Biotechnology and Bioprocess Engineering

, Volume 19, Issue 2, pp 231–238 | Cite as

Isolation and characterization of a novel heterotrophic nitrifying and aerobic denitrifying bacterium Pseudomonas stutzeri KTB for bioremediation of wastewater

  • Maohong Zhou
  • Hairen Ye
  • Xiaowei ZhaoEmail author
Research Paper


A novel heterotrophic nitrifying and aerobic denitrifying bacterium, KTB, was isolated from activated sludge flocci collected from a biological aerated filter according to the modified Takaya method and identified as Pseudomonas stutzeri by 16S rDNA gene sequence analysis. When shaking-cultured in the presence of 4.331 mmol/L of nitrate, 4.511 mmol/L of nitrite and 4.438 mmol/L of ammonium, the strain grew fast, with μmax being 0.42, 0.45, and 0.56/h, and displayed high nitrogen removal efficiency, with nitrogen removal rate being 0.239, 0.362, and 0.361 mmol/L/h and nitrogen removal ratio being 99.1, 100.0, and 100.0% in 18 h, respectively. The removal mainly occurred in the logarithmic phase. Nitrite accumulation did not affect denitrification performance. Nitrate concentration was below the detectable limit during the whole growth cycle when ammonium was used as sole nitrogen source. It tolerated high DO level and exhibited excellent aggregation ability. A possible pathway involved in the nitrogen removal process, which demonstrated a full nitrification and denitrification route, was speculated. The strain might be a great candidate for biological removal of nitrogen compounds from wastewater.


nitriogen removal heterotrophic nitrification aerobic denitrification aggregation Pseudomonas stutzeri 


