Skip to main content
Log in

Identification and monitoring of nitrification and denitrification genes in Klebsiella pneumoniae EGD-HP19-C for its ability to perform heterotrophic nitrification and aerobic denitrification

  • Original Paper
  • Published:
Functional & Integrative Genomics Aims and scope Submit manuscript

Abstract

Microbes capable of performing heterotrophic nitrification and aerobic denitrification simultaneously have application in nitrogen level management in effluent treatment plants. Klebsiella pneumoniae EGD-HP19-C is a metabolically versatile bacterium capable of utilising NH3–N, NO2–N and NO3–N as sole sources of nitrogen. The annotation was done for the genes involved in N-assimilation and N-dissimilation pathways from the draft genome sequences of this bacterium (NCBI GenBank accession no. AUTW02000000.1). The sequence data also suggested possible existence of plasmid associated with this bacterium. Multiple gene sequence alignments of glutamine synthetase (gln), hydroxylamine reductase (har), nitrite reductase (nir), nitric oxide reductase (nor), assimilatory nitrate reductase (nas) and respiratory nitrate reductase (nar) genes from EGD-HP19-C genome were performed to compare sequence identities with that of closely related bacterial species. The metabolic pathways were mapped using KAAS and 3D structures for representative enzyme sub-units were also elucidated. The study suggested that the organism, though it has incomplete nitrification and denitrification pathways still removes the inorganic nitrogen content from the system via ammonification reaction.

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

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

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

Similar content being viewed by others

References

  • Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215(3):403–410

    Article  CAS  PubMed  Google Scholar 

  • APHA, AWWA, and WEF, (2005) Standard methods for the examination of water and wastewater, 21st ed. American Public Health Association, Washington, D.C

  • Aziz RK, Bartels D, Best AA et al (2008) The RAST server: rapid annotations using subsystems technology. BMC Genomics 9:75

    Article  PubMed Central  PubMed  Google Scholar 

  • Berman HM (2008) The protein data bank: a historical perspective. Acta Crystallogr A 64(Pt 1):88–95

    Article  CAS  PubMed  Google Scholar 

  • Chen Y, Zhou W, Li Y et al (2014) Nitrite reductase genes as functional markers to investigate diversity of denitrifying bacteria during agricultural waste composting. Appl Microbiol Biotechnol 98(9):4233–4243

    Article  CAS  PubMed  Google Scholar 

  • Darling AC, Mau B, Blattner FR, Perna NT (2004) Mauve: multiple alignment of conserved genomic sequence with rearrangements. Genome Res 14(7):1394–1403

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • De AK (2003) Environmental chemistry, 5th ed. New Age International

  • Dias Z, Dias U, Setubal JC (2012) SIS: a program to generate draft genome sequence scaffolds for prokaryotes. BMC Bioinforma 13:96

    Article  Google Scholar 

  • EPA (1993) Nitrogen control. USEPA, Washington, DC

  • Ge S, Peng Y, Qiu S, Zhu A, Ren N (2014) Complete nitrogen removal from municipal wastewater via partial nitrification by appropriately alternating anoxic/aerobic conditions in a continuous plug-flow step feed process. Water Res 15; 55: 95–105

  • Hall TA (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucl Acids Symp Ser 41:95–98

    CAS  Google Scholar 

  • Joo HS, Hirai M, Shoda M (2005) Characteristics of ammonium removal by heterotrophic nitrification-aerobic denitrification by Alcaligenes faecalis no. 4. J Biosci Bioeng 100(2):184–191

    Article  CAS  PubMed  Google Scholar 

  • Kapley A, Purohit HJ (2009) Diagnosis of treatment efficiency in industrial wastewater treatment plants: a case study at a refinery ETP. Environ Sci Technol 43(10):3789–3795

    Article  CAS  PubMed  Google Scholar 

  • Kelley LA, Sternberg MJ (2009) Protein structure prediction on the Web: a case study using the Phyre server. Nat Protoc 4(3):363--71

