skip to main content
DOE PAGES title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Identification of a novel cyanobacterial group as active diazotrophs in a coastal microbial mat using NanoSIMS analysis

Abstract

N2 fixation is a key process in photosynthetic microbial mats to support the nitrogen demands associated with primary production. Despite its importance, groups that actively fix N2 and contribute to the input of organic N in these ecosystems still remain largely unclear. To investigate the active diazotrophic community in microbial mats from the Elkhorn Slough estuary, Monterey Bay, CA, USA, we conducted an extensive combined approach, including biogeochemical, molecular and high-resolution secondary ion mass spectrometry (NanoSIMS) analyses. Detailed analysis of dinitrogenase reductase (nifH) transcript clone libraries from mat samples that fixed N2 at night indicated that cyanobacterial nifH transcripts were abundant and formed a novel monophyletic lineage. Independent NanoSIMS analysis of 15N2-incubated samples revealed significant incorporation of 15N into small, non-heterocystous cyanobacterial filaments. Mat-derived enrichment cultures yielded a unicyanobacterial culture with similar filaments (named Elkhorn Slough Filamentous Cyanobacterium-1 (ESFC-1)) that contained nifH gene sequences grouping with the novel cyanobacterial lineage identified in the transcript clone libraries, displaying up to 100% amino-acid sequence identity. The 16S rRNA gene sequence recovered from this enrichment allowed for the identification of related sequences from Elkhorn Slough mats and revealed great sequence diversity in this cluster. Furthermore, by combining 15N2 tracer experiments, fluorescence in situmore » hybridization and NanoSIMS, in situ N2 fixation activity by the novel ESFC-1 group was demonstrated, suggesting that this group may be the most active cyanobacterial diazotroph in the Elkhorn Slough mat. Pyrotag sequences affiliated with ESFC-1 were recovered from mat samples throughout 2009, demonstrating the prevalence of this group. Here, this work illustrates that combining standard and single-cell analyses can link phylogeny and function to identify previously unknown key functional groups in complex ecosystems.« less

Authors:
 [1];  [2];  [3];  [3];  [3];  [4];  [5];  [4];  [6]
  1. Stanford Univ., CA (United States). Dept. of Chemical Engineering, and of Civil and Environmental Engineering; NASA Ames Research Center (ARC), Moffett Field, Mountain View, CA (United States). Exobiology Branch; Univ. of Vienna (Austria). Dept. of Microbial Ecology
  2. Stanford Univ., CA (United States). Dept. of Chemical Engineering, and of Civil and Environmental Engineering; NASA Ames Research Center (ARC), Moffett Field, Mountain View, CA (United States). Exobiology Branch
  3. NASA Ames Research Center (ARC), Moffett Field, Mountain View, CA (United States). Exobiology Branch
  4. Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States). Physical and Life Sciences Directorate
  5. Stanford Univ., CA (United States). Dept. of Chemical Engineering, and of Civil and Environmental Engineering
  6. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Earth Sciences Division
Publication Date:
Research Org.:
Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
Sponsoring Org.:
USDOE; German Research Foundation (DFG)
OSTI Identifier:
1396226
Report Number(s):
LLNL-JRNL-501172
Journal ID: ISSN 1751-7362
Grant/Contract Number:  
AC52-07NA27344; AC02-05CH11231; SCW1039
Resource Type:
Accepted Manuscript
Journal Name:
The ISME Journal
Additional Journal Information:
Journal Volume: 6; Journal Issue: 7; Journal ID: ISSN 1751-7362
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 54 ENVIRONMENTAL SCIENCES; 58 GEOSCIENCES; cyanobacteria; dinitrogenase reductase (nifH); microbial mats; NanoSIMS; N2 fixation

