Nanopore Sequencing Is a Credible Alternative to Recover Complete Genomes of Geminiviruses

• Published in: Microorganisms
• Main Authors: Selim Ben Chehida, Denis Filloux, Emmanuel Fernandez, Oumaima Moubset, Murielle Hoareau, Charlotte Julian, Laurence Blondin, Jean-Michel Lett, Philippe Roumagnac, Pierre Lefeuvre
• Format: Article
• Language: English
• Published: MDPI AG 2021-04-01
• Subjects: MinION; nanopore sequencing; rolling circle amplification; viral metagenomics; CRESS DNA; capulavirus
• Online Access: https://www.mdpi.com/2076-2607/9/5/903

Next-generation sequencing (NGS), through the implementation of metagenomic protocols, has led to the discovery of thousands of new viruses in the last decade. Nevertheless, these protocols are still laborious and costly to implement, and the technique has not yet become routine for everyday virus characterization. Within the context of CRESS DNA virus studies, we implemented two alternative long-read NGS protocols, one that is agnostic to the sequence (without a priori knowledge of the viral genome) and the other that use specific primers to target a virus (with a priori). Agnostic and specific long read NGS-based assembled genomes of two capulavirus strains were compared to those obtained using the gold standard technique of Sanger sequencing. Both protocols allowed the detection and accurate full genome characterization of both strains. Globally, the assembled genomes were very similar (99.5–99.7% identity) to the Sanger sequences consensus, but differences in the homopolymeric tracks of these sequences indicated a specific lack of accuracy of the long reads NGS approach that has yet to be improved. Nevertheless, the use of the bench-top sequencer has proven to be a credible alternative in the context of CRESS DNA virus study and could offer a new range of applications not previously accessible.