Application of nanopore adaptive sampling for metagenomic detection of tick-borne RNA viruses

Nanopore sequencing is a powerful tool for real-time pathogen detection and genomic characterization; however, its application to individual ticks is limited by abundant host-derived nucleic acids and low viral RNA levels. In this study, we applied nanopore adaptive sampling (NAS) to sequence viral RNA from individual Haemaphysalis (H.) ticks collected in the Republic of Korea (ROK). By combining NAS with long-read sequencing, high-resolution genome assembly can be achieved from samples containing low-abundance viral RNA and relatively short complementary DNA (cDNA) fragments generated during library preparation. These results indicate that NAS remains effective under suboptimal fragment-size conditions and improves genome assembly compared to conventional nanopore workflows. Phylogenetic analyses revealed that the detected Dabieshan tick virus (DTV) sequences were clustered with isolates from China and Japan, suggesting regional circulation facilitated by the widespread distribution of H. longicornis. Unlike previous studies relying on pooled samples without selective sequencing, NAS allowed high-resolution viral genome assembly from single ticks. These findings confirm the presence and genotypes of DTV for the first time in the ROK and demonstrate NAS as a practical, scalable approach for tick-borne RNA virus surveillance in single ticks, improving genomic assembly and supporting the monitoring of emerging tick-borne viruses in endemic regions.