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Title: Activity and interactions of methane seep microorganisms assessed by parallel transcription and FISH-NanoSIMS analyses

Abstract

To characterize the activity and interactions of methanotrophic archaea (ANME) and Deltaproteo-bacteria at a methane-seeping mud volcano, we used two complimentary measures of microbial activity: a community-level analysis of the transcription of four genes (16S rRNA, methyl coenzyme M reductase A (mcrA), adenosine-5'-phosphosulfate reductase α-subunit (aprA), dinitrogenase reductase (nifH)), and a single-cell-level analysis of anabolic activity using fluorescence in situ hybridization coupled to nanoscale secondary ion mass spectrometry (FISH-NanoSIMS). Transcript analysis revealed that members of the deltaproteobacterial groups Desulfosarcina/Desulfococcus (DSS) and Desulfobulbaceae (DSB) exhibit increased rRNA expression in incubations with methane, suggestive of ANME-coupled activity. Direct analysis of anabolic activity in DSS cells in consortia with ANME by FISH-NanoSIMS confirmed their dependence on methanotrophy, with no 15NH$$+\atop{4}$$ assimilation detected without methane. In contrast, DSS and DSB cells found physically independent of ANME (i.e., single cells) were anabolically active in incubations both with and without methane. These single cells therefore comprise an active ‘free-living’ population, and are not dependent on methane or ANME activity. We investigated the possibility of N 2 fixation by seep Deltaproteobacteria and detected nifH transcripts closely related to those of cultured diazotrophic Deltaproteobacteria. However, nifH expression was methane-dependent. 15N 2 incorporation was not observed in single DSS cells, but was detected in single DSB cells. Interestingly, 15N 2 incorporation in single DSB cells was methane-dependent, raising the possibility that DSB cells acquired reduced 15N products from diazotrophic ANME while spatially coupled, and then subsequently dissociated. In conclusion, with this combined data set we address several outstanding questions in methane seep microbial ecosystems and highlight the benefit of measuring microbial activity in the context of spatial associations.

Authors:
 [1];  [1];  [1];  [1];  [1]
  1. California Inst. of Technology (CalTech), Pasadena, CA (United States). Division of Geological and Planetary Sciences
Publication Date:
Research Org.:
California Inst. of Technology (CalTech), Pasadena, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
OSTI Identifier:
1435726
Grant/Contract Number:  
SC0003940; SC0004949; AC52-07NA27344
Resource Type:
Accepted Manuscript
Journal Name:
The ISME Journal
Additional Journal Information:
Journal Volume: 10; Journal Issue: 3; Journal ID: ISSN 1751-7362
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Dekas, Anne E., Connon, Stephanie A., Chadwick, Grayson L., Trembath-Reichert, Elizabeth, and Orphan, Victoria J. Activity and interactions of methane seep microorganisms assessed by parallel transcription and FISH-NanoSIMS analyses. United States: N. p., 2015. Web. doi:10.1038/ismej.2015.145.
Dekas, Anne E., Connon, Stephanie A., Chadwick, Grayson L., Trembath-Reichert, Elizabeth, & Orphan, Victoria J. Activity and interactions of methane seep microorganisms assessed by parallel transcription and FISH-NanoSIMS analyses. United States. doi:10.1038/ismej.2015.145.
Dekas, Anne E., Connon, Stephanie A., Chadwick, Grayson L., Trembath-Reichert, Elizabeth, and Orphan, Victoria J. Tue . "Activity and interactions of methane seep microorganisms assessed by parallel transcription and FISH-NanoSIMS analyses". United States. doi:10.1038/ismej.2015.145. https://www.osti.gov/servlets/purl/1435726.
@article{osti_1435726,
title = {Activity and interactions of methane seep microorganisms assessed by parallel transcription and FISH-NanoSIMS analyses},
author = {Dekas, Anne E. and Connon, Stephanie A. and Chadwick, Grayson L. and Trembath-Reichert, Elizabeth and Orphan, Victoria J.},
abstractNote = {To characterize the activity and interactions of methanotrophic archaea (ANME) and Deltaproteo-bacteria at a methane-seeping mud volcano, we used two complimentary measures of microbial activity: a community-level analysis of the transcription of four genes (16S rRNA, methyl coenzyme M reductase A (mcrA), adenosine-5'-phosphosulfate reductase α-subunit (aprA), dinitrogenase reductase (nifH)), and a single-cell-level analysis of anabolic activity using fluorescence in situ hybridization coupled to nanoscale secondary ion mass spectrometry (FISH-NanoSIMS). Transcript analysis revealed that members of the deltaproteobacterial groups Desulfosarcina/Desulfococcus (DSS) and Desulfobulbaceae (DSB) exhibit increased rRNA expression in incubations with methane, suggestive of ANME-coupled activity. Direct analysis of anabolic activity in DSS cells in consortia with ANME by FISH-NanoSIMS confirmed their dependence on methanotrophy, with no 15NH$+\atop{4}$ assimilation detected without methane. In contrast, DSS and DSB cells found physically independent of ANME (i.e., single cells) were anabolically active in incubations both with and without methane. These single cells therefore comprise an active ‘free-living’ population, and are not dependent on methane or ANME activity. We investigated the possibility of N2 fixation by seep Deltaproteobacteria and detected nifH transcripts closely related to those of cultured diazotrophic Deltaproteobacteria. However, nifH expression was methane-dependent. 15N2 incorporation was not observed in single DSS cells, but was detected in single DSB cells. Interestingly, 15N2 incorporation in single DSB cells was methane-dependent, raising the possibility that DSB cells acquired reduced 15N products from diazotrophic ANME while spatially coupled, and then subsequently dissociated. In conclusion, with this combined data set we address several outstanding questions in methane seep microbial ecosystems and highlight the benefit of measuring microbial activity in the context of spatial associations.},
doi = {10.1038/ismej.2015.145},
journal = {The ISME Journal},
number = 3,
volume = 10,
place = {United States},
year = {2015},
month = {9}
}

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    Works referencing / citing this record:

    Synthetic microbial consortia for biosynthesis and biodegradation: promises and challenges
    journal, July 2019