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Towards standards for human fecal sample processing in metagenomic studies

by Pons, Nicolas , Patil, Kiran Raosaheb , Singh, Bhagirath , Coelho, Luis Pedro , Hercog, Rajna , Hayward, Matthew R , Daigneault, Michelle , Popova, Milena , Hattori, Masahira , de Vos, Willem M , Flint, Harry J , Druesne, Anne , Langella, Philippe , Zeller, Georg , Le Chatelier, Emmanuelle , Heilig, Hans , Bertrand, Laurie , Derrien, Muriel , Dore, Joel , Pelletier, Eric , Klymiuk, Ingeborg , Alberti, Adriana , Tramontano, Melanie , Guarner, Francisco , Slezak, Kathleen , Luna, Ruth Ann , Orvain, Céline , Martin, Jennifer C , Persson, Søren , Blaut, Michael , van Hylckama Vlieg, Johan , Driessen, Marja , O'Toole, Paul W , Junick, Jana , Bork, Peer , Jung, Ferris-Elias , Kultima, Jens Roat , Allen-Vercoe, Emma , Versalovic, James , Mai, Volker , Scott, Karen P , Carton, Thomas , Mery, Clémentine , Brown, Jillian R M , Zoetendal, Erwin G , Levenez, Florence , Cools-Portier, Stéphanie , Finlay, B Brett , Saulnier, Delphine , Costea, Paul I , Penders, John , Morita, Hidetoshi , Ehrlich, S Dusko , Manichanh, Chaysavanh , Veiga, Patrick , Zhao, Liping , Sunagawa, Shinichi , Salonen, Anne

in 13/62 / 45/23 / 631/326/2522 / 631/326/2565/2142 / Agriculture / analysis / Bacteria / Bacteria - genetics / Bioinformatics / Biomedical Engineering/Biotechnology / Biomedicine / Biotechnology / Chemical Fractionation - methods / Communities / Community composition / Computational Biology / Deoxyribonucleic acid / DNA / DNA - chemistry / Feces / Feces - chemistry / Gram-negative bacteria / Gram-positive bacteria / Humans / Intestinal microflora / Laboratorium voor Microbiologie / Life Sciences / Metagenomics / Microbiological Laboratory / Microbiologie / Microbiology / Microbiomes / Microbiota / Microorganisms / Quality Control / Species Specificity / Stool specimen analysis / VLAG / WIMEK

2017

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2017

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2017

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Journal Article

Towards standards for human fecal sample processing in metagenomic studies

Pons, Nicolas,

Patil, Kiran Raosaheb,

Singh, Bhagirath,

Coelho, Luis Pedro,

Hercog, Rajna,

Hayward, Matthew R,

Daigneault, Michelle,

Popova, Milena,

Hattori, Masahira,

de Vos, Willem M,

Flint, Harry J,

Druesne, Anne,

Langella, Philippe,

Zeller, Georg,

Le Chatelier, Emmanuelle,

Heilig, Hans,

Bertrand, Laurie,

Derrien, Muriel,

Dore, Joel,

Pelletier, Eric,

Klymiuk, Ingeborg,

Alberti, Adriana,

Tramontano, Melanie,

Guarner, Francisco,

Slezak, Kathleen,

Luna, Ruth Ann,

Orvain, Céline,

Martin, Jennifer C,

Persson, Søren,

Blaut, Michael,

van Hylckama Vlieg, Johan,

Driessen, Marja,

O'Toole, Paul W,

Junick, Jana,

Bork, Peer,

Jung, Ferris-Elias,

Kultima, Jens Roat,

Allen-Vercoe, Emma,

Versalovic, James,

Mai, Volker,

Scott, Karen P,

Carton, Thomas,

Mery, Clémentine,

Brown, Jillian R M,

Zoetendal, Erwin G,

Levenez, Florence,

Cools-Portier, Stéphanie,

Finlay, B Brett,

Saulnier, Delphine,

Costea, Paul I,

Penders, John,

Morita, Hidetoshi,

Ehrlich, S Dusko,

Manichanh, Chaysavanh,

Veiga, Patrick,

Zhao, Liping,

Sunagawa, Shinichi,

Salonen, Anne

2017

Overview

Testing 21 different fecal DNA extraction protocols in multiple laboratories results in a standardized protocol with the potential to improve comparability across human gut microbiome studies. Technical variation in metagenomic analysis must be minimized to confidently assess the contributions of microbiota to human health. Here we tested 21 representative DNA extraction protocols on the same fecal samples and quantified differences in observed microbial community composition. We compared them with differences due to library preparation and sample storage, which we contrasted with observed biological variation within the same specimen or within an individual over time. We found that DNA extraction had the largest effect on the outcome of metagenomic analysis. To rank DNA extraction protocols, we considered resulting DNA quantity and quality, and we ascertained biases in estimates of community diversity and the ratio between Gram-positive and Gram-negative bacteria. We recommend a standardized DNA extraction method for human fecal samples, for which transferability across labs was established and which was further benchmarked using a mock community of known composition. Its adoption will improve comparability of human gut microbiome studies and facilitate meta-analyses.

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Publisher

Nature Publishing Group US,Nature Publishing Group

Subject

/ 45/23