Physical enrichment of polyphosphate-accumulating organisms in activated sludge

Chun Hsiung Hung, Jordan Peccia, Julie L. Zilles, Daniel R. Noguera

Research output: Contribution to journalArticlepeer-review


Two methods that physically separate polyphosphate-accumulating organisms (PAO) from other organisms in activated sludge were developed. The first method used 4′6-diamidino-2-phenylindole dihydrochloride (DAPI) to selectively stain PAO. When excited with light at 340 nm, polyphosphate granules in DAPI-stained cells fluoresce yellow while cells without polyphosphate fluoresce blue. This difference in fluorescent response was used to separate PAO from non-PAO using flow cytometry. The second method consisted of a simple gradient centrifugation to physically separate PAO from non-PAO based on their density differences. Both methods produced cell suspensions with an increased PAO concentration. From an average PAO concentration of approximately 14% in a full-scale process, the DAPI-flow cytometry method produced sorted samples with PAO representing more than 70% of the total cells, while the density gradient method produced an approximate 43 to 48% PAO enrichment. The physical enrichment methods described herein should facilitate the identification and study of PAO that are relevant in full-scale enhanced biological phosphorus removal processes.

Original languageEnglish (US)
Pages (from-to)354-361
Number of pages8
JournalWater Environment Research
Issue number4
StatePublished - Jan 1 2002
Externally publishedYes


  • 4′6-diamidino-2-phenylindole dihydrochloride
  • Enhanced biological phosphorus removal
  • Flow cytometry
  • Gradient centrifugation
  • Polyphosphate-accumulating organisms

ASJC Scopus subject areas

  • Environmental Chemistry
  • Ecological Modeling
  • Water Science and Technology
  • Waste Management and Disposal
  • Pollution


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