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Application of stable‐isotope labelling techniques for the detection of active diazotrophs

Investigating active participants in the fixation of dinitrogen gas is vital as N is often a limiting factor for primary production. Biological nitrogen fixation is performed by a diverse guild of bacteria and archaea (diazotrophs), which can be free‐living or symbionts. Free‐living diazotrophs are...

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Autores principales: Angel, Roey, Panhölzl, Christopher, Gabriel, Raphael, Herbold, Craig, Wanek, Wolfgang, Richter, Andreas, Eichorst, Stephanie A., Woebken, Dagmar
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5814836/
https://www.ncbi.nlm.nih.gov/pubmed/29027346
http://dx.doi.org/10.1111/1462-2920.13954
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author Angel, Roey
Panhölzl, Christopher
Gabriel, Raphael
Herbold, Craig
Wanek, Wolfgang
Richter, Andreas
Eichorst, Stephanie A.
Woebken, Dagmar
author_facet Angel, Roey
Panhölzl, Christopher
Gabriel, Raphael
Herbold, Craig
Wanek, Wolfgang
Richter, Andreas
Eichorst, Stephanie A.
Woebken, Dagmar
author_sort Angel, Roey
collection PubMed
description Investigating active participants in the fixation of dinitrogen gas is vital as N is often a limiting factor for primary production. Biological nitrogen fixation is performed by a diverse guild of bacteria and archaea (diazotrophs), which can be free‐living or symbionts. Free‐living diazotrophs are widely distributed in the environment, yet our knowledge about their identity and ecophysiology is still limited. A major challenge in investigating this guild is inferring activity from genetic data as this process is highly regulated. To address this challenge, we evaluated and improved several (15)N‐based methods for detecting N(2) fixation activity (with a focus on soil samples) and studying active diazotrophs. We compared the acetylene reduction assay and the (15)N(2) tracer method and demonstrated that the latter is more sensitive in samples with low activity. Additionally, tracing (15)N into microbial RNA provides much higher sensitivity compared to bulk soil analysis. Active soil diazotrophs were identified with a (15)N‐RNA‐SIP approach optimized for environmental samples and benchmarked to (15)N‐DNA‐SIP. Lastly, we investigated the feasibility of using SIP‐Raman microspectroscopy for detecting (15)N‐labelled cells. Taken together, these tools allow identifying and investigating active free‐living diazotrophs in a highly sensitive manner in diverse environments, from bulk to the single‐cell level.
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spelling pubmed-58148362018-02-27 Application of stable‐isotope labelling techniques for the detection of active diazotrophs Angel, Roey Panhölzl, Christopher Gabriel, Raphael Herbold, Craig Wanek, Wolfgang Richter, Andreas Eichorst, Stephanie A. Woebken, Dagmar Environ Microbiol Research Articles Investigating active participants in the fixation of dinitrogen gas is vital as N is often a limiting factor for primary production. Biological nitrogen fixation is performed by a diverse guild of bacteria and archaea (diazotrophs), which can be free‐living or symbionts. Free‐living diazotrophs are widely distributed in the environment, yet our knowledge about their identity and ecophysiology is still limited. A major challenge in investigating this guild is inferring activity from genetic data as this process is highly regulated. To address this challenge, we evaluated and improved several (15)N‐based methods for detecting N(2) fixation activity (with a focus on soil samples) and studying active diazotrophs. We compared the acetylene reduction assay and the (15)N(2) tracer method and demonstrated that the latter is more sensitive in samples with low activity. Additionally, tracing (15)N into microbial RNA provides much higher sensitivity compared to bulk soil analysis. Active soil diazotrophs were identified with a (15)N‐RNA‐SIP approach optimized for environmental samples and benchmarked to (15)N‐DNA‐SIP. Lastly, we investigated the feasibility of using SIP‐Raman microspectroscopy for detecting (15)N‐labelled cells. Taken together, these tools allow identifying and investigating active free‐living diazotrophs in a highly sensitive manner in diverse environments, from bulk to the single‐cell level. John Wiley and Sons Inc. 2017-12-15 2018-01 /pmc/articles/PMC5814836/ /pubmed/29027346 http://dx.doi.org/10.1111/1462-2920.13954 Text en © 2017 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd This is an open access article under the terms of the Creative Commons Attribution (http://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
spellingShingle Research Articles
Angel, Roey
Panhölzl, Christopher
Gabriel, Raphael
Herbold, Craig
Wanek, Wolfgang
Richter, Andreas
Eichorst, Stephanie A.
Woebken, Dagmar
Application of stable‐isotope labelling techniques for the detection of active diazotrophs
title Application of stable‐isotope labelling techniques for the detection of active diazotrophs
title_full Application of stable‐isotope labelling techniques for the detection of active diazotrophs
title_fullStr Application of stable‐isotope labelling techniques for the detection of active diazotrophs
title_full_unstemmed Application of stable‐isotope labelling techniques for the detection of active diazotrophs
title_short Application of stable‐isotope labelling techniques for the detection of active diazotrophs
title_sort application of stable‐isotope labelling techniques for the detection of active diazotrophs
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5814836/
https://www.ncbi.nlm.nih.gov/pubmed/29027346
http://dx.doi.org/10.1111/1462-2920.13954
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