Metagenomic high-throughput sequencing (mHTS) is a hypothesis-free, universal pathogen detection technique for determination of the DNA/RNA sequences in a variety of sample types and infectious syndromes. mHTS is still in its early stages of translating into clinical application. To support the development, implementation and standardization of mHTS procedures for virus diagnostics, the European Society for Clinical Virology (ESCV) Network on Next-Generation Sequencing (ENNGS) has been established. The aim of ENNGS is to bring together professionals involved in mHTS for viral diagnostics to share methodologies and experiences, and to develop application recommendations. This manuscript aims to provide practical recommendations for the wet lab procedures necessary for implementation of mHTS for virus diagnostics and to give recommendations for development and validation of laboratory methods, including mHTS quality assurance, control and quality assessment protocols.

Center of Virology Medical University Vienna Vienna Austria

Clinical Virology Laboratory Medicine University Hospital Basel Basel Switzerland; Transplantation and Clinical Virology Department Biomedicine University of Basel Basel Switzerland

Department of Clinical Microbiology University Hospital of Region Zealand Slagelse Denmark

Department of Medical Microbiology Leiden University Medical Center Leiden the Netherlands

Department of Microbiology and Ecology Faculty of Medicine University of Valencia Valencia Spain

Department of Paediatrics and Medical Microbiology 2nd Faculty of Medicine Charles University Prague Czech Republic

Department of Preservation and Food Safety Technologies IATA CSIC Paterna Valencia Spain

Department of Virology Helsinki University Hospital Laboratory and University of Helsinki Helsinki Finland

Department of Virology University Medical Center Utrecht Utrecht the Netherlands

Dokuz Eylul University Faculty of Medicine Department of Medical Microbiology Division of Medical Virology Izmir Turkey

ErasmusMC Department of Viroscience Erasmus Medical Center Rotterdam the Netherlands

Faculty of Biology Medicine and Health Division of Infection Immunity and Respiratory Medicine University of Manchester Manchester United Kingdom

Friedrich Loeffler Institut Federal Research Institute for Animal Health Greifswald Insel Riems Germany

Institute of Biomedicine University of Turku Finland

Institute of Medical Microbiology Virology and Hygiene University Medical Center Hamburg Eppendorf Hamburg Germany

Institute of Medical Virology University of Zurich Zurich Switzerland

Institute of Virology Leipzig University Leipzig Germany

Laboratorium Klinische en Epidemiologische Virologie Leuven Belgium

Laboratory of Molecular Biology and Microbiology Instituto Tecnologico Agrario de Castilla y Leon Valladolid Spain

Life and Health Sciences Research Institute Porto Portugal

Medical Microbiology and Infection Control Amsterdam UMC Amsterdam the Netherlands; Laboratory of Experimental Virology Medical Microbiology and Infection Control Amsterdam UMC Amsterdam the Netherlands

Microbiology Department and NGS Platform University Hospital Henri Mondor Créteil France

Microbiology Services NHS Blood and Transplant London United Kingdom

Microbiology Virology and Infection Prevention and Control Great Ormond Street Hospital for Children NHS Foundation Trust United Kingdom

Nuffield Department of Medicine University of Oxford Oxford United Kingdom

University of Groningen University Medical Center Groningen Department of Medical Microbiology Groningen the Netherlands; Milner Centre for Evolution Department of Biology and Biochemistry University of Bath Bath United Kingdom

Virology Laboratory Genomics and Health Area Centre for Public Health Research Valencia Spain; CIBERESP Instituto de Salud Carlos 3 Madrid Spain

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