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Consistent, comprehensive and computationally efficient OTU definitions

, , , , , , , , , , , , , , ,
1 Center for Microbial Genetics and Genomics, Northern Arizona University, Flagstaff, AZ, USA
2 Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA
3 School of Public Health and Tropical Medicine, Southern Medical University, Guangzhou, Guangdong, China
4 Department of Computer Science, University of Colorado Boulder, Boulder, CO, USA
5 Department of Molecular, Cellular, and Developmental Biology, University of Colorado at Boulder, Boulder, CO, USA
6 Department of Chemistry and Biochemistry, University of Colorado Boulder, Boulder, CO, USA
7 Graduate Program in Biophysical Sciences, University of Chicago, Chicago, IL, U.S.A.
8 Institute for Genomics and Systems Biology, Argonne National Laboratory, Lemont, IL, U.S.A.
9 Department of Biological Sciences, Northern Arizona University, Flagstaff, AZ, United States
10 BioFrontiers Institute, University of Colorado at Boulder, Boulder, CO, USA
11 Department of Ecology and Evolution, University of Chicago, Chicago, Illinois, USA
12 Department of Pathology and Laboratory Medicine, Brown University, Providence, RI, USA
13 Howard Hughes Medical Institute, Boulder, CO, USA
DOI
10.7287/peerj.preprints.411v2
Published
Accepted
Subject Areas
Bioinformatics, Ecology, Microbiology
Keywords
OTU picking, microbial ecology, microbiome, qiime, bioinformatics
Copyright
© 2014 Rideout et al.
Licence
This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ PrePrints) and either DOI or URL of the article must be cited.
Cite this article
Rideout JR, He Y, Navas-Molina JA, Walters WA, Ursell LK, Gibbons SM, Chase JH, McDonald D, Gonzalez A, Robbins-Pianka A, Clemente JC, Gilbert J, Huse SM, Zhou H, Knight R, Caporaso JG. 2014. Consistent, comprehensive and computationally efficient OTU definitions. PeerJ PrePrints 2:e411v2

Abstract

We present a performance-optimized algorithm, subsampled open-reference OTU picking, for assigning marker gene (e.g., 16S rRNA) sequences generated on next-generation sequencing platforms to operational taxonomic units (OTUs) for microbial community analysis. This algorithm provides benefits over de novo OTU picking (clustering can be performed largely in parallel, reducing runtime) and closed-reference OTU picking (all reads are clustered, not only those that match a reference database sequence with high similarity). Because more of our algorithm can be run in parallel relative to “classic” open-reference OTU picking, it makes open-reference OTU picking tractable on massive amplicon sequence data sets (though on smaller data sets, “classic” open-reference OTU clustering is often faster). We illustrate that here by applying it to the first 15,000 samples sequenced for the Earth Microbiome Project (1.3 billion V4 16S rRNA amplicons). To the best of our knowledge, this is the largest OTU picking run ever performed, and we estimate that our new algorithm runs in less than 1/5 the time than would be required of “classic” open reference OTU picking. We show that subsampled open-reference OTU picking yields results that are highly correlated with those generated by “classic” open-reference OTU picking through comparisons on three well-studied datasets. An implementation of this algorithm is provided in the popular QIIME software package, which uses uclust for read clustering. All analyses were performed using QIIME’s uclust wrappers, though we provide details (aided by the open-source code in our GitHub repository) that will allow implementation of subsampled open-reference OTU picking independently of QIIME (e.g., in a compiled programming language, where runtimes should be further reduced). Our analyses should generalize to other implementations of these OTU picking algorithms. Finally, we present a comparison of parameter settings in QIIME’s OTU picking workflows and make recommendations on settings for these free parameters to optimize runtime without reducing the quality of the results. These optimized parameters can vastly decrease the runtime of uclust-based OTU picking in QIIME.

Author Comment

This includes changes in response to reviewer comments.

Supplemental Information

Supplementary Data

Description of where to obtain data for this study.

DOI: 10.7287/peerj.preprints.411v2/supp-1

Additional Information

Competing Interests

There are no competing interests.

Author Contributions

Jai Ram Rideout performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, prepared figures and/or tables, reviewed drafts of the paper.

Yan He performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, wrote the paper, prepared figures and/or tables, reviewed drafts of the paper.

Jose Antonio Navas-Molina performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, wrote the paper, reviewed drafts of the paper.

William A Walters performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, reviewed drafts of the paper.

Luke K Ursell performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, wrote the paper, reviewed drafts of the paper.

Sean M Gibbons performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, reviewed drafts of the paper.

John H Chase performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, reviewed drafts of the paper.

Daniel McDonald performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, reviewed drafts of the paper.

Antonio Gonzalez performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, reviewed drafts of the paper.

Adam Robbins-Pianka performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, reviewed drafts of the paper.

Jose C Clemente conceived and designed the experiments, contributed reagents/materials/analysis tools, prepared figures and/or tables, reviewed drafts of the paper.

Jack Gilbert conceived and designed the experiments, contributed reagents/materials/analysis tools, reviewed drafts of the paper.

Susan M Huse conceived and designed the experiments, contributed reagents/materials/analysis tools, reviewed drafts of the paper.

Hong-Wei Zhou conceived and designed the experiments, contributed reagents/materials/analysis tools, reviewed drafts of the paper.

Rob Knight conceived and designed the experiments, contributed reagents/materials/analysis tools, wrote the paper, reviewed drafts of the paper.

J Gregory Caporaso conceived and designed the experiments, performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, wrote the paper, prepared figures and/or tables, reviewed drafts of the paper.

Human Ethics

The following information was supplied relating to ethical approvals (i.e., approving body and any reference numbers):

All of the human subjects data used in this study was previously published and is publicly accessible. The citations for each dataset contains the IRB protocol information.

Data Deposition

The following information was supplied regarding the deposition of related data:

https://github.com/gregcaporaso/cloaked-octo-ninja

Funding

SMG was supported by an EPA STAR Graduate Fellowship. DM was supported in part by NSF IGERT (award number: 1144807). This work was partially supported by a grant from Arizona's Technology and Research Initiative Fund to JGC (no award number), and by a grant from the Alfred P. Sloan Foundation to JGC and RK (award number: 2012-5-42 MBRP). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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Consistent, comprehensive and computationally efficient OTU definitions
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