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 Deltaproteobacteria 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 ^(15)NH^+_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. ^(15)N_2 incorporation was not observed in single DSS cells, but was detected in single DSB cells. Interestingly, ^(15)N_2 incorporation in single DSB cells was methane-dependent, raising the possibility that DSB cells acquired reduced ^(15)N products from diazotrophic ANME while spatially coupled, and then subsequently dissociated. 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.
Additional Information
© 2015 International Society for Microbial Ecology. This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article's Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/ Received 31 January 2015; Revised 29 May 2015; Accepted 5 July 2015; Advance online publication 22 September 2015. We thank the pilots, crew and science party of AT15-11 and AT15-59, especially Lisa Levin, Jake Bailey and Shana Goffredi; Abigail Green-Saxena and Joshua Steele for valuable discussions; Bethany Jenkins for consultation with RNA extraction protocol development; John Eiler and Yunbin Guan for assistance with NanoSIMS measurements. We additionally thank three anonymous reviewers for their careful critique of the manuscript. Funding was provided by the US Department of Energy, Office of Science, Office of Biological and Environmental Research (DE-SC0003940 and DE-SC0004949 to VJO), the National Science Foundation (MCB-0348492 to VJO and a Graduate Research Fellowship to AED) and the Gordon and Betty Moore Foundation (GBMF no. 3780 to VJO, and via support for The Caltech Center for Microanalysis housing the CAMECA NanoSIMS 50 L). The writing of this manuscript by AED was partially performed while funded by a Lawrence Postdoctoral Fellowship under the auspices of the US Department of Energy by Lawrence Livermore National Laboratory under Contract DE-AC52-07NA27344. The authors declare no conflict of interest.Attached Files
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Additional details
- PMCID
- PMC4817681
- Eprint ID
- 60573
- Resolver ID
- CaltechAUTHORS:20150928-110118499
- DE-SC0003940
- Department of Energy (DOE)
- DE-SC0004949
- Department of Energy (DOE)
- MCB-0348492
- NSF
- NSF Graduate Research Fellowship
- 3780
- Gordon and Betty Moore Foundation
- DE-AC52-07NA27344
- Department of Energy (DOE)
- Created
-
2015-09-29Created from EPrint's datestamp field
- Updated
-
2021-11-10Created from EPrint's last_modified field
- Caltech groups
- Division of Geological and Planetary Sciences