Comparison of targeted next-generation sequencing for whole-genome sequencing of Hantaan orthohantavirus in Apodemus agrarius lung tissues

초록

Orthohantaviruses, negative-sense single-strand tripartite RNA viruses, are a global public health threat. In humans, orthohantavirus infection causes hemorrhagic fever with renal syndrome or hantavirus cardiopulmonary syndrome. Whole-genome sequencing of the virus helps in identification and characterization of emerging or re-emerging viruses. Next-generation sequencing (NGS) is a potent method to sequence the viral genome, using molecular enrichment methods, from clinical specimens containing low virus titers. Hence, a comparative study on the target enrichment NGS methods is required for whole-genome sequencing of orthohantavirus in clinical samples. In this study, we used the sequence-independent, single-primer amplification, target capture, and amplicon NGS for whole-genome sequencing of Hantaan orthohantavirus (HTNV) from rodent specimens. We analyzed the coverage of the HTNV genome based on the viral RNA copy number, which is quantified by real-time quantitative PCR. Target capture and amplicon NGS demonstrated a high coverage rate of HTNV in Apodemus agrarius lung tissues containing up to 10(3)-10(4) copies/mu L of HTNV RNA. Furthermore, the amplicon NGS showed a 10-fold (10(2) copies/mu L) higher sensitivity than the target capture NGS. This report provides useful insights into target enrichment NGS for whole-genome sequencing of orthohantaviruses without cultivating the viruses.

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