Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Ammonium removal performance of anaerobic ammonium-oxidizing bacteria immobilized in polyethylene glycol gel carrier

Anammox bacteria immobilized in gel carrier


Anaerobic ammonium-oxidizing (anammox) bacteria were immobilized in polyethylene glycol gel carriers. A small amount of seed sludge [0.24% (w/v)] was entrapped in the carriers, and continuous feeding tests were performed. Nitrogen removal activity increased gradually, reaching 3.7 kg N/m3 reactor per day on day 67. The average of nitrogen conversion rate was calculated as 3.4 kg N/m3 reactor per day. Microscopic examination clearly showed that small red clusters formed in the gel carrier. Moreover, fluorescence in situ hybridization analysis revealed that these clusters consisted of anammox bacteria. From real-time polymerase chain reaction analysis, the growth of anammox bacteria in the gel carriers was clearly shown by increased concentration of 16S rRNA gene of planctomycete from 4.3 × 108 to 4.2 × 109 copies/ml between days 41 and 55. To determine the effects of inoculation on the start-up of the reactor, the amount of seed sludge in the gel carrier was varied and it was found that the start-up period could be reduced to as little as 25 days when a sludge concentration of 1.4% (w/v) was used. This is the first report of successful immobilization and cultivation of anammox bacteria in a gel carrier.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8


  1. Amann RI (1995) In situ identification of micro-organism by whole cell hybridization with rRNA-targeted nucleic probes. In: Akkerman ADC, van Elsas JD, de Bruijn FJ (eds) Molecular microbial ecology manual. Kluwer, Dordrecht, pp 1–15

  2. Asano H, Myoga H, Asano M, Toyao M (1992) A study of nitrification utilizing whole microorganisms immobilized by the PVA-freezing method. Water Sci Technol 26:1037–1046

  3. Furukawa K, Rouse JD, Bhatti ZI, Imajo U, Nakamura K, Ishida H (2002) Anaerobic oxidation of ammonium confirmed in continuous flow treatment using a nonwoven biomass carrier. Jpn J Water Treat Biol 38:87–94

  4. Imajo U, Tokutomi T, Furukawa K (2005) Evaluation of various reactor types for the anammox process. J Jpn Soc Water Environ 28:185–190

  5. Isaka K, Date Y, Sumino T, Yoshie S, Tsuneda S (2006) Growth characteristic of anaerobic ammonium-oxidizing (anammox) bacteria in an anaerobic biological filtrated (ABF) reactor. Appl Microbiol Biotechnol 70:47–52

  6. Mulder A, Van de Graaf AA, Robertson LA, Kuenen JG (1995) Anaerobic ammonium oxidation discovered in a denitrifying fluidized bed reactor. FEMS Microbiol Ecol 16:177–184

  7. Neef A, Amann R, Schlesner H, Schleifer KH (1998) Monitoring a widespread bacterial group: in situ detection of planctomycetes with 16S rRNA-targeted probes. Microbiology 144:3257–3266

  8. Noda N, Ikuta H, Ebie Y, Hirata A, Tsuneda S, Matsumura M, Sumino T, Inamori Y (2000) Rapid quantification and in situ detection of nitrifying bacteria in biofilms by monoclonal antibody method. Water Sci Technol 65:3182–3191

  9. Okabe S, Satoh H, Watanabe Y (1999) In situ analysis of nitrifying biofilms as determined by in situ hybridization and the use of microelectrodes. Appl Environ Microbiol 65:3182–3191

  10. Saiki Y, Iwabuchi C, Katami A, Kitagawa Y (2002) Microbial analyses by fluorescence in situ hybridization of well-settled granular sludge in brewery wastewater treatment plants. J Biosci Bioeng 93:601–606

  11. Sliekers AO, Third KA, Abma W, Kuenen JG, Jetten MSM (2003) CANON and anammox in a gas-lift reactor. FEMS Microbiol Lett 218:339–344

  12. Strous M, Van Gerven E, Zheng P, Kuenen JG, Jetten MSM (1997) Ammonium removal from concentrated waste streams with the anaerobic ammonium oxidation (ANAMMOX) process in different reactor configurations. Water Res 31:1955–1962

  13. Strous M, Heijnen JJ, Kuenen JG, Jetten MSM (1998) The sequencing batch reactor as a powerful tool for the study of slowly growing anaerobic ammonium-oxidizing microorganisms. Appl Microbiol Biotechnol 50:589–596

  14. Strous M, Fuerst JA, Kramer EHM, Logemann S, Muyzer G, Van de Pas-Schoonen KT, Webb R, Kuenen JG, Jetten MSM (1999) Missing lithotroph identified as new planctomycete. Nature 400:446–449

  15. Sumino T, Nakamura H, Mori N (1992) Immobilization of activated sludge by polyethylene glycol prepolymer. J Ferment Bioeng 73:37–42

  16. Van de Graaf AA, de Bruijn P, Robertson LA, Jetten MSM, Kuenen JG (1996) Autotrophic growth of anaerobic ammonium-oxidizing microorganisms in a fluidized bed reactor. Microbiol 142:2187–2196

  17. Van de Graaf AA, Peter de Bruijn, Robertson LA, Jetten MSM, Kuenen JG (1997) Metabolic pathway of anaerobic ammonium oxidation on the basis of 15N studies in a fluidized bed reactor. Microbiology 143:2415–2421

  18. Vogelsang C, Husby A, Osgaard K (1997) Functional stability of temperature-compensated nitrification in domestic wastewater treatment obtained with PVA-SBQ/alginate gel entrapment. Water Res 31:1659–1664

  19. Weatherburn MW (1967) Phenol-hypochlorite reaction for determination of ammonia. J Anal Chem 39:971–974

Download references

Author information

Correspondence to Kazuichi Isaka.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Isaka, K., Date, Y., Sumino, T. et al. Ammonium removal performance of anaerobic ammonium-oxidizing bacteria immobilized in polyethylene glycol gel carrier. Appl Microbiol Biotechnol 76, 1457–1465 (2007).

Download citation


  • Immobilization
  • Anammox
  • Ammonium
  • Nitrite
  • Nitrogen removal
  • Gel entrapment