Phylogenetic placement of exact amplicon sequences improves associations with clinical information

Stefan Janssen, Daniel McDonald, Antonio Gonzalez, Jose A. Navas-Molina, Lingjing Jiang, Zhenjiang Zech Xu, Kevin Winker, Deborah M. Kado, Eric Orwoll, Mark Manary, Siavash Mirarab, Rob Knight

Research output: Contribution to journalArticlepeer-review

300 Scopus citations


Recent algorithmic advances in amplicon-based microbiome studies enable the inference of exact amplicon sequence fragments. These new methods enable the investigation of sub-operational taxonomic units (sOTU) by removing erroneous sequences. However, short (e.g., 150-nucleotide [nt]) DNA sequence fragments do not contain sufficient phylogenetic signal to reproduce a reasonable tree, introducing a barrier in the utilization of critical phylogenetically aware metrics such as Faith's PD or UniFrac. Although fragment insertion methods do exist, those methods have not been tested for sOTUs from high-throughput amplicon studies in insertions against a broad reference phylogeny. We benchmarked the SATé-enabled phylogenetic placement (SEPP) technique explicitly against 16S V4 sequence fragments and showed that it outperforms the conceptually problematic but often-used practice of reconstructing de novo phylogenies. In addition, we provide a BSD-licensed QIIME2 plugin ( for SEPP and integration into the microbial study management platform QIITA. IMPORTANCE The move from OTU-based to sOTU-based analysis, while providing additional resolution, also introduces computational challenges. We demonstrate that one popular method of dealing with sOTUs (building a de novo tree from the short sequences) can provide incorrect results in human gut metagenomic studies and show that phylogenetic placement of the new sequences with SEPP resolves this problem while also yielding other benefits over existing methods.

Original languageEnglish (US)
Article numbere00021-18
Issue number3
StatePublished - May 1 2018


  • Amplicon sequencing
  • Microbial community analysis
  • Phylogenetic placement
  • SEPP

ASJC Scopus subject areas

  • Microbiology
  • Physiology
  • Biochemistry
  • Ecology, Evolution, Behavior and Systematics
  • Modeling and Simulation
  • Molecular Biology
  • Genetics
  • Computer Science Applications


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