Rapid whole-genome based typing and surveillance of avipoxviruses using nanopore sequencing.

Avian pox is an infectious disease caused by avipoxviruses (APV), resulting in cutaneous and/or tracheal lesions. Poxviruses share large genome sizes (from 130 to 360 kb), featuring repetitions, deletions or insertions as a result of a long-term recombination history. The increasing performances of next-generation sequencing (NGS) opened new opportunities for surveillance of poxviruses, based on timely and affordable workflows. We investigated the application of the 3rd generation Oxford Nanopore Minion technology to achieve real-time whole-genome sequencing directly from lesions, without any enrichment or isolation step. Fowlpox lesions were sampled on hens, total DNA was extracted and processed for sequencing on a MinION, Oxford Nanopore. We readily generated whole APV genomes from cutaneous or tracheal lesions, without any isolation or PCR-based enrichment: Fowlpox virus reads loads ranged from 0.75% to 2.62% and reads up to 61 kbp were generated and readily assembled into 3 APV complete genomes. This long read size eases the assembly step and lowers the bioinformatics capacity requirements and processing time compared to huge sets of short reads. The complete genome analysis confirmed that these Fowlpox viruses cluster within clade A1 and host full length reticuloendotheliovirus (REV) inserts. The pathobiological relevance of REV insert, although a classical feature of fowlpoxviruses (FPVs), should be further investigated. Surveillance of emerging poxviruses could greatly benefit from real-time whole genome sequencing.