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).