  • Khardenavis AA, Kapley A, Purohit HJ (2007) Simultaneous nitrification and denitrification by diverse Diaphobacter sp. Appl Microbiol Biotechnol 77:403–409

    Article  CAS  PubMed  Google Scholar 

  • Kim YJ, Yoshizawa M, Takenaka S, Murakami S, Aoki K (2002) Isolation and culture conditions of a Klebsiella pneumoniae strain that can utilize ammonium and nitrate ions simultaneously with controlled iron and molybdate ion concentrations. Biosci Biotechnol Biochem 6(5):996–1001

    Article  Google Scholar 

  • Kim M, Jeong SY, Yoon SJ et al (2008) Aerobic denitrification of Pseudomonas putida AD-21 at different C/N ratios. J Biosci Bioeng 106(5):498–502

    Article  CAS  PubMed  Google Scholar 

  • Laskowski RA, MacArthur MW, Moss DS, Thornton JM (1993) PROCHECK: a program to check the stereochemical quality of protein structures. J Appl Cryst 26:283–291

    Article  CAS  Google Scholar 

  • Lin JT, Goldman BS, Stewart V (1993) Structures of genes nasA and nasB, encoding assimilatory nitrate and nitrite reductases in klebsiella pneumoniae M5al. J Bacteriol 175(8):2370–2378

    CAS  PubMed Central  PubMed  Google Scholar 

  • Liu Y, Zang Y, Long X et al (2013) High nitrite removal capacity of an aerobic denitrifier Klebsiella oxytoca DF-1 isolated from aquaculture ponds in coastal mudflats. Afr J Microbiol Res 7(16):1527–1534

    CAS  Google Scholar 

  • Liu P, Lia P, Jiangb X, Bia D, Xiea Y, Taia C, Denga Z, Kumar R, Oua H (2012) Complete genome sequence of Klebsiella pneumoniae subsp. pneumoniae HS11286, a multidrug-resistant strain isolated from human sputum. J Bacteriol 194(7):1841–1842

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Minoru K, Susumu G, Yoko S, Miho F, Mao T (2000) KEGG: Kyoto encyclopaedia of genes and genomes. Nucleic Acids Res 28:27–30

    Article  Google Scholar 

  • Moriya Y, Itoh M, Okuda S, Yoshizawa A, Kanehisa M (2007) KAAS: an automatic genome annotation and pathway reconstruction server. Nucleic Acids Res 35:W182–W185

    Article  PubMed Central  PubMed  Google Scholar 

  • Nagore D, Sanz B, Soria J, Llarena M, Llama MJ, Calvete JJ, Serra JL (2006) The nitrate/nitrite ABC transporter of Phormidium laminosum: phosphorylation state of NrtA is not involved in its substrate binding activity. Biochim Biophys Acta 1760(2):172--81

  • Notredame C, Higgins DG, Heringa J (2000) T-coffee: a novel method for fast and accurate multiple sequence alignment. J Mol Biol 302:205–217

    Article  CAS  PubMed  Google Scholar 

  • Padhi SK, Tripathy S, Sen R, Mahapatra AS, MohantyS MNK (2013) Characterisation of heterotrophic nitrifying and aerobic denitrifying Klebsiella pneumoniae CF-S9 strain for bioremediation of wastewater. Int Biodeterior Biodegrad 78:67–73

    Article  CAS  Google Scholar 

  • Paliwal V, Puranik S, Purohit HJ (2012) Integrated perspective for effective bioremediation. Appl Biochem Biotechnol 166(4):903–924

    Article  CAS  PubMed  Google Scholar 

  • Patureau D, Zumstein E, Delgenes JP, Moletta R (2000) Aerobic denitrifiers isolated from diverse natural and managed ecosystem. Microb Ecol 39:145–152

