Abstract

Nitrification, the oxidation of ammonia via nitrite to nitrate, has always been considered to be a two-step process catalysed by chemolithoautotrophic microorganisms oxidizing either ammonia or nitrite. No known nitrifier carries out both steps, although complete nitrification should be energetically advantageous. This functional separation has puzzled microbiologists for a century. Here we report on the discovery and cultivation of a completely nitrifying bacterium from the genus Nitrospira, a globally distributed group of nitrite oxidizers. The genome of this chemolithoautotrophic organism encodes the pathways both for ammonia and nitrite oxidation, which are concomitantly activated during growth by ammonia oxidation to nitrate. Genes affiliated with the phylogenetically distinct ammonia monooxygenase and hydroxylamine dehydrogenase genes of Nitrospira are present in many environments and were retrieved on Nitrospira-contigs in new metagenomes from engineered systems. These findings fundamentally change our picture of nitrification and point to completely nitrifying Nitrospira as key components of nitrogen-cycling microbial communities.

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European Nucleotide Archive

Data deposits

All raw sequence data is available in the European Nucleotide Archive (ENA) under the project accession number PRJEB10139. The genome sequence of Ca. N. inopinata has been deposited at ENA under the project PRJEB10818, sequence accession LN885086. The draft genome of the betaproteobacterium from ENR4 and ENR6 is available in the JGI Integrated Microbial Genomes Database (https://img.jgi.doe.gov/cgi-bin/m/main.cgi) under the IMG Genome ID 2636415980.

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Acknowledgements

We thank T. K. Lee and M. Steinberger for help with PCR analyses, N. V. Grigor’eva and M. Pogoda for assistance with culture maintenance, N. A. Kostrikina for assistance with electron microscopy, K. Kitzinger for support with FISH analyses, M. Mooshammer for help with chemical analyses, R. Hatzenpichler for designing probe Nmir1009, K. Eismann for help with proteome sample preparation, B. Scheer for help with mass spectrometer maintenance, Purena GmbH (Wolfenbüttel, Germany) for cooperation, N. Chernyh and J. Rosenthal for taking samples, and H. Koch and E. Bock for discussion. The authors are grateful for using the analytical facilities of the Centre for Chemical Microscopy (ProVIS) (Helmholtz Centre for Environmental Research), which is headed by H. Richnow (Department of Isotope Biochemistry) and supported by European Regional Development Funds (EFRE–Europe funds Saxony) and the Helmholtz Association. P.P. and H.D. were supported by the Austrian Science Fund (FWF) projects P27319-B21 and P25231-B21 (to H.D.). M.P., J.V., P.H., and M.W. were supported by the European Research Council Advanced Grant project NITRICARE 294343 (to M.W.). M.A., R.H.K., and P.H.N. were supported by the Danish Council for Independent Research, DFF – 4005-00369 and Innovation Fund Denmark (EcoDesign).

Author information

Affiliations

  1. Department of Microbiology and Ecosystem Science, Division of Microbial Ecology, University of Vienna, Althanstrasse 14, 1090 Vienna, Austria

    • Holger Daims
    • , Petra Pjevac
    • , Ping Han
    • , Craig Herbold
    • , Marton Palatinszky
    • , Julia Vierheilig
    •  & Michael Wagner
  2. Winogradsky Institute of Microbiology, Research Center of Biotechnology of the Russian Academy of Sciences, Leninsky Ave. 33, bld. 2, 119071 Moscow, Russia

    • Elena V. Lebedeva
    •  & Alexandr Bulaev
  3. Center for Microbial Communities, Department of Chemistry and Bioscience, Aalborg University, Fredrik Bajers Vej 7H, 9220 Aalborg, Denmark

    • Mads Albertsen
    • , Rasmus H. Kirkegaard
    •  & Per H. Nielsen
  4. Helmholtz-Centre for Environmental Research - UFZ, Department of Proteomics, Permoserstrasse 15, 04318 Leipzig, Germany

    • Nico Jehmlich
    •  & Martin von Bergen
  5. Helmholtz-Centre for Environmental Research - UFZ, Department of Metabolomics, Permoserstrasse 15, 04318 Leipzig, Germany

    • Martin von Bergen
  6. Department of Microbiology and Ecosystem Science, Division of Computational Systems Biology, University of Vienna, Althanstrasse 14, 1090 Vienna, Austria

    • Thomas Rattei
  7. DVGW-Forschungsstelle TUHH, Hamburg University of Technology, 21073 Hamburg, Germany

    • Bernd Bendinger

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Contributions

H.D. did (meta)genomic analysis of Ca. N. inopinata and comammox Nitrospira, contributed to phylogenetic and proteomics data analyses, designed the study and wrote the paper; E.V.L. enriched Ca. N. inopinata; E.V.L., P.P., P.H., A.B. and M.P. performed physiological experiments, analysed data, and characterized enrichments; M.A., R.H.K. and P.H.N. carried out metagenome sequencing, assembly and binning; C.H. performed bioinformatic and phylogenetic analyses; N.J. and M.vB. performed proteomics measurements and data analysis; T.R. performed bioinformatic analyses; P.H., M.P. and J.V. maintained enrichment cultures and performed experiments; J.V. carried out database analyses; B.B. organized sampling and characterized environmental samples; M.W. designed the study, analysed data, and wrote the paper. All authors discussed the results and commented the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Michael Wagner.

Extended data

Supplementary information

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  1. 1.

    Supplementary Tables

    This file contains Supplementary Tables 1-3 and Supplementary Table 8.

Excel files

  1. 1.

    Supplementary Table 4

    This file contains Supplementary Table 4, which lists marker genes and their copy numbers detected by CheckM in the closed Ca. N. inopinata genome.

  2. 2.

    Supplementary Table 5

    This file contains Supplementary Table 5, which lists marker genes and their copy numbers detected by CheckM in the genome of the betaproteobacterium found in enrichment cultures ENR4 and ENR6.

  3. 3.

    Supplementary Table 6

    This file contains Supplementary Table 6, which lists marker genes and their copy numbers detected by CheckM in the genome of the alphaproteobacterium found in enrichment culture ENR4.

  4. 4.

    Supplementary Table 7

    This file contains Supplementary Table 7, which lists marker genes and their copy numbers detected by CheckM in the genome of the actinobacterium found in enrichment culture ENR4.

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https://doi.org/10.1038/nature16461

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