Ecological Patterns of <italic toggle="yes">nifH</italic> Genes in Four Terrestrial Climatic Zones Explored with Targeted Metagenomics Using FrameBot, a New Informatics Tool

ABSTRACT Biological nitrogen fixation is an important component of sustainable soil fertility and a key component of the nitrogen cycle. We used targeted metagenomics to study the nitrogen fixation-capable terrestrial bacterial community by targeting the gene for nitrogenase reductase (nifH). We obt...

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Autores principales:	Qiong Wang, John F. Quensen, Jordan A. Fish, Tae Kwon Lee, Yanni Sun, James M. Tiedje, James R. Cole
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2013
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/6eb8ef141cfc40b3a4e8da84ddd638b1
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spelling	oai:doaj.org-article:6eb8ef141cfc40b3a4e8da84ddd638b12021-11-15T15:42:48ZEcological Patterns of <italic toggle="yes">nifH</italic> Genes in Four Terrestrial Climatic Zones Explored with Targeted Metagenomics Using FrameBot, a New Informatics Tool10.1128/mBio.00592-132150-7511https://doaj.org/article/6eb8ef141cfc40b3a4e8da84ddd638b12013-11-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00592-13https://doaj.org/toc/2150-7511ABSTRACT Biological nitrogen fixation is an important component of sustainable soil fertility and a key component of the nitrogen cycle. We used targeted metagenomics to study the nitrogen fixation-capable terrestrial bacterial community by targeting the gene for nitrogenase reductase (nifH). We obtained 1.1 million nifH 454 amplicon sequences from 222 soil samples collected from 4 National Ecological Observatory Network (NEON) sites in Alaska, Hawaii, Utah, and Florida. To accurately detect and correct frameshifts caused by indel sequencing errors, we developed FrameBot, a tool for frameshift correction and nearest-neighbor classification, and compared its accuracy to that of two other rapid frameshift correction tools. We found FrameBot was, in general, more accurate as long as a reference protein sequence with 80% or greater identity to a query was available, as was the case for virtually all nifH reads for the 4 NEON sites. Frameshifts were present in 12.7% of the reads. Those nifH sequences related to the Proteobacteria phylum were most abundant, followed by those for Cyanobacteria in the Alaska and Utah sites. Predominant genera with nifH sequences similar to reads included Azospirillum, Bradyrhizobium, and Rhizobium, the latter two without obvious plant hosts at the sites. Surprisingly, 80% of the sequences had greater than 95% amino acid identity to known nifH gene sequences. These samples were grouped by site and correlated with soil environmental factors, especially drainage, light intensity, mean annual temperature, and mean annual precipitation. FrameBot was tested successfully on three ecofunctional genes but should be applicable to any. IMPORTANCE High-throughput phylogenetic analysis of microbial communities using rRNA-targeted sequencing is now commonplace; however, such data often allow little inference with respect to either the presence or the diversity of genes involved in most important ecological processes. To study the gene pool for these processes, it is more straightforward to assess the genes directly responsible for the ecological function (ecofunctional genes). However, analyzing these genes involves technical challenges beyond those seen for rRNA. In particular, frameshift errors cause garbled downstream protein translations. Our FrameBot tool described here both corrects frameshift errors in query reads and determines their closest matching protein sequences in a set of reference sequences. We validated this new tool with sequences from defined communities and demonstrated the tool’s utility on nifH gene fragments sequenced from soils in well-characterized and major terrestrial ecosystem types.Qiong WangJohn F. QuensenJordan A. FishTae Kwon LeeYanni SunJames M. TiedjeJames R. ColeAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 4, Iss 5 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Qiong Wang John F. Quensen Jordan A. Fish Tae Kwon Lee Yanni Sun James M. Tiedje James R. Cole Ecological Patterns of <italic toggle="yes">nifH</italic> Genes in Four Terrestrial Climatic Zones Explored with Targeted Metagenomics Using FrameBot, a New Informatics Tool