    Article  CAS  PubMed  Google Scholar 

  • Phale PS, Paliwal V, Raju SC, Modak A, Purohit HJ (2013) Genome sequence of naphthalene-degrading soil bacterium Pseudomonas putida CSV86. Genome Announc 1(1):e00234–12. doi:10.1128/genomeA.00234-12

    Article  PubMed Central  PubMed  Google Scholar 

  • Purohit HJ, Raje DV, Kapley A, Padmanabhan P, Singh RN (2003) Genomics tools in environmental impact assessment. Environ Sci Technol 37(19):356A–363A

    Article  PubMed  Google Scholar 

  • Roberts KR, Kessler AJ, Grace MR, Cook PLM (2014) Increased rates of dissimilatory nitrate reduction to ammonium (DNRA) under oxic conditions in a periodically hypoxic estuary. Geochim Cosmochim Acta 133: 313--324

  • Robertson LA, Van Niel EWJ, Torremans RAM, Kuenen JG (1988) Simultaneous nitrification and denitrification in aerobic chemostat cultures of Thiosphaera pantotropha. Appl Environ Microbiol 54(11):2812–2818

    CAS  PubMed Central  PubMed  Google Scholar 

  • Shi W, Lu W, Liu Q, Zhi Y, Zhou P (2014) The identification of the nitrate assimilation related genes in the novel Bacillus megaterium NCT-2 accounts for its ability to use nitrate as its only source of nitrogen. Funct Integr Genomics 14(1):219--27

  • Su JJ, Liu BY, Liu CY (2001) Comparison of aerobic denitrification under high oxygen atmosphere by Thiosphaera pantotropha ATCC 35512 and Pseudomonas stutzeri SU2 newly isolated from the activated sludge of a piggery wastewater treatment system. J Appl Microbiol 90:457–462

    Article  CAS  PubMed  Google Scholar 

  • Wu Q, Stewart V (1998) NasFED proteins mediate assimilatory nitrate and nitrite transport in Klebsiella oxytoca (pneumoniae) M5al. J Bacteriol 180(5):1311–1322

    CAS  PubMed Central  PubMed  Google Scholar 

  • Wu S, Zhu Z, Fu L, Niu B, Li W (2011) WebMGA: a customizable web server for fast metagenomic sequence analysis. BMC Genomics 12:444

    Article  PubMed Central  PubMed  Google Scholar 

  • Zhao B, Liang He Y, Hughes J, Zhang XF (2010) Heterotrophic nitrogen removal by a newly isolated Acinetobacter calcoaceticus HNR. Bioresour Technol 101:5194–5200

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

Mr. Rajesh Pal, acknowledges the Senior Research Fellowship (SRF) from the University Grant Commission (UGC) of India for carrying out this work. Grant from CSIR network project ESC0108 is also gratefully acknowledged. Authors would also like to thank Mr. Ravi P. More and Mr. Ramkrishna Ojha for their constructive suggestions during discussions on the present study.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Hemant J. Purohit.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary Fig. 1

(DOCX 15 kb)

Supplementary Fig. 2

(DOCX 474 kb)

Supplementary Fig. 3

(DOCX 99 kb)

Supplementary Fig. 4

(DOCX 1398 kb)

Supplementary Fig. 5

(DOCX 1128 kb)

Supplementary Fig. 6

(DOCX 1496 kb)

Supplementary Fig. 7

(DOCX 347 kb)

Supplementary Table 1

(DOCX 15 kb)

Supplementary Table 2

(DOCX 15 kb)

Supplementary Table 3

(DOCX 15 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Pal, R.R., Khardenavis, A.A. & Purohit, H.J. Identification and monitoring of nitrification and denitrification genes in Klebsiella pneumoniae EGD-HP19-C for its ability to perform heterotrophic nitrification and aerobic denitrification. Funct Integr Genomics 15, 63–76 (2015). https://doi.org/10.1007/s10142-014-0406-z

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10142-014-0406-z

Keywords

Navigation