Wang, J.

Infectious salmon anaemia virus (ISAV) poses a major threat to Atlantic salmon (Salmo salar), causing significant economic losses in aquaculture. Enhancing the genetic acquisition of resistance to ISAV is therefore important for sustainable fish farming. While previous genome-wide CRISPR screens have identified potential genetic targets for ISAV resistance in salmon, no causative genes have been functionally validated. In the current study, we investigated three key genes-st6gal1, ccdc115 and sec14l1-involved in the ISAV infection pathway as putative candidates for resistance. The role of these genes in ISA resistance was assessed by CRISPR/Cas9-mediated knockout (KO) in Atlantic salmon kidney (ASK) cells. We achieved high editing efficiencies with large fragment deletions (up to 90 bp) using double sgRNAs at these three loci (89%, 100%, and 100%) with KO scores of 100, 71, and 100, respectively. Although gene KO induced varying levels of cell mortality, the deletion of these genes significantly reduced productive ISAV replication. Our study provides a functional validation of antiviral candidate genes in Atlantic salmon and demonstrates the utility of gene editing as a tool to dissect the genetic basis of disease resistance. The findings offer valuable insights into the mechanisms underlying viral resistance in salmon and may guide future breeding and genetic engineering efforts to improve disease resistance in aquaculture.