Whole genome sequencing of hepatitis C virus from dried blood spots by hybrid capture for genotype and antiviral resistance determination.

BACKGROUND

Dried blood spot (DBS) testing enables hepatitis C virus (HCV) testing in hard‑to‑reach patient populations but typically lacks full genotype and drug resistance information. This study evaluates a commercially available hybrid‑capture whole‑genome sequencing (WGS) workflow for HCV from DBS to determine suitability for clinical diagnostic and surveillance purposes.

METHODS

We analysed 147 DBS and 24 plasma samples known to be HCV‑RNA positive using the QIAGEN QIAseq xHYB HepC panel. Whole genome sequences were generated using an in-house custom designed iterative bioinformatic pipeline. We assessed coverage characteristics, genotyping accuracy, resistance‑associated mutation (RAM) detection, and analytical sensitivity to determine panel performance.

RESULTS

WGS success rates were high for both DBS (95%) and plasma (96%). Genome enrichment was uniform across major HCV genotypes, with no systematic dropouts. Genotyping showed 100% concordance with reference laboratory results including for novel genotype 1 subtypes. RAMs were detected in 27% of samples assessed, with full concordance with reference laboratory results in those with known resistance profiles. Analytical sensitivity was calculated at < 7301 IU/ml for WGS with slightly poorer performance when assessing the NS5a gene in isolation.

CONCLUSION

This protocol reliably generates high‑quality HCV genomes from DBS, enabling accurate genotype, subtype, and resistance profiling. This approach overcomes limitations of venous sampling in hard-to-reach populations and could support decentralised diagnostics and enhancing genomic surveillance in populations underserved by traditional healthcare pathways.