Comparison of the Performance of Two Targeted Metagenomic Virus Capture Probe-Based Methods Using Synthetic Viral Sequences and Clinical Samples

Viral enrichment by probe hybridization has been reported to significantly increase the sensitivity of viral metagenomics.This study compares the analytical performance of two targeted metagenomic virus capture probe-based methods: i) SeqCap EZ HyperCap by Roche (ViroCap) and ii) Twist Comprehensive Viral Research Panel workflow, for diagnostic use. Sensitivity, specificity, limit of detection, and effect of human background DNA were analysed, using synthetic viral sequences, clinical and reference samples with known viral loads.Sensitivity and specificity were 95% and higher for both methods. Combining thresholds for viral sequence read counts and genome coverage (respectively 500 reads per million and 10% coverage) resulted in optimal prediction of true positive results. Limits of detection were approximately 50-500 copies/ml for both methods. Increasing proportions of spike-in cell free human background sequences did not negatively affect viral detection.These data show analytical performances in ranges applicable to clinical samples, for both probe hybridization metagenomic approaches. This study supports further steps towards more widespread use of viral metagenomics for pathogen detection, in clinical and surveillance settings using low biomass samples.### Competing Interest StatementThe authors have declared no competing interest.### Funding StatementThis study did not receive any funding.### Author DeclarationsI confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained.YesThe details of the IRB/oversight body that provided approval or exemption for the research described are given below:Ethics committee/IRB of the Leiden University Medical Center (IRB Leiden/The Hague/Delft) gave ethical approval for this work (CME number B20.002, 2020/2022).I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals.YesI understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance).YesI have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable.YesAll data produced in the present study are available upon reasonable request to the authors.