Human and Extracellular DNA Depletion for Metagenomic Analysis of Complex Clinical Infection Samples Yields Optimized Viable Microbiome Profiles

Maria T. Nelson, Christopher E. Pope, Robyn L. Marsh, Daniel J. Wolter, Eli J. Weiss, Kyle R. Hager, Anh T. Vo, Mitchell J. Brittnacher, Matthew C. Radey, Hillary S. Hayden, Alexander Eng, Samuel I. Miller, Elhanan Borenstein, Lucas R. Hoffman

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    Abstract

    Metagenomic sequencing is a promising approach for identifying and characterizing organisms and their functional characteristics in complex, polymicrobial infections, such as airway infections in people with cystic fibrosis. These analyses are often hampered, however, by overwhelming quantities of human DNA, yielding only a small proportion of microbial reads for analysis. In addition, many abundant microbes in respiratory samples can produce large quantities of extracellular bacterial DNA originating either from biofilms or dead cells. We describe a method for simultaneously depleting DNA from intact human cells and extracellular DNA (human and bacterial) in sputum, using selective lysis of eukaryotic cells and endonuclease digestion. We show that this method increases microbial sequencing depth and, consequently, both the number of taxa detected and coverage of individual genes such as those involved in antibiotic resistance. This finding underscores the substantial impact of DNA from sources other than live bacteria in microbiological analyses of complex, chronic infection specimens.

    Original languageEnglish
    Pages (from-to)2227-2240.e1-e5
    Number of pages19
    JournalCell Reports
    Volume26
    Issue number8
    DOIs
    Publication statusPublished - 19 Feb 2019

    Fingerprint

    Metagenomics
    Microbiota
    Bacterial DNA
    DNA
    Infection
    Endonucleases
    Eukaryotic Cells
    Biofilms
    Microbial Drug Resistance
    Sputum
    Coinfection
    Cystic Fibrosis
    Digestion
    Bacteria
    Genes
    Cells
    Anti-Bacterial Agents

    Cite this

    Nelson, Maria T. ; Pope, Christopher E. ; Marsh, Robyn L. ; Wolter, Daniel J. ; Weiss, Eli J. ; Hager, Kyle R. ; Vo, Anh T. ; Brittnacher, Mitchell J. ; Radey, Matthew C. ; Hayden, Hillary S. ; Eng, Alexander ; Miller, Samuel I. ; Borenstein, Elhanan ; Hoffman, Lucas R. / Human and Extracellular DNA Depletion for Metagenomic Analysis of Complex Clinical Infection Samples Yields Optimized Viable Microbiome Profiles. In: Cell Reports. 2019 ; Vol. 26, No. 8. pp. 2227-2240.e1-e5.
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    abstract = "Metagenomic sequencing is a promising approach for identifying and characterizing organisms and their functional characteristics in complex, polymicrobial infections, such as airway infections in people with cystic fibrosis. These analyses are often hampered, however, by overwhelming quantities of human DNA, yielding only a small proportion of microbial reads for analysis. In addition, many abundant microbes in respiratory samples can produce large quantities of extracellular bacterial DNA originating either from biofilms or dead cells. We describe a method for simultaneously depleting DNA from intact human cells and extracellular DNA (human and bacterial) in sputum, using selective lysis of eukaryotic cells and endonuclease digestion. We show that this method increases microbial sequencing depth and, consequently, both the number of taxa detected and coverage of individual genes such as those involved in antibiotic resistance. This finding underscores the substantial impact of DNA from sources other than live bacteria in microbiological analyses of complex, chronic infection specimens.",
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    author = "Nelson, {Maria T.} and Pope, {Christopher E.} and Marsh, {Robyn L.} and Wolter, {Daniel J.} and Weiss, {Eli J.} and Hager, {Kyle R.} and Vo, {Anh T.} and Brittnacher, {Mitchell J.} and Radey, {Matthew C.} and Hayden, {Hillary S.} and Alexander Eng and Miller, {Samuel I.} and Elhanan Borenstein and Hoffman, {Lucas R.}",
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    Nelson, MT, Pope, CE, Marsh, RL, Wolter, DJ, Weiss, EJ, Hager, KR, Vo, AT, Brittnacher, MJ, Radey, MC, Hayden, HS, Eng, A, Miller, SI, Borenstein, E & Hoffman, LR 2019, 'Human and Extracellular DNA Depletion for Metagenomic Analysis of Complex Clinical Infection Samples Yields Optimized Viable Microbiome Profiles', Cell Reports, vol. 26, no. 8, pp. 2227-2240.e1-e5. https://doi.org/10.1016/j.celrep.2019.01.091

    Human and Extracellular DNA Depletion for Metagenomic Analysis of Complex Clinical Infection Samples Yields Optimized Viable Microbiome Profiles. / Nelson, Maria T.; Pope, Christopher E.; Marsh, Robyn L.; Wolter, Daniel J.; Weiss, Eli J.; Hager, Kyle R.; Vo, Anh T.; Brittnacher, Mitchell J.; Radey, Matthew C.; Hayden, Hillary S.; Eng, Alexander; Miller, Samuel I.; Borenstein, Elhanan; Hoffman, Lucas R.

    In: Cell Reports, Vol. 26, No. 8, 19.02.2019, p. 2227-2240.e1-e5.

    Research output: Contribution to journalArticleResearchpeer-review

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    AU - Vo, Anh T.

    AU - Brittnacher, Mitchell J.

    AU - Radey, Matthew C.

    AU - Hayden, Hillary S.

    AU - Eng, Alexander

    AU - Miller, Samuel I.

    AU - Borenstein, Elhanan

    AU - Hoffman, Lucas R.

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    AB - Metagenomic sequencing is a promising approach for identifying and characterizing organisms and their functional characteristics in complex, polymicrobial infections, such as airway infections in people with cystic fibrosis. These analyses are often hampered, however, by overwhelming quantities of human DNA, yielding only a small proportion of microbial reads for analysis. In addition, many abundant microbes in respiratory samples can produce large quantities of extracellular bacterial DNA originating either from biofilms or dead cells. We describe a method for simultaneously depleting DNA from intact human cells and extracellular DNA (human and bacterial) in sputum, using selective lysis of eukaryotic cells and endonuclease digestion. We show that this method increases microbial sequencing depth and, consequently, both the number of taxa detected and coverage of individual genes such as those involved in antibiotic resistance. This finding underscores the substantial impact of DNA from sources other than live bacteria in microbiological analyses of complex, chronic infection specimens.

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    KW - microbiome

    KW - microbiota

    KW - polymicrobial infections

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