Comparison of direct RNA sequencing of Orthoavulavirus javaense using two different chemistries on the MinION platform.

Rapidly identifying and sequencing viral pathogens in poultry flocks can substantially reduce economic loss especially during disease outbreaks. Current next generation sequencing technologies require multi-step laboratory-intensive workflows to generate sequence data which precludes field adaptation. In this study, we hypothesized that direct RNA sequencing (DRS) using an Oxford Nanopore Technology (ONT) MinION device would enable sequencing of the full-length viral RNA genome of Orthoavulavirus javaense (OAVJ), the causative of Newcastle disease, a major poultry challenge. The data demonstrate that a custom OAVJ-specific adapter paired with the ONT DRS kits enables capture and sequencing of OAVJ viral RNAs. Further, the new ONT SQK-RNA004 chemistry and flow cells, paired with the associated super accurate base calling workflow improves on read quality and length compared to the previous SQK-RNA002 chemistry. This is the first report of a method to sequence near full-length viral RNA genome of a member of the Paramyxoviridae family. While additional improvements in DRS are needed before widespread adaptation of this method for rapid field sequencing, DRS of OAVJ has the potential to enable further studies into the viral epitranscriptome and its role in infection and pathogenesis.