Evaluating methods for Avian avulavirus-1 whole genome sequencing

Avian avulavirus-1 (AAvV-1, previously Newcastle Disease Virus) is responsible for poultry and wild birds' disease outbreaks. Numerous whole genome sequencing methods were reported for this virus. These methods included cloning, specific primers amplification, shotgun PCR approaches, Sequence Independent Single Primer Amplification and next generation sequencing platform kits. Three methods were used to sequence 173 Israeli Avian avulavirus-1 field isolates and one vaccine strain (VH). The sequencing was performed on Proton and Ion Torrent Personal Genome Machine and to a lesser extent, Illumina MiSeq and NextSeq sequencers. Target specific primers (SP) and Sequence Independent Single Primer Amplification (SISPA) products sequenced via the Ion torrent sequencer had a high error rate and truncated genomes. All the next generation sequencing platform sequencing kits generated high sequence accuracy and near-complete genomic size. A high level of mutations was observed in the intergenic regions between the avian avulavirus-1 genes. Within genes, multiple regions are more mutated than the Fusion region currently used for typing. Our findings suggest that the whole genome sequencing by the Ion torrent sequencing kit is sufficient. However, when higher fidelity is desired, the Illumina NextSeq and Proton torrent sequencing kits were found to be preferable. •PCR amplification of the viral genome, results in partial genome sequences (72% and 98% respectively).•Primer based sequencing methods have 5 to 5.6 times more N reads compared to NGS platform sequencing kits, indicating poor sequence quality.•The Ion Torrent PGM sequencing results in sequencing 99% of the viral genome.•ProtonTorrent and Illumina Nextseq sequenced 99-100% of the viral genome, leading a to full genome sequence at high coverage.•The virus is mostly mutated within a host at intergenic regions.