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Next-generation sequencing in neuropathological diagnosis of infections of the nervous system

Steven L. Salzberg, Florian P. Breitwieser, Anupama Kumar, Haiping Hao, Peter Burger, Fausto J. Rodriguez, Michael Lim, Alfredo Quiñones-Hinojosa, Gary L. Gallia, Jeffrey A. Tornheim, Michael T. Melia, Cynthia L. Sears, Carlos A. Pardo
doi: https://doi.org/10.1101/039222
Steven L. Salzberg
1Center for Computational Biology, McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins School of Medicine
2Departments of Biomedical Engineering, Computer Science, and Biostatistics, Johns Hopkins University
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Florian P. Breitwieser
1Center for Computational Biology, McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins School of Medicine
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Anupama Kumar
3Department of Neurology, Johns Hopkins University School of Medicine
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Haiping Hao
4Deep Sequencing and Microarray Core, Johns Hopkins University School of Medicine
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Peter Burger
5Department of Pathology, Johns Hopkins University School of Medicine
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Fausto J. Rodriguez
5Department of Pathology, Johns Hopkins University School of Medicine
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Michael Lim
6Department of Neurosurgery, Johns Hopkins University School of Medicine
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Alfredo Quiñones-Hinojosa
6Department of Neurosurgery, Johns Hopkins University School of Medicine
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Gary L. Gallia
6Department of Neurosurgery, Johns Hopkins University School of Medicine
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Jeffrey A. Tornheim
7Department of Medicine, Johns Hopkins University School of Medicine
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Michael T. Melia
7Department of Medicine, Johns Hopkins University School of Medicine
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Cynthia L. Sears
7Department of Medicine, Johns Hopkins University School of Medicine
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Carlos A. Pardo
3Department of Neurology, Johns Hopkins University School of Medicine
5Department of Pathology, Johns Hopkins University School of Medicine
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  • For correspondence: cpardov1@jhmi.edu
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Abstract

Objective To determine the feasibility of next-generation sequencing (NGS) microbiome approaches in the diagnosis of infectious disorders in brain or spinal cord biopsies in patients with suspected central nervous system (CNS) infections.

Methods In a prospective-pilot study, we applied NGS in combination with a new computational analysis pipeline to detect the presence of pathogenic microbes in brain or spinal cord biopsies from ten patients with neurological problems indicating possible infection but for whom conventional clinical and microbiology studies yielded negative or inconclusive results.

Results Direct DNA and RNA sequencing of brain tissue biopsies generated 8.3 million to 29.1 million sequence reads per sample, which successfully identified with high confidence the infectious agent in three patients, identified possible pathogens in two more, and helped to understand neuropathological processes in three others, demonstrating the power of large-scale unbiased sequencing as a novel diagnostic tool. Validation techniques confirmed the pathogens identified by NGS in each of the three positive cases. Clinical outcomes were consistent with the findings yielded by NGS on the presence or absence of an infectious pathogenic process in eight of ten cases, and were non-contributory in the remaining two.

Conclusions NGS-guided metagenomic studies of brain, spinal cord or meningeal biopsies offer the possibility for dramatic improvements in our ability to detect (or rule out) a wide range of CNS pathogens, with potential benefits in speed, sensitivity, and cost. NGS-based microbiome approaches present a major new opportunity to investigate the potential role of infectious pathogens in the pathogenesis of neuroinflammatory disorders.

Footnotes

  • Study Funding: NIH R01 HG006677, U. S. Army Research Office W911NF-14-1-0490 and The Bart McLean Fund for Neuroimmunology Research-Johns Hopkins Project Restore.

  • Search Terms: Microbiome, metagenomics, brain biopsy, infections (135), sequencing.

  • Authors Contributions: Steven L. Salzberg: Study concept or design, computational biology data analysis, study supervision, drafting/revising the manuscript, obtaining funding.

    Florian P. Breitwieser: Computational biology data analysis, revising manuscript

    Anupama Kumar: Tissue processing, clinical data analysis, revising manuscript

    Haiping Hao: Tissue processing, DNA/RNA sequencing

    Peter Burger: Neuropathology analysis, revising manuscript

    Fausto J. Rodriguez: Neuropathology analysis, revising manuscript

    Michael Lim: Neurosurgical procedure, revising manuscript

    Alfredo Quiñones-Hinojosa: Neurosurgical procedure, revising manuscript

    Gary L. Gallia: Neurosurgical procedure, revising manuscript

    Jeffrey A Tornheim: Clinical data analysis, revising manuscript

    Michael T. Melia: Clinical data analysis, revising manuscript

    Cynthia L. Sears: Clinical data analysis, revising manuscript

    Carlos A. Pardo: Study concept or design, neuropathology analysis, clinical data analysis, study supervision, drafting/revising the manuscript, obtaining funding.

  • Study Funding: This research was supported in part by NIH under grant ROI HG006677 (S.L.S.) by the U. S. Army Research Office under grant number W911NF-14-1-0490 (S.L.S.) and The Bart McLean Fund for Neuroimmunology Research-Johns Hopkins Project Restore (CAP).

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-ND 4.0 International license.
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Posted February 09, 2016.
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Next-generation sequencing in neuropathological diagnosis of infections of the nervous system
Steven L. Salzberg, Florian P. Breitwieser, Anupama Kumar, Haiping Hao, Peter Burger, Fausto J. Rodriguez, Michael Lim, Alfredo Quiñones-Hinojosa, Gary L. Gallia, Jeffrey A. Tornheim, Michael T. Melia, Cynthia L. Sears, Carlos A. Pardo
bioRxiv 039222; doi: https://doi.org/10.1101/039222
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Next-generation sequencing in neuropathological diagnosis of infections of the nervous system
Steven L. Salzberg, Florian P. Breitwieser, Anupama Kumar, Haiping Hao, Peter Burger, Fausto J. Rodriguez, Michael Lim, Alfredo Quiñones-Hinojosa, Gary L. Gallia, Jeffrey A. Tornheim, Michael T. Melia, Cynthia L. Sears, Carlos A. Pardo
bioRxiv 039222; doi: https://doi.org/10.1101/039222

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