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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doaj-5cae92b7109240d78171c196d697b3372020-11-25T01:43:12ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2018-09-01910.3389/fmicb.2018.02146356564A Comparative Analysis of Microbial DNA Preparation Methods for Use With Massive and Branching Coral Growth FormsAlejandra Hernandez-Agreda0Alejandra Hernandez-Agreda1William Leggat2William Leggat3William Leggat4Tracy D. Ainsworth5Tracy D. Ainsworth6Australian Research Council Centre of Excellence for Coral Reef Studies, James Cook University, Townsville, QLD, AustraliaCollege of Public Health, Medical and Veterinary Sciences, James Cook University, Townsville, QLD, AustraliaAustralian Research Council Centre of Excellence for Coral Reef Studies, James Cook University, Townsville, QLD, AustraliaCollege of Public Health, Medical and Veterinary Sciences, James Cook University, Townsville, QLD, AustraliaSchool of Environmental and Life Sciences, The University of Newcastle, Ourimbah, NSW, AustraliaAustralian Research Council Centre of Excellence for Coral Reef Studies, James Cook University, Townsville, QLD, AustraliaSchool of Biological, Earth and Environmental Sciences, University of New South Wales, Sydney, NSW, AustraliaIn 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.https://www.frontiersin.org/article/10.3389/fmicb.2018.02146/fullcoral microbiomebacteriamicrobial ecologyDESSparaformaldehydesnap frozen |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Alejandra Hernandez-Agreda Alejandra Hernandez-Agreda William Leggat William Leggat William Leggat Tracy D. Ainsworth Tracy D. Ainsworth |
spellingShingle |
Alejandra Hernandez-Agreda Alejandra Hernandez-Agreda William Leggat William Leggat William Leggat Tracy D. Ainsworth Tracy D. Ainsworth A Comparative Analysis of Microbial DNA Preparation Methods for Use With Massive and Branching Coral Growth Forms Frontiers in Microbiology coral microbiome bacteria microbial ecology DESS paraformaldehyde snap frozen |
author_facet |
Alejandra Hernandez-Agreda Alejandra Hernandez-Agreda William Leggat William Leggat William Leggat Tracy D. Ainsworth Tracy D. Ainsworth |
author_sort |
Alejandra Hernandez-Agreda |
title |
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_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_sort |
comparative analysis of microbial dna preparation methods for use with massive and branching coral growth forms |
publisher |
Frontiers Media S.A. |
series |
Frontiers in Microbiology |
issn |
1664-302X |
publishDate |
2018-09-01 |
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. |
topic |
coral microbiome bacteria microbial ecology DESS paraformaldehyde snap frozen |
url |
https://www.frontiersin.org/article/10.3389/fmicb.2018.02146/full |
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