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 Daigneault; Muriel 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

doi:10.1038/nbt.3960

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

Research output: Contribution to journal › Article

136 Citations (Scopus)

Abstract

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 language	English
Pages (from-to)	1069–1076
Number of pages	8
Journal	Nature Biotechnology
Volume	35
Early online date	2 Oct 2017
DOIs	https://doi.org/10.1038/nbt.3960
Publication status	Published - 2017

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Keywords

applied microbiology
metagenomics

Cite this

Costea, P. I., Zeller, G., Sunagawa, S., Pelletier, E., Alberti, A., Levenez, F., Tramontano, M., Driessen, M., Hercog, R., Jung, F-E., Kultima, J. R., Hayward, M. R., Coelho, L. P., Allen-Vercoe, E., Bertrand, L., Blaut, M., Brown, J. R. M., Carton, T., Cools-Portier, S., ... Bork, P. (2017). Towards standards for human fecal sample processing in metagenomic studies. Nature Biotechnology, 35, 1069–1076. https://doi.org/10.1038/nbt.3960

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

In: Nature Biotechnology, Vol. 35, 2017, p. 1069–1076.

Research output: Contribution to journal › Article

Costea, PI, Zeller, G, Sunagawa, S, Pelletier, E, Alberti, A, Levenez, F, Tramontano, M, Driessen, M, Hercog, R, Jung, F-E, Kultima, JR, Hayward, MR, Coelho, LP, Allen-Vercoe, E, Bertrand, L, Blaut, M, Brown, JRM, Carton, T, Cools-Portier, S, Daigneault, M, Derrien, M, Druesne, A, de Vos, WM, Finlay, BB, Flint, HJ, Guarner, F, Hattori, M, Heilig, H, Luna, RA, van Hylckama Vlieg, J, Junick, J, Klymiuk, I, Langella, P, Le Chatelier, E, Mai, V, Manichanh, C, Martin, JC, Mery, C, Morita, H, O'Toole, PW, Orvain, C, Patil, KR, Penders, J, Persson, S, Pons, N, Popova, M, Salonen, A, Saulnier, D, Scott, KP, Singh, B, Slezak, K, Veiga, P, Versalovic, J, Zhao, L, Zoetendal, EG, Ehrlich, SD, Dore, J & Bork, P 2017, 'Towards standards for human fecal sample processing in metagenomic studies', Nature Biotechnology, vol. 35, pp. 1069–1076. https://doi.org/10.1038/nbt.3960

Costea, Paul I ; Zeller, Georg ; Sunagawa, Shinichi ; Pelletier, Eric ; Alberti, Adriana ; Levenez, Florence ; Tramontano, Melanie ; Driessen, Marja ; Hercog, Rajna ; Jung, Ferris-Elias ; Kultima, Jens Roat ; Hayward, Matthew R ; Coelho, Luis Pedro ; Allen-Vercoe, Emma ; Bertrand, Laurie ; Blaut, Michael ; Brown, Jillian R M ; Carton, Thomas ; Cools-Portier, Stéphanie ; Daigneault, Michelle ; Derrien, Muriel ; Druesne, Anne ; de Vos, Willem M ; Finlay, B Brett ; Flint, Harry J ; Guarner, Francisco ; Hattori, Masahira ; Heilig, Hans ; Luna, Ruth Ann ; van Hylckama Vlieg, Johan ; Junick, Jana ; Klymiuk, Ingeborg ; Langella, Philippe ; Le Chatelier, Emmanuelle ; Mai, Volker ; Manichanh, Chaysavanh ; Martin, Jennifer C ; Mery, Clémentine ; Morita, Hidetoshi ; O'Toole, Paul W ; Orvain, Céline ; Patil, Kiran Raosaheb ; Penders, John ; Persson, Søren ; Pons, Nicolas ; Popova, Milena ; Salonen, Anne ; Saulnier, Delphine ; Scott, Karen P ; Singh, Bhagirath ; Slezak, Kathleen ; Veiga, Patrick ; Versalovic, James ; Zhao, Liping ; Zoetendal, Erwin G ; Ehrlich, S Dusko ; Dore, Joel ; Bork, Peer. / Towards standards for human fecal sample processing in metagenomic studies. In: Nature Biotechnology. 2017 ; Vol. 35. pp. 1069–1076.

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

note = "We thank S. Burz and K. Weizer for editing and web-posting the SOPs. We thank D. Ordonez and N.P. Gabrielli Lopez for advice on flow cytometry, which was provided by the Flow Cytometry Core Facility, EMBL. This study was funded by the European Community's Seventh Framework Programme via International Human Microbiome Standards (HEALTH-F4-2010-261376) grant. We also received support from Scottish Government Rural and Environmental Science and Analytical Services as well as from EMBL.",

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AU - Zeller, Georg

AU - Sunagawa, Shinichi

AU - Pelletier, Eric

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AU - Tramontano, Melanie

AU - Driessen, Marja

AU - Hercog, Rajna

AU - Jung, Ferris-Elias

AU - Kultima, Jens Roat

AU - Hayward, Matthew R

AU - Coelho, Luis Pedro

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AU - Manichanh, Chaysavanh

AU - Martin, Jennifer C

AU - Mery, Clémentine

AU - Morita, Hidetoshi

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AB - 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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