Towards standards for human fecal sample processing in metagenomic studies

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

*Corresponding author for this work

    Research output: Contribution to journalArticlepeer-review

    396 Citations (Scopus)


    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.

    Original languageEnglish
    Pages (from-to)1069-1076
    Number of pages8
    JournalNature Biotechnology
    Issue number11
    Publication statusPublished - 2017 Nov 1

    ASJC Scopus subject areas

    • Biotechnology
    • Bioengineering
    • Applied Microbiology and Biotechnology
    • Molecular Medicine
    • Biomedical Engineering


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