ArticleMicrobiome2017
Optimization and validation of sample preparation for metagenomic sequencing of viruses in clinical samples.
Article in Microbiome, 2017. The graph could read no effect estimate from its abstract, so it casts no vote on the map. An erratum has been issued. Cited by 44 papers.
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Who cites it
44 citing papers in PubMed.
- Empowering One Health with metagenomics.One health outlook · 2026Review
- One health viral metagenomics for pathogen surveillance: robust mNGS workflows for viral detection and genome recovery from swab and tissue specimens.BMC microbiology · 2026Article
- DNA Viral Size Fraction Metagenomics for Human Stool Samples.Journal of visualized experiments : JoVE · 2026Article
- Metagenomic characterization of the porcine respiratory virome on farms in northern Kazakhstan.Frontiers in veterinary science · 2026Article
- A comparison of methods for the optimal recovery of the human fecal virome.ISME communications · 2026Article
- Viral metagenomic analysis of human bocavirus in pediatric pneumonia: detection pattern and genetic characterization.Frontiers in cellular and infection microbiology · 2026Article
- teamNGS Balances Sensitivity for Viruses with Comprehensive Microbial Detection in Clinical Specimens.Microorganisms · 2025Article
- A tombus-like virus in patients with lower respiratory tract infection: an observational study based on meta-transcriptomic sequencing.Emerging microbes & infections · 2025Observational
- Development and performance evaluation of a clinical metagenomics approach for identifying pathogens in the whole blood from patients with undifferentiated fever.Frontiers in cellular and infection microbiology · 2025Article
- MetaAll: integrative bioinformatics workflow for analysing clinical metagenomic data.Briefings in bioinformatics · 2024Article
- Genomics for Arbovirus Surveillance: Considerations for Routine Use in Public Health Laboratories.Viruses · 2024Review
- Comparison of the performance of two targeted metagenomic virus capture probe-based methods using reference control materials and clinical samples.Journal of clinical microbiology · 2024Article
- Retrospective Genotyping of Enteroviruses Using a Diagnostic Nanopore Sequencing Workflow.Pathogens (Basel, Switzerland) · 2024Article
- Findings and methodologies in oral phageome research: a systematic review.Journal of oral microbiology · 2024Review
- Case report: Local bacteriophage therapy for fracture-related infection with polymicrobial multi-resistant bacteria: hydrogel application and postoperative phage analysis through metagenomic sequencing.Frontiers in medicine · 2024Article
- Next-Generation Sequencing for the Detection of Microbial Agents in Avian Clinical Samples.Veterinary sciences · 2023Review
- The International Virus Bioinformatics Meeting 2023.Viruses · 2023Article
- Evaluation of extraction and enrichment methods for recovery of respiratory RNA viruses in a metagenomics approach.Journal of virological methods · 2023Article
- Article
- Next Generation Sequencing Approaches to Characterize the Respiratory Tract Virome.Microorganisms · 2022Review
Corrections and comments
- Erratum issued
Authors and funding
9 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundSequence-specific PCR is the most common approach for virus identification in diagnostic laboratories. However, as specific PCR only detects pre-defined targets, novel virus strains or viruses not included in routine test panels will be missed. Recently, advances in high-throughput sequencing allow for virus-sequence-independent identification of entire virus populations in clinical samples, yet standardized protocols are needed to allow broad application in clinical diagnostics. Here, we describe a comprehensive sample preparation protocol for high-throughput metagenomic virus sequencing using random amplification of total nucleic acids from clinical samples.
resultsIn order to optimize metagenomic sequencing for application in virus diagnostics, we tested different enrichment and amplification procedures on plasma samples spiked with RNA and DNA viruses. A protocol including filtration, nuclease digestion, and random amplification of RNA and DNA in separate reactions provided the best results, allowing reliable recovery of viral genomes and a good correlation of the relative number of sequencing reads with the virus input. We further validated our method by sequencing a multiplexed viral pathogen reagent containing a range of human viruses from different virus families. Our method proved successful in detecting the majority of the included viruses with high read numbers and compared well to other protocols in the field validated against the same reference reagent. Our sequencing protocol does work not only with plasma but also with other clinical samples such as urine and throat swabs.
conclusionsThe workflow for virus metagenomic sequencing that we established proved successful in detecting a variety of viruses in different clinical samples. Our protocol supplements existing virus-specific detection strategies providing opportunities to identify atypical and novel viruses commonly not accounted for in routine diagnostic panels.
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