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  1. 1.
    Zhu, L., W. Ding, L. J. Feng, Y. Kong, J. Xu, and X. Y. Xu (2012) Isolation of aerobic denitrifiers and characterization for their potential application in the bioremediation of oligotrophic ecosystem. Bioresour. Technol. 108: 1–7.CrossRefGoogle Scholar
  2. 2.
    Yang, X. P., S. M. Wang, D. W. Zhang, and L. X. Zhou (2011) Isolation and nitrogen removal characteristics of an aerobic heterotrophic nitrifying-denitrifying bacterium Bacillus subtilis A1. Bioresour. Technol. 102: 854–862.CrossRefGoogle Scholar
  3. 3.
    Chen, Q. and J. R. Ni (2011) Heterotrophic nitrification-aerobic denitrification by novel isolated bacteria. J. Ind. Microbiol. Biotechnol. 38: 1305–1310.CrossRefGoogle Scholar
  4. 4.
    Ludwig, W., G. Mittenhuber, and C. G. Friedrich (1993) Transfer of Thiosphaera pantotropha to Paracoccus denitrificans. Int. J. Syst. Bacteriol. 43: 363–367.CrossRefGoogle Scholar
  5. 5.
    Rainey, F. A., D. P. Kelly, E. Stackebrandt, J. Burghardt, A. Hiraishi, Y. Katayama, and A. P. Wood (1999) A re-evaluation of the taxonomy of Paracoccus denitrificans and a proposal for the combination Paracoccus pantotrophus comb. Nov. Int. J. Syst. Bacteriol. 49: 645–651.CrossRefGoogle Scholar
  6. 6.
    Robertson, L. A. and J. G. Kuenen (1983) Thiosphaera pantotropha gen. nov. sp. nov., a facultatively anaerobic, facultatively autotrophic sulfur bacterium. J. Gen. Microbiol. 129: 2847–2855.Google Scholar
  7. 7.
    Taylor, S. M., Y. L. He, B. Zhao, and J. Huang (2009) Heterotrophic ammonium removal characteristics of an aerobic heterotrophic nitrifying-denitrifying bacterium, Providencia rettgeri YL. J. Environ. Sci. 21: 1336–1341.CrossRefGoogle Scholar
  8. 8.
    Padhi, S. K., S. Tripathy, R. Sen, A. S. Mahapatra, S. Mohanty, and N. K. Maiti (2013) Characterisation of heterotrophic nitrifying and aerobic denitrifying Klebsiella pneumoniae CF-S9 strain for bioremediation of wastewater. Int. Biodeter. Biodegr. 78: 67–73.CrossRefGoogle Scholar
  9. 9.
    Takaya, N., M. A. B. Catalan-Sakairi, Y. Sakaguchi, I. Kato, Z. M. Zhou, and H. Shoun (2003) Aerobic denitrifying bacteria that produce low levels of nitrous oxide. Appl. Environ. Microbiol. 69: 3152–3157.CrossRefGoogle Scholar
  10. 10.
    Frear, D. S. and R. C. Burrell (1955) Spectrophotometric method for determining hydroxylamine reductase activity in higher plants. Anal. Chem. 27: 1664–1665.CrossRefGoogle Scholar
  11. 11.
    Michal, G., H. O. Beutler, G. Lang, and U. Guentner (1976) Enzymatic determination of succinic acid in foodstuffs. Fresenius Z. Anal. Chem. 279: 137–138.CrossRefGoogle Scholar
  12. 12.
    Rickard, A. H., S. A. Leach, L. S. Hall, C. M. Buswell, N. J. High, and P. S. Handley (2002) Phylogenetic relationships and coaggregation ability of freshwater biofilm bacteria. Appl. Environ. Microbiol. 68: 3644–3650.CrossRefGoogle Scholar
  13. 13.
    Rosenberg, M., D. Gutnick, and E. Rosenberg (1980) Adherence of bacteria to hydrocarbons: a simple method for measuring cell-surface hydrophobicity. FEMS Microbiol. Lett. 9: 29–33.CrossRefGoogle Scholar
  14. 14.
    Zhang, Q. L., Y. Liu, G. M. Ai, L. L. Miao, H. Y. Zheng, and Z. P. Liu (2012) The characteristics of a novel heterotrophic nitrification-aerobic denitrification bacterium, Bacillus methylotrophicus strain L7. Bioresour. Technol. 108: 35–44.CrossRefGoogle Scholar
  15. 15.
    Joo, H. S., M. Hirai, and M. Shoda (2005) Characteristics of ammonium removal by heterotrophic nitrification-aerobic denitrification by Alcaligenes faecalis no. 4. J. Biosci. Bioeng. 100: 184–191.CrossRefGoogle Scholar
  16. 16.
    Castignetti, D. and T. C. Hollocher (1984) Heterotrophic nitrification among denitrifiers. Appl. Environ. Microbiol. 47: 620–623.Google Scholar
  17. 17.
    Marshall, K. C. and M. Alexander (1962) Nitrification by Aspergillus flavus. J. Bacteriol. 83: 572–577.Google Scholar
  18. 18.
    Oh, J. and J. Silverstein (1999) Acetate limitation and nitrite accumulation during denitrification. J. Environ. Eng. 125: 234–242.CrossRefGoogle Scholar
  19. 19.
    Münch, E. V., P. Lant, and J. Keller (1996) Simultaneous nitrification and denitrification in bench-scale sequencing batch reactors. Water Res. 30: 277–284.CrossRefGoogle Scholar
  20. 20.
    Conley, D. J., H. W. Paerl, R. W. Howarth, D. F. Boesch, S. P. Seitzinger, K. E. Havens, C. Lancelot, and G. E. Likens (2009) Controlling eutrophication, nitrogen and phosphorus. Sci. 323: 1014–1015.CrossRefGoogle Scholar
  21. 21.
    Robertson, L. A. and J. G. Kuenen (1990) Combined heterotrophic nitrification and aerobic denitrification in Thiosphaera pantotropha and other bacteria. Antonie Van Leeuwenhoek 57: 139–152.CrossRefGoogle Scholar
  22. 22.
    van Niel, E. W. J., K. J. Braber, L. A. Robertson, and J. G. Kuenen (1992) Heterotrophic nitrification and aerobic denitrification in Alcaligenes-faecalis strain TUD. Antonie Van Leeuwenhoek 62: 231–237.CrossRefGoogle Scholar
  23. 23.
    Khardenavis, A. A., A. Kapley, and H. J. Purohit (2007) Simultaneous nitrification and denitrification by diverse Diaphorobacter sp. Appl. Microbiol. Biotechnol. 77: 403–409.CrossRefGoogle Scholar
  24. 24.
    Gibbs, B. M., K. A. Shephard, K. A. Third, and R. Cord-Ruwisch (2004) The presence of ammonium facilitates nitrite reduction under PHB driven simultaneous nitrification and denitrification. Water Sci. Technol. 50: 181–189.Google Scholar
  25. 25.
    Lloyd, D., L. Boddy, and K. J. P. Davies (1987) Persistence of bacterial denitrification capacity under aerobic conditions: The rule rather than the exception. FEMS Microbiol. Lett. 45: 185–190.CrossRefGoogle Scholar
  26. 26.
    Hochstein, L. I., M. Betlach, and G. Kritikos (1984) The effect of oxygen on denitrification during steady-state growth of Paracoccus halodenitrificans. Arch. Microbiol. 137: 74–78.CrossRefGoogle Scholar
  27. 27.
    Robertson, L. A., E. W. J. van Niel, R. A. M. Torremans, and J. G. Kuenen (1988) Simultaneous nitrification and denitrification in aerobic chemostat cultures of Thiosphaera pantotropha. Appl. Environ. Microbiol. 54: 2812–2818.Google Scholar
  28. 28.
    Mével, G. and D. Prieur (2000) Heterotrophic nitrification by a thermophilic Bacillus species as influenced by different culture conditions. Can. J. Microbiol. 46: 465–473.CrossRefGoogle Scholar
  29. 29.
    Chen, P. Z., J. Li, Q. X. Li, Y. C. Wang, S. P. Li, T. Z. Ren, and L. G. Wang (2012) Simultaneous heterotrophic nitrification and aerobic denitrification by bacterium Rhodococcus sp. CPZ24. Bioresour. Technol. 116: 266–270.CrossRefGoogle Scholar
  30. 30.
    Su, J. J., K. S. Yeh, and P. W. Tseng (2006) A Strain of Pseudomonas sp. isolated from piggery wastewater treatment systems with heterotrophic nitrification capability in Taiwan. Curr. Microbiol. 53: 77–81.CrossRefGoogle Scholar

Copyright information

© The Korean Society for Biotechnology and Bioengineering and Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  1. 1.Zhejiang Provincial Key Laboratory for Subtropical Water Environment and Marine Biological Resources Protection, College of Life and Environmental ScienceWenzhou UniversityWenzhouChina

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