The effects of freezing on faecal microbiota as determined using MiSeq sequencing and culture-based investigations

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dc.contributor.authorFouhy, Fiona
dc.contributor.authorDeane, Jennifer
dc.contributor.authorRea, Mary C.
dc.contributor.authorO'Sullivan, Orla
dc.contributor.authorRoss, R. Paul
dc.contributor.authorO'Callaghan, Grace
dc.contributor.authorPlant, Barry J.
dc.contributor.authorStanton, Catherine
dc.contributor.funderTeagasc
dc.contributor.funderScience Foundation Ireland
dc.contributor.funderEuropean Commission
dc.date.accessioned2016-02-17T10:07:59Z
dc.date.available2016-02-17T10:07:59Z
dc.date.issued2015
dc.description.abstractBackground: High-throughput sequencing has enabled detailed insights into complex microbial environments, including the human gut microbiota. The accuracy of the sequencing data however, is reliant upon appropriate storage of the samples prior to DNA extraction. The aim of this study was to conduct the first MiSeq sequencing investigation into the effects of faecal storage on the microbiota, compared to fresh samples. Culture-based analysis was also completed. Methods: Seven faecal samples were collected from healthy adults. Samples were separated into fresh (DNA extracted immediately), snap frozen on dry ice and frozen for 7 days at -80 degrees C prior to DNA extraction or samples frozen at -80 degrees C for 7 days before DNA extraction. Sequencing was completed on the Illumina MiSeq platform. Culturing of total aerobes, anaerobes and bifidobacteria was also completed. Results: No significant differences at phylum or family levels between the treatment groups occurred. At genus level only Faecalibacterium and Leuconostoc were significantly different in the fresh samples compared to the snap frozen group (p = 0.0298; p = 0.0330 respectively). Diversity analysis indicated that samples clustered based on the individual donor, rather than by storage group. No significant differences occurred in the culture-based analysis between the fresh, snap or -80 degrees C frozen samples. Conclusions: Using the MiSeq platform coupled with culture-based analysis, this study highlighted that limited significant changes in microbiota occur following rapid freezing of faecal samples prior to DNA extraction. Thus, rapid freezing of samples prior to DNA extraction and culturing, preserves the integrity of the microbiota.en
dc.description.sponsorshipTeagasc (Walsh Fellowship); Science Foundation Ireland (SFI-CSET02/CE/B124; SFI/12/RC/2273); European Commission (FP7 Grant Agreement No. 603038);en
dc.description.statusPeer revieweden
dc.description.versionPublished Versionen
dc.format.mimetypeapplication/pdfen
dc.identifier.articleide0119355
dc.identifier.citationFouhy F, Deane J, Rea MC, O’Sullivan Ó, Ross RP, O’Callaghan G, et al. (2015) The Effects of Freezing on Faecal Microbiota as Determined Using MiSeq Sequencing and Culture-Based Investigations. PLoS ONE 10(3): e0119355. doi:10.1371/journal.pone.0119355
dc.identifier.doi10.1371/journal.pone.0119355
dc.identifier.issn1932-6203
dc.identifier.issued3en
dc.identifier.journaltitlePLOS ONEen
dc.identifier.urihttps://hdl.handle.net/10468/2315
dc.identifier.volume10en
dc.language.isoenen
dc.publisherPublic Library of Scienceen
dc.rights© 2015 Fouhy et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are crediteden
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/en
dc.subjectGut microbiotaen
dc.subjectStorage conditionsen
dc.subjectDna extractionen
dc.subjectBacterialen
dc.subjectSamplesen
dc.subjectCollectionen
dc.subjectDiversityen
dc.subjectDiseaseen
dc.subjectFloraen
dc.titleThe effects of freezing on faecal microbiota as determined using MiSeq sequencing and culture-based investigationsen
dc.typeArticle (peer-reviewed)en
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