Identification and genetic characterization of a novel Orthobunyavirus species by a straightforward high-throughput sequencing-based approach

Shifman, Ohad; Cohen-Gihon, Inbar; Beth-Din, Adi; Zvi, Anat; Laskar, Orly; Paran, Nir; Epstein, Eyal; Stein, Dana; Dorozko, Marina; Wolf, Dana; Yitzhaki, Shmuel; Shapira, Shmuel C.; Melamed, Sharon; Israeli, Ofir

Identification and characterization of novel unknown viruses is of great importance. The introduction of high-throughput sequencing (HTS)-based methods has paved the way for genomics-based detection of pathogens without any prior assumptions about the characteristics of the organisms. However, the use of HTS for the characterization of viral pathogens from clinical samples remains limited. Here, we report the identification of a novel Orthobunyavirus species isolated from horse plasma. The identification was based on a straightforward HTS approach. Following enrichment in cell culture, RNA was extracted from the growth medium and rapid library preparation, HTS and primary bioinformatic analyses were performed in less than 12 hours. Taxonomical profiling of the sequencing reads did not reveal sequence similarities to any known virus. Subsequent application of de novo assembly tools to the sequencing reads produced contigs, of which three showed some similarity to the L, M, and S segments of viruses belonging to the Orthobunyavirus genus. Further refinement of these contigs resulted in high-quality, full-length genomic sequences of the three genomic segments (L, M and S) of a novel Orthobunyavirus. Characterization of the genomic sequence, including the prediction of open reading frames and the inspection of consensus genomic termini and phylogenetic analysis, further confirmed that the novel virus is indeed a new species, which we named Ness Ziona virus.