title: Unbiased Detection of Respiratory Viruses by Use of RNA Sequencing-Based Metagenomics: a Systematic Comparison to a Commercial PCR Panel.
abstract: Current infectious disease molecular tests are largely pathogen specific, requiring test selection based on the patient's symptoms. For many syndromes caused by a large number of viral, bacterial, or fungal pathogens, such as respiratory tract infections, this necessitates large panels of tests and has limited yield. In contrast, next-generation sequencing-based metagenomics can be used for unbiased detection of any expected or unexpected pathogen. However, barriers for its diagnostic implementation include incomplete understanding of analytical performance and complexity of sequence data analysis. We compared detection of known respiratory virus-positive (n= 42) and unselected (n= 67) pediatric nasopharyngeal swabs using an RNA sequencing (RNA-seq)-based metagenomics approach and Taxonomer, an ultrarapid, interactive, web-based metagenomics data analysis tool, with an FDA-cleared respiratory virus panel (RVP; GenMark eSensor). Untargeted metagenomics detected 86% of known respiratory virus infections, and additional PCR testing confirmed RVP results for only 2 (33%) of the discordant samples. In unselected samples, untargeted metagenomics had excellent agreement with the RVP (93%). In addition, untargeted metagenomics detected an additional 12 viruses that were either not targeted by the RVP or missed due to highly divergent genome sequences. Normalized viral read counts for untargeted metagenomics correlated with viral burden determined by quantitative PCR and showed high intrarun and interrun reproducibility. Partial or full-length viral genome sequences were generated in 86% of RNA-seq-positive samples, allowing assessment of antiviral resistance, strain-level typing, and phylogenetic relatedness. Overall, untargeted metagenomics had high agreement with a sensitive RVP, detected viruses not targeted by the RVP, and yielded epidemiologically and clinically valuable sequence information.
authors.affiliation: University of Utah School of Medicine, Department of Pathology, Salt Lake City, Utah, USA.
authors.forename: Erin H
authors.initials: EH
authors.lastname: Graf
authors.affiliation: University of Utah School of Medicine, Department of Biomedical Informatics, Salt Lake City, Utah, USA ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, Utah, USA.
authors.forename: Keith E
authors.initials: KE
authors.lastname: Simmon
authors.affiliation: ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, Utah, USA.
authors.forename: Keith D
authors.initials: KD
authors.lastname: Tardif
authors.affiliation: ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, Utah, USA.
authors.forename: Weston
authors.initials: W
authors.lastname: Hymas
authors.affiliation: Department of Human Genetics, University of Utah, Salt Lake City, Utah, USA.
authors.forename: Steven
authors.initials: S
authors.lastname: Flygare
authors.affiliation: University of Utah School of Medicine, Department of Biomedical Informatics, Salt Lake City, Utah, USA USTAR Center for Genetic Discovery, Salt Lake City, Utah, USA.
authors.forename: Karen
authors.initials: K
authors.lastname: Eilbeck
authors.affiliation: Department of Human Genetics, University of Utah, Salt Lake City, Utah, USA USTAR Center for Genetic Discovery, Salt Lake City, Utah, USA.
authors.forename: Mark
authors.initials: M
authors.lastname: Yandell
authors.affiliation: University of Utah School of Medicine, Department of Pathology, Salt Lake City, Utah, USA ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, Utah, USA robert.schlaberg@path.utah.edu.
authors.forename: Robert
authors.initials: R
authors.lastname: Schlaberg
mesh_terms: D002675:Child, Preschool; D019295:Computational Biology / Q000379:methods; D005260:Female; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D008297:Male; D056186:Metagenomics / Q000379:methods*; D009305:Nasopharynx / Q000821:virology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012141:Respiratory Tract Infections / Q000175:diagnosis* / Q000821:virology*; D012189:Retrospective Studies; D017423:Sequence Analysis, RNA / Q000379:methods*; D014780:Viruses / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*
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