Tilapia lake virus (TiLV) is a 10 -segmented single-stranded negative-sense RNA virus first identified in 2014. TiLV can cause up to 90% mortality in tilapia and poses a severe threat to the global tilapia industry . However, scientific knowledge and research relating to TiLV are very limited. The unavailability of a reverse genetics system (RGS) has made it challenging to elucidate the molecular mechanisms underlying TiLV replication and pathogenesis. Firstly, we identified that TiLV can infect and replicate in the Vero E6 cells after susceptibility testing of many mammalian cell lines. Based on this discovery and through sequence analyses of the TiLV genome, we successfully generated recombinant TiLVs . This was achieved by transfecting 10 synthetic plasmids containing the complete TiLV reference genome ( GCF_001630085.1, TiLV isolate Til-4-2011) into Vero E6 cells that were co-cultured with fish-derived E11 cells. Sequence analysis of the genome of the rescued virus and comparison with parental TiLV genomic segments confirmed the successful rescue. We then sequenced the complete genome of a wild-type TiLV strain originating from Israel (named TiLV-Israel-HK) , the new recombinant TiLV (TiLV -Israel-HK) was also successfully recovered. Furthermore, we used our system to generate reassortant viruses of TiLV . All TiLVs rescued using the RGS caused a clear cytopathic effect in E11 cells, which was indistinguishable from wild-type virus. This RGS will allow in-depth characterization of this emerged virus and facilitate the development of vaccines and antiviral therapeutics.
In summary , we have established the first RGS for TiLV. We believe t his RGS will bring a lot of convenience for studying TiLV and accelerate research relating to TiLV.