description	ABSTRACT Biological nitrogen fixation is an important component of sustainable soil fertility and a key component of the nitrogen cycle. We used targeted metagenomics to study the nitrogen fixation-capable terrestrial bacterial community by targeting the gene for nitrogenase reductase (nifH). We obtained 1.1 million nifH 454 amplicon sequences from 222 soil samples collected from 4 National Ecological Observatory Network (NEON) sites in Alaska, Hawaii, Utah, and Florida. To accurately detect and correct frameshifts caused by indel sequencing errors, we developed FrameBot, a tool for frameshift correction and nearest-neighbor classification, and compared its accuracy to that of two other rapid frameshift correction tools. We found FrameBot was, in general, more accurate as long as a reference protein sequence with 80% or greater identity to a query was available, as was the case for virtually all nifH reads for the 4 NEON sites. Frameshifts were present in 12.7% of the reads. Those nifH sequences related to the Proteobacteria phylum were most abundant, followed by those for Cyanobacteria in the Alaska and Utah sites. Predominant genera with nifH sequences similar to reads included Azospirillum, Bradyrhizobium, and Rhizobium, the latter two without obvious plant hosts at the sites. Surprisingly, 80% of the sequences had greater than 95% amino acid identity to known nifH gene sequences. These samples were grouped by site and correlated with soil environmental factors, especially drainage, light intensity, mean annual temperature, and mean annual precipitation. FrameBot was tested successfully on three ecofunctional genes but should be applicable to any. IMPORTANCE High-throughput phylogenetic analysis of microbial communities using rRNA-targeted sequencing is now commonplace; however, such data often allow little inference with respect to either the presence or the diversity of genes involved in most important ecological processes. To study the gene pool for these processes, it is more straightforward to assess the genes directly responsible for the ecological function (ecofunctional genes). However, analyzing these genes involves technical challenges beyond those seen for rRNA. In particular, frameshift errors cause garbled downstream protein translations. Our FrameBot tool described here both corrects frameshift errors in query reads and determines their closest matching protein sequences in a set of reference sequences. We validated this new tool with sequences from defined communities and demonstrated the tool’s utility on nifH gene fragments sequenced from soils in well-characterized and major terrestrial ecosystem types.
format	article
author	Qiong Wang John F. Quensen Jordan A. Fish Tae Kwon Lee Yanni Sun James M. Tiedje James R. Cole
author_facet	Qiong Wang John F. Quensen Jordan A. Fish Tae Kwon Lee Yanni Sun James M. Tiedje James R. Cole
author_sort	Qiong Wang
title	Ecological Patterns of <italic toggle="yes">nifH</italic> Genes in Four Terrestrial Climatic Zones Explored with Targeted Metagenomics Using FrameBot, a New Informatics Tool
title_short	Ecological Patterns of <italic toggle="yes">nifH</italic> Genes in Four Terrestrial Climatic Zones Explored with Targeted Metagenomics Using FrameBot, a New Informatics Tool
title_full	Ecological Patterns of <italic toggle="yes">nifH</italic> Genes in Four Terrestrial Climatic Zones Explored with Targeted Metagenomics Using FrameBot, a New Informatics Tool
title_fullStr	Ecological Patterns of <italic toggle="yes">nifH</italic> Genes in Four Terrestrial Climatic Zones Explored with Targeted Metagenomics Using FrameBot, a New Informatics Tool
title_full_unstemmed	Ecological Patterns of <italic toggle="yes">nifH</italic> Genes in Four Terrestrial Climatic Zones Explored with Targeted Metagenomics Using FrameBot, a New Informatics Tool
title_sort	ecological patterns of <italic toggle="yes">nifh</italic> genes in four terrestrial climatic zones explored with targeted metagenomics using framebot, a new informatics tool
publisher	American Society for Microbiology
publishDate	2013
url	https://doaj.org/article/6eb8ef141cfc40b3a4e8da84ddd638b1
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Ecological Patterns of <italic toggle="yes">nifH</italic> Genes in Four Terrestrial Climatic Zones Explored with Targeted Metagenomics Using FrameBot, a New Informatics Tool

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