Laboratory validation of a clinical metagenomic sequencing assay for pathogen detection in cerebrospinal fluid.
Miller S, Naccache SN, Samayoa E, Messacar K, Arevalo S, Federman S, Stryke D, Pham E, Fung B, Bolosky WJ, Ingebrigtsen D, Lorizio W, Paff SM, Leake JA, Pesano R, DeBiasi R, Dominguez S, Chiu CY
Genome Res. Apr 2019. doi: 10.1101/gr.238170.118
COMMENT: In this work metagenomics massive sequencing is used to detect pathogens (viruses, bacteria, fungi, and parasites) in cases of encephalitis and meningitis. This kind of metagenomics assay provides a method theoretically able to detect any type of organism in only one assay. The existence of better bioinformatics pipelines and quicker NGS technologies allow doing the assay with the sufficient quality and in clinically acceptable time.
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.
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.
Challenge study: The Aseptic Meningitis and Encephalitis Study (AMES) is a prospective cohort study enrolling children presenting to CHCO with culture-negative meningitis and encephalitis since 2012. A subset of CSF samples (n = 20) with sufficient residual volume of 600 µL from subjects with known and unknown etiologies was coded for mNGS testing as a challenge set. Samples were processed in a blinded fashion at UCSF and results discussed in clinical context with site investigators at CHCO over web-based teleconferencing
mNGS: metagenomics Next Generation Sequencing
CSF: CerebroSpinal Fluid
CHCO: Children’s Hospital Colorado
UCSF: University of California, San Francisco