Citation Formats

Woebken, Dagmar, Burow, Luke C., Prufert-Bebout, Leslie, Bebout, Brad M., Hoehler, Tori M., Pett-Ridge, Jennifer, Spormann, Alfred M., Weber, Peter K., and Singer, Steven W. Identification of a novel cyanobacterial group as active diazotrophs in a coastal microbial mat using NanoSIMS analysis. United States: N. p., 2012. Web. doi:10.1038/ismej.2011.200.
Woebken, Dagmar, Burow, Luke C., Prufert-Bebout, Leslie, Bebout, Brad M., Hoehler, Tori M., Pett-Ridge, Jennifer, Spormann, Alfred M., Weber, Peter K., & Singer, Steven W. Identification of a novel cyanobacterial group as active diazotrophs in a coastal microbial mat using NanoSIMS analysis. United States. doi:10.1038/ismej.2011.200.
Woebken, Dagmar, Burow, Luke C., Prufert-Bebout, Leslie, Bebout, Brad M., Hoehler, Tori M., Pett-Ridge, Jennifer, Spormann, Alfred M., Weber, Peter K., and Singer, Steven W. Thu . "Identification of a novel cyanobacterial group as active diazotrophs in a coastal microbial mat using NanoSIMS analysis". United States. doi:10.1038/ismej.2011.200. https://www.osti.gov/servlets/purl/1396226.
@article{osti_1396226,
title = {Identification of a novel cyanobacterial group as active diazotrophs in a coastal microbial mat using NanoSIMS analysis},
author = {Woebken, Dagmar and Burow, Luke C. and Prufert-Bebout, Leslie and Bebout, Brad M. and Hoehler, Tori M. and Pett-Ridge, Jennifer and Spormann, Alfred M. and Weber, Peter K. and Singer, Steven W.},
abstractNote = {N2 fixation is a key process in photosynthetic microbial mats to support the nitrogen demands associated with primary production. Despite its importance, groups that actively fix N2 and contribute to the input of organic N in these ecosystems still remain largely unclear. To investigate the active diazotrophic community in microbial mats from the Elkhorn Slough estuary, Monterey Bay, CA, USA, we conducted an extensive combined approach, including biogeochemical, molecular and high-resolution secondary ion mass spectrometry (NanoSIMS) analyses. Detailed analysis of dinitrogenase reductase (nifH) transcript clone libraries from mat samples that fixed N2 at night indicated that cyanobacterial nifH transcripts were abundant and formed a novel monophyletic lineage. Independent NanoSIMS analysis of 15N2-incubated samples revealed significant incorporation of 15N into small, non-heterocystous cyanobacterial filaments. Mat-derived enrichment cultures yielded a unicyanobacterial culture with similar filaments (named Elkhorn Slough Filamentous Cyanobacterium-1 (ESFC-1)) that contained nifH gene sequences grouping with the novel cyanobacterial lineage identified in the transcript clone libraries, displaying up to 100% amino-acid sequence identity. The 16S rRNA gene sequence recovered from this enrichment allowed for the identification of related sequences from Elkhorn Slough mats and revealed great sequence diversity in this cluster. Furthermore, by combining 15N2 tracer experiments, fluorescence in situ hybridization and NanoSIMS, in situ N2 fixation activity by the novel ESFC-1 group was demonstrated, suggesting that this group may be the most active cyanobacterial diazotroph in the Elkhorn Slough mat. Pyrotag sequences affiliated with ESFC-1 were recovered from mat samples throughout 2009, demonstrating the prevalence of this group. Here, this work illustrates that combining standard and single-cell analyses can link phylogeny and function to identify previously unknown key functional groups in complex ecosystems.},
doi = {10.1038/ismej.2011.200},
journal = {The ISME Journal},
number = 7,
volume = 6,
place = {United States},
year = {2012},
month = {1}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record

Citation Metrics:
Cited by: 39 works
Citation information provided by
Web of Science

Save / Share:

Works referenced in this record:

Application of a nifH oligonucleotide microarray for profiling diversity of N 2 -fixing microorganisms in marine microbial mats
journal, October 2006


Carbon and nitrogen fluxes associated with the cyanobacterium Aphanizomenon sp. in the Baltic Sea
journal, April 2010

  • Ploug, Helle; Musat, Niculina; Adam, Birgit
  • The ISME Journal, Vol. 4, Issue 9
  • DOI: 10.1038/ismej.2010.53

In situ studies on N2 fixation using the acetylene reduction technique.
journal, November 1967

  • Stewart, W. D.; Fitzgerald, G. P.; Burris, R. H.
  • Proceedings of the National Academy of Sciences, Vol. 58, Issue 5
  • DOI: 10.1073/pnas.58.5.2071

Nitrogen fixation and transfer in open ocean diatom–cyanobacterial symbioses
journal, March 2011

  • Foster, Rachel A.; Kuypers, Marcel M. M.; Vagner, Tomas
  • The ISME Journal, Vol. 5, Issue 9
  • DOI: 10.1038/ismej.2011.26

A single-cell view on the ecophysiology of anaerobic phototrophic bacteria
journal, November 2008

  • Musat, N.; Halm, H.; Winterholler, B.
  • Proceedings of the National Academy of Sciences, Vol. 105, Issue 46
  • DOI: 10.1073/pnas.0809329105

Fluorescence In Situ Hybridization and Catalyzed Reporter Deposition for the Identification of Marine Bacteria
journal, June 2002


Nitrogen fixation associated with the cyanobacterial mat of a marine laminated microbial ecosystem
journal, January 1984

  • Stal, L. J.; Grossberger, S.; Krumbein, W. E.
  • Marine Biology, Vol. 82, Issue 3
  • DOI: 10.1007/BF00392402

Photosynthesis and structure of benthic microbial mats: Microelectrode and SEM studies of four cyanobacterial communities1: Photosynthesis of cyanobacterial mats
journal, November 1983

  • Jorgensen, Bo Barker; Revsbech, Niels Peter; Cohen, Yehuda
  • Limnology and Oceanography, Vol. 28, Issue 6
  • DOI: 10.4319/lo.1983.28.6.1075

Seasonal nitrogen fixation dynamics in a marine microbial mat: Potential roles of cyanobacteria and microheterotrophs
journal, May 1996

  • Paerl, Hans W.; Fitzpatrick, Matthew; Bebout, Brad M.
  • Limnology and Oceanography, Vol. 41, Issue 3
  • DOI: 10.4319/lo.1996.41.3.0419

Spatial and temporal variability in nitrogenase activity and diazotrophic community composition in coastal microbial mats
journal, November 2010

  • Severin, I.; Stal, Lj
  • Marine Ecology Progress Series, Vol. 417
  • DOI: 10.3354/meps08759

Aerobic nitrogen fixation in pure cultures of a benthic marine Oscillatoria (cyanobacteria)
journal, August 1981


Biogeochemical cycles of carbon, sulfur, and free oxygen in a microbial mat
journal, August 1993


Proteogenomic basis for ecological divergence of closely related bacteria in natural acidophilic microbial communities
journal, February 2010

  • Denef, V. J.; Kalnejais, L. H.; Mueller, R. S.
  • Proceedings of the National Academy of Sciences, Vol. 107, Issue 6
  • DOI: 10.1073/pnas.0907041107

Consortial N2 fixation: a strategy for meeting nitrogen requirements of marine and terrestrial cyanobacterial mats
journal, November 1996


Optimizing fluorescent in situ hybridization with rRNA-targeted oligonucleotide probes for flow cytometric identification of microorganisms
journal, January 1993


Simultaneous analysis of microbial identity and function using NanoSIMS
journal, March 2008


Unexpected Diversity and Complexity of the Guerrero Negro Hypersaline Microbial Mat
journal, May 2006


Fixation and fate of C and N in the cyanobacterium Trichodesmium using nanometer-scale secondary ion mass spectrometry
journal, March 2009

  • Finzi-Hart, J. A.; Pett-Ridge, J.; Weber, P. K.
  • Proceedings of the National Academy of Sciences, Vol. 106, Issue 15
  • DOI: 10.1073/pnas.0810547106

Spatial-temporal variability in diazotroph assemblages in Chesapeake Bay using an oligonucleotide nifH microarray: Chesapeake Bay nifH microarray
journal, April 2007


Microbial mats and the early evolution of life
journal, May 1990


Imaging and 3D Elemental Characterization of Intact Bacterial Spores by High-Resolution Secondary Ion Mass Spectrometry
journal, August 2008

