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Keywords: Diagnostics; Genome sequencing; Genetic testing; Genomics

C.R. Marshall 1; N.J. Lennon 2; S. Chowdhury 3; R.J. Taft 4; D.J. Stavropoulos 1; M.S. Lebo 5; S.M. Harrison 5; J.G. Buchan 6; P. Liu 7; S. Kulkarni 7; D. Dimmock 3; J.W. Belmont 4; D. Bick 8; E.A. Worthey 8; R. Rowsey 9; E.W. Klee 9; H.M. Kearney 9; Medical Genome Initiative

1) Genome Diagnostics, The Hospital for Sick Children, Toronto, ON, Canada; 2) Broad Institute of MIT and Harvard, Cambridge, MA, USA; 3) 3Rady Children's Institute for Genomic Medicine, San Diego, CA, USA; 4) Illumina Inc., San Diego, CA, USA; 5) Laboratory for Molecular Medicine, Partners HealthCare Personalized Medicine, Cambridge, MA, USA; 6) Stanford Medicine Clinical Genomics Program, Stanford Health Care, Stanford, CA; 7) Baylor Genetics Laboratories, Houston, TX, USA; 8) HudsonAlpha Institute for Biotechnology, Huntsville, Alabama, USA; 9) Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN, USA

Current standard-of-care for genetically heterogeneous phenotypes often employs serial targeted gene testing, chromosomal microarray analysis and/or whole exome sequencing (WES), resulting in a lengthy time to diagnosis. Whole genome sequencing (WGS) addresses many of the technical limitations of these methodologies and can detect most forms of variation in a single test. Our group and others have demonstrated the diagnostic superiority of WGS compared with conventional testing in pediatric patients and critically ill infants.

Although WGS is poised to become a first-tier test for diagnosis of patients with rare genetic disorders, technical challenges as well as a lack of standards in both defining and deploying a clinical WGS test remain. To address these challenges, the Medical Genome Initiative (https://medgenomeinitiative.org/) was formed as a consortium of leading health care and research organizations in the US and Canada with an overarching mission to expand access to high quality clinical WGS. The work presented here focuses on the analytical validation of a clinical WGS test and aims to provide practical recommendations based on the consensus of group members

An analytical validation working group was formed to share common challenges faced by laboratories during the implementation of clinical WGS which led to the identification of practical solutions employed by the group. A series of discussions and surveys yielded consensus on several important topics: 1) Intended use and composition of a clinical WGS test including recommendations for the types of variation that should be reported; 2) the types of controls required for analytical validation of a clinical WGS test including reference standards and either commercially available or laboratory held positive controls for each variant type; 3) measurement of genome completeness (coverage and uniformity) to define WGS performance (callability) that can be tied to variant calling sensitivity; 4) a validation framework that includes accounting for genome complexity and variant type since both will affect test accuracy; 5) use of a comprehensive set of performance metrics for ongoing monitoring of quality and periodic use of positive controls dependent on sample volume.

These consensus recommendations may reduce the burden on new-to-WGS laboratories who wish to introduce WGS into clinical practice, and more importantly may serve to support safe and effective WGS testing for genetic disease.