
0099-2240/09/$08.00+0     doi:10.1128/AEM.01540-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.