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A Comparative Analysis of Microbial DNA Preparation Methods for Use With Massive and Branching Coral Growth Forms

In the last two decades, over 100 studies have investigated the structure of the coral microbiome. However, as yet there are no standardized methods applied to sample preservation and preparation, with different studies using distinct methods. There have also been several comparisons made of microbi...

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Autores principales: Hernandez-Agreda, Alejandra, Leggat, William, Ainsworth, Tracy D.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6137167/
https://www.ncbi.nlm.nih.gov/pubmed/30245683
http://dx.doi.org/10.3389/fmicb.2018.02146
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author Hernandez-Agreda, Alejandra
Leggat, William
Ainsworth, Tracy D.
author_facet Hernandez-Agreda, Alejandra
Leggat, William
Ainsworth, Tracy D.
author_sort Hernandez-Agreda, Alejandra
collection PubMed
description In the last two decades, over 100 studies have investigated the structure of the coral microbiome. However, as yet there are no standardized methods applied to sample preservation and preparation, with different studies using distinct methods. There have also been several comparisons made of microbiome data generated across different studies, which have not addressed the influence of the methodology employed over each of the microbiome datasets. Here, we assess three different preservation methods; salt saturated dimethyl sulfoxide (DMSO) – EDTA, snap freezing with liquid nitrogen and 4% paraformaldehyde solution, and two different preparation methodologies; bead beating and crushing, that have been applied to study the coral microbiome. We compare the resultant bacterial assemblage data for two coral growth forms, the massive coral Goniastrea edwardsi and the branching coral Isopora palifera. We show that microbiome datasets generated from differing preservation and processing protocols are comparable in composition (presence/absence). Significant discrepancies between preservation and homogenization methods are observed in structure (relative abundance), and in the occurrence and dominance of taxa, with rare (low abundance and low occurrence) phylotypes being the most variable fraction of the microbial community. Finally, we provide evidence to support chemical preservation with DMSO as effective as snap freezing samples for generating reliable and robust microbiome datasets. In conclusion, we recommend where possible a standardized preservation and extraction method be taken up by the field to provide the best possible practices for detailed assessments of symbiotic and conserved bacterial associations.
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spelling pubmed-61371672018-09-21 A Comparative Analysis of Microbial DNA Preparation Methods for Use With Massive and Branching Coral Growth Forms Hernandez-Agreda, Alejandra Leggat, William Ainsworth, Tracy D. Front Microbiol Microbiology In the last two decades, over 100 studies have investigated the structure of the coral microbiome. However, as yet there are no standardized methods applied to sample preservation and preparation, with different studies using distinct methods. There have also been several comparisons made of microbiome data generated across different studies, which have not addressed the influence of the methodology employed over each of the microbiome datasets. Here, we assess three different preservation methods; salt saturated dimethyl sulfoxide (DMSO) – EDTA, snap freezing with liquid nitrogen and 4% paraformaldehyde solution, and two different preparation methodologies; bead beating and crushing, that have been applied to study the coral microbiome. We compare the resultant bacterial assemblage data for two coral growth forms, the massive coral Goniastrea edwardsi and the branching coral Isopora palifera. We show that microbiome datasets generated from differing preservation and processing protocols are comparable in composition (presence/absence). Significant discrepancies between preservation and homogenization methods are observed in structure (relative abundance), and in the occurrence and dominance of taxa, with rare (low abundance and low occurrence) phylotypes being the most variable fraction of the microbial community. Finally, we provide evidence to support chemical preservation with DMSO as effective as snap freezing samples for generating reliable and robust microbiome datasets. In conclusion, we recommend where possible a standardized preservation and extraction method be taken up by the field to provide the best possible practices for detailed assessments of symbiotic and conserved bacterial associations. Frontiers Media S.A. 2018-09-07 /pmc/articles/PMC6137167/ /pubmed/30245683 http://dx.doi.org/10.3389/fmicb.2018.02146 Text en Copyright © 2018 Hernandez-Agreda, Leggat and Ainsworth. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Microbiology
Hernandez-Agreda, Alejandra
Leggat, William
Ainsworth, Tracy D.
A Comparative Analysis of Microbial DNA Preparation Methods for Use With Massive and Branching Coral Growth Forms
title A Comparative Analysis of Microbial DNA Preparation Methods for Use With Massive and Branching Coral Growth Forms
title_full A Comparative Analysis of Microbial DNA Preparation Methods for Use With Massive and Branching Coral Growth Forms
title_fullStr A Comparative Analysis of Microbial DNA Preparation Methods for Use With Massive and Branching Coral Growth Forms
title_full_unstemmed A Comparative Analysis of Microbial DNA Preparation Methods for Use With Massive and Branching Coral Growth Forms
title_short A Comparative Analysis of Microbial DNA Preparation Methods for Use With Massive and Branching Coral Growth Forms
title_sort comparative analysis of microbial dna preparation methods for use with massive and branching coral growth forms
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6137167/
https://www.ncbi.nlm.nih.gov/pubmed/30245683
http://dx.doi.org/10.3389/fmicb.2018.02146
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