  • Ghosal, Sutapa; Fallon, Stewart J.; Leighton, Terrance J.
  • Analytical Chemistry, Vol. 80, Issue 15
  • DOI: 10.1021/ac8006279

Nitrate sources and sinks in Elkhorn Slough, California: Results from long-term continuous in situ nitrate analyzers
journal, October 2004

  • Chapin, Thomas P.; Caffrey, Jane M.; Jannasch, Hans W.
  • Estuaries, Vol. 27, Issue 5
  • DOI: 10.1007/BF02912049

Comparison of diazotroph community structure in Lyngbya sp. and Microcoleus chthonoplastes dominated microbial mats from Guerrero Negro, Baja, Mexico
journal, March 2004


Cyanobacterial-bacterial mat consortia: examining the functional unit of microbial survival and growth in extreme environments
journal, February 2000


Regulation of nif gene expression and the energetics of N2 fixation over the diel cycle in a hot spring microbial mat
journal, March 2008

  • Steunou, Anne-Soisig; Jensen, Sheila I.; Brecht, Eric
  • The ISME Journal, Vol. 2, Issue 4
  • DOI: 10.1038/ismej.2007.117

The Domain-specific Probe EUB338 is Insufficient for the Detection of all Bacteria: Development and Evaluation of a more Comprehensive Probe Set
journal, September 1999


Microbial Community Diversity Associated with Carbon and Nitrogen Cycling in Permeable Shelf Sediments
journal, September 2006

  • Hunter, E. M.; Mills, H. J.; Kostka, J. E.
  • Applied and Environmental Microbiology, Vol. 72, Issue 9
  • DOI: 10.1128/AEM.03007-05

Nitrogenase gene diversity and microbial community structure: a cross-system comparison
journal, July 2003


Horizontal transfer of the nitrogen fixation gene cluster in the cyanobacterium Microcoleus chthonoplastes
journal, September 2009

  • Bolhuis, Henk; Severin, Ina; Confurius-Guns, Veronique
  • The ISME Journal, Vol. 4, Issue 1
  • DOI: 10.1038/ismej.2009.99

Quantitative Imaging of Nitrogen Fixation by Individual Bacteria Within Animal Cells
journal, September 2007


Co-occurrence of denitrification and nitrogen fixation in a meromictic lake, Lake Cadagno (Switzerland)
journal, August 2009


Determination of Nitrogen-Fixing Phylotypes in Lyngbya sp. and Microcoleus chthonoplastes Cyanobacterial Mats from Guerrero Negro, Baja California, Mexico
journal, April 2004


Nitrogen cycling in coastal marine ecosystems
journal, October 1999


Hydrogen production in photosynthetic microbial mats in the Elkhorn Slough estuary, Monterey Bay
journal, October 2011

  • Burow, Luke C.; Woebken, Dagmar; Bebout, Brad M.
  • The ISME Journal, Vol. 6, Issue 4
  • DOI: 10.1038/ismej.2011.142

Carbon and nitrogen fixation and metabolite exchange in and between individual cells of Anabaena oscillarioides
journal, July 2007

  • Popa, Radu; Weber, Peter K.; Pett-Ridge, Jennifer
  • The ISME Journal, Vol. 1, Issue 4
  • DOI: 10.1038/ismej.2007.44

    Works referencing / citing this record:

    NanoSIMS imaging of extracellular electron transport processes during microbial iron(III) reduction
    journal, June 2018

    • Newsome, Laura; Lopez Adams, Rebeca; Downie, Helen F.
    • FEMS Microbiology Ecology, Vol. 94, Issue 8
    • DOI: 10.1093/femsec/fiy104

    Processes and Microorganisms Involved in the Marine Nitrogen Cycle: Knowledge and Gaps
    journal, November 2019


    NanoSIMS imaging of extracellular electron transport processes during microbial iron(III) reduction
    journal, June 2018

    • Newsome, Laura; Lopez Adams, Rebeca; Downie, Helen F.
    • FEMS Microbiology Ecology, Vol. 94, Issue 8
    • DOI: 10.1093/femsec/fiy104

    Processes and Microorganisms Involved in the Marine Nitrogen Cycle: Knowledge and Gaps
    journal, November 2019