Establishing microbial composition measurement standards with reference frames

James T. Morton, Clarisse Marotz, Alex Washburne, Justin Silverman, Livia S. Zaramela, Anna Edlund, Karsten Zengler, Rob Knight

    Research output: Contribution to journalArticlepeer-review

    224 Scopus citations


    Differential abundance analysis is controversial throughout microbiome research. Gold standard approaches require laborious measurements of total microbial load, or absolute number of microorganisms, to accurately determine taxonomic shifts. Therefore, most studies rely on relative abundance data. Here, we demonstrate common pitfalls in comparing relative abundance across samples and identify two solutions that reveal microbial changes without the need to estimate total microbial load. We define the notion of “reference frames”, which provide deep intuition about the compositional nature of microbiome data. In an oral time series experiment, reference frames alleviate false positives and produce consistent results on both raw and cell-count normalized data. Furthermore, reference frames identify consistent, differentially abundant microbes previously undetected in two independent published datasets from subjects with atopic dermatitis. These methods allow reassessment of published relative abundance data to reveal reproducible microbial changes from standard sequencing output without the need for new assays.

    Original languageEnglish (US)
    Article number2719
    JournalNature communications
    Issue number1
    StatePublished - Dec 1 2019

    All Science Journal Classification (ASJC) codes

    • Chemistry(all)
    • Biochemistry, Genetics and Molecular Biology(all)
    • Physics and Astronomy(all)


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