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Reliability of stool microbiome methods for DNA yields and sequencing among infants and young children
With the emergence of large‐scale epidemiologic human microbiome studies, there is a need to understand the reproducibility of microbial DNA sequencing and the impact of specimen collection and processing methods on measures of microbial community composition and structure, with reproducibility stud...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
John Wiley and Sons Inc.
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7221451/ https://www.ncbi.nlm.nih.gov/pubmed/32166902 http://dx.doi.org/10.1002/mbo3.1018 |
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author | Antosca, Katherine Hoen, Anne G. Palys, Thomas Hilliard, Margaret Morrison, Hilary G. Coker, Modupe Madan, Juliette Karagas, Margaret R. |
author_facet | Antosca, Katherine Hoen, Anne G. Palys, Thomas Hilliard, Margaret Morrison, Hilary G. Coker, Modupe Madan, Juliette Karagas, Margaret R. |
author_sort | Antosca, Katherine |
collection | PubMed |
description | With the emergence of large‐scale epidemiologic human microbiome studies, there is a need to understand the reproducibility of microbial DNA sequencing and the impact of specimen collection and processing methods on measures of microbial community composition and structure, with reproducibility studies in infants and young children particularly lacking. Here, we examined batch‐to‐batch variability and reliability of collection, handling, and processing protocols, testing replicate stool samples from infants and young children using Illumina MiSeq sequencing of the bacterial 16S rRNA gene V4‐V5 hypervariable region, evaluating 33 conditions with different protocols and extraction methods. We detected no evidence of batch effects in replicate DNA samples or extractions from the same stool sample. Variability in DNA yield and alpha diversity was observed between the different collection, handling, and processing protocols. However, across all protocols, subject variability was the dominant contributor to microbiome structure, with comparatively little impact of the protocol used. While collection method and DNA extraction kit may affect DNA yield, and correspondingly alpha diversity, our findings suggest that characterization of the structure and composition of the fecal microbiome of infants and young children are reliably measurable by standardized collection, handling, and processing protocols and DNA extraction methods within an individual longitudinal study. |
format | Online Article Text |
id | pubmed-7221451 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | John Wiley and Sons Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-72214512020-05-15 Reliability of stool microbiome methods for DNA yields and sequencing among infants and young children Antosca, Katherine Hoen, Anne G. Palys, Thomas Hilliard, Margaret Morrison, Hilary G. Coker, Modupe Madan, Juliette Karagas, Margaret R. Microbiologyopen Original Articles With the emergence of large‐scale epidemiologic human microbiome studies, there is a need to understand the reproducibility of microbial DNA sequencing and the impact of specimen collection and processing methods on measures of microbial community composition and structure, with reproducibility studies in infants and young children particularly lacking. Here, we examined batch‐to‐batch variability and reliability of collection, handling, and processing protocols, testing replicate stool samples from infants and young children using Illumina MiSeq sequencing of the bacterial 16S rRNA gene V4‐V5 hypervariable region, evaluating 33 conditions with different protocols and extraction methods. We detected no evidence of batch effects in replicate DNA samples or extractions from the same stool sample. Variability in DNA yield and alpha diversity was observed between the different collection, handling, and processing protocols. However, across all protocols, subject variability was the dominant contributor to microbiome structure, with comparatively little impact of the protocol used. While collection method and DNA extraction kit may affect DNA yield, and correspondingly alpha diversity, our findings suggest that characterization of the structure and composition of the fecal microbiome of infants and young children are reliably measurable by standardized collection, handling, and processing protocols and DNA extraction methods within an individual longitudinal study. John Wiley and Sons Inc. 2020-03-12 /pmc/articles/PMC7221451/ /pubmed/32166902 http://dx.doi.org/10.1002/mbo3.1018 Text en © 2020 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd This is an open access article under the terms of the http://creativecommons.org/licenses/by-nc-nd/4.0/ License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non‐commercial and no modifications or adaptations are made. |
spellingShingle | Original Articles Antosca, Katherine Hoen, Anne G. Palys, Thomas Hilliard, Margaret Morrison, Hilary G. Coker, Modupe Madan, Juliette Karagas, Margaret R. Reliability of stool microbiome methods for DNA yields and sequencing among infants and young children |
title | Reliability of stool microbiome methods for DNA yields and sequencing among infants and young children |
title_full | Reliability of stool microbiome methods for DNA yields and sequencing among infants and young children |
title_fullStr | Reliability of stool microbiome methods for DNA yields and sequencing among infants and young children |
title_full_unstemmed | Reliability of stool microbiome methods for DNA yields and sequencing among infants and young children |
title_short | Reliability of stool microbiome methods for DNA yields and sequencing among infants and young children |
title_sort | reliability of stool microbiome methods for dna yields and sequencing among infants and young children |
topic | Original Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7221451/ https://www.ncbi.nlm.nih.gov/pubmed/32166902 http://dx.doi.org/10.1002/mbo3.1018 |
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