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Laboratory validation of a clinical metagenomic sequencing assay for pathogen detection in cerebrospinal fluid.
Miller, Steve; Naccache, Samia N; Samayoa, Erik; Messacar, Kevin; Arevalo, Shaun; Federman, Scot; Stryke, Doug; Pham, Elizabeth; Fung, Becky; Bolosky, William J; Ingebrigtsen, Danielle; Lorizio, Walter; Paff, Sandra M; Leake, John A; Pesano, Rick; DeBiasi, Roberta; Dominguez, Samuel; Chiu, Charles Y.
Afiliación
  • Miller S; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California 94143, USA.
  • Naccache SN; UCSF-Abbott Viral Diagnostics and Discovery Center, San Francisco, California 94143, USA.
  • Samayoa E; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California 94143, USA.
  • Messacar K; UCSF-Abbott Viral Diagnostics and Discovery Center, San Francisco, California 94143, USA.
  • Arevalo S; Department of Pathology and Laboratory Medicine, Children's Hospital Los Angeles, Los Angeles, California 90027, USA.
  • Federman S; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California 94143, USA.
  • Stryke D; Department of Pediatrics, Children's Hospital Colorado and University of Colorado School of Medicine, Aurora, Colorado 80045, USA.
  • Pham E; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California 94143, USA.
  • Fung B; UCSF-Abbott Viral Diagnostics and Discovery Center, San Francisco, California 94143, USA.
  • Bolosky WJ; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California 94143, USA.
  • Ingebrigtsen D; UCSF-Abbott Viral Diagnostics and Discovery Center, San Francisco, California 94143, USA.
  • Lorizio W; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California 94143, USA.
  • Paff SM; UCSF-Abbott Viral Diagnostics and Discovery Center, San Francisco, California 94143, USA.
  • Leake JA; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California 94143, USA.
  • Pesano R; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California 94143, USA.
  • DeBiasi R; Microsoft Research, Redmond, Washington 98052, USA.
  • Dominguez S; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California 94143, USA.
  • Chiu CY; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, California 94143, USA.
Genome Res ; 29(5): 831-842, 2019 05.
Article en En | MEDLINE | ID: mdl-30992304
Metagenomic next-generation sequencing (mNGS) for pan-pathogen detection has been successfully tested in proof-of-concept case studies in patients with acute illness of unknown etiology but to date has been largely confined to research settings. Here, we developed and validated a clinical mNGS assay for diagnosis of infectious causes of meningitis and encephalitis from cerebrospinal fluid (CSF) in a licensed microbiology laboratory. A customized bioinformatics pipeline, SURPI+, was developed to rapidly analyze mNGS data, generate an automated summary of detected pathogens, and provide a graphical user interface for evaluating and interpreting results. We established quality metrics, threshold values, and limits of detection of 0.2-313 genomic copies or colony forming units per milliliter for each representative organism type. Gross hemolysis and excess host nucleic acid reduced assay sensitivity; however, spiked phages used as internal controls were reliable indicators of sensitivity loss. Diagnostic test accuracy was evaluated by blinded mNGS testing of 95 patient samples, revealing 73% sensitivity and 99% specificity compared to original clinical test results, and 81% positive percent agreement and 99% negative percent agreement after discrepancy analysis. Subsequent mNGS challenge testing of 20 positive CSF samples prospectively collected from a cohort of pediatric patients hospitalized with meningitis, encephalitis, and/or myelitis showed 92% sensitivity and 96% specificity relative to conventional microbiological testing of CSF in identifying the causative pathogen. These results demonstrate the analytic performance of a laboratory-validated mNGS assay for pan-pathogen detection, to be used clinically for diagnosis of neurological infections from CSF.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Encefalitis / Metagenómica / Secuenciación de Nucleótidos de Alto Rendimiento / Meningitis Aséptica / Mielitis Tipo de estudio: Diagnostic_studies / Prognostic_studies Límite: Child / Humans Idioma: En Revista: Genome Res Asunto de la revista: BIOLOGIA MOLECULAR / GENETICA Año: 2019 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Encefalitis / Metagenómica / Secuenciación de Nucleótidos de Alto Rendimiento / Meningitis Aséptica / Mielitis Tipo de estudio: Diagnostic_studies / Prognostic_studies Límite: Child / Humans Idioma: En Revista: Genome Res Asunto de la revista: BIOLOGIA MOLECULAR / GENETICA Año: 2019 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos