Infectious Salmon Anaemia Virus (ISAV) causes a notifiable disease in Atlantic salmon that represents a major problem for salmon breeders and producers worldwide. Current prevention and therapeutic methods are not fully effective, and therefore selective breeding to produce ISAV-resistant strains of Atlantic salmon (Salmo salar) is a high priority for the industry. Genomic selection and potentially genome editing can be applied to enhance host resistance. Both approaches can benefit from increased knowledge on the genetic and functional mechanisms of resistance to ISAV. Here we have combined used bulk and single-cell RNA sequencing in in vitro and in vivo models to study Atlantic salmon immune response against ISAV.
First, we studied the transcriptomic changes in response to ISAV in four different tissues (heart, gills, head kidney and spleen) at three timepoints (pre-challenge, 7 and 14 dpi), finding a clear but unique response to ISAV in each tissue. Comparisons between four resistant and four susceptible fish per timepoint, selected based on their genomic breeding values estimated from a disease challenge in 1,353 fish from the same population, revealed a large number of differentially expressed genes in the head kidney, while the differences in the other tissues were small.
To better understand the host-response interaction at the cellular level, and evaluate the importance of potential candidate resistant genes, we investigated the Atlantic salmon response to ISAV at the single cell level using an in vitro model (SHK-1 cells, Atlantic salmon head kidney 1). Cells were challenged with ISAV and single-cell sequenced at 0, 24, 48 and 92h post-infection. Early infection (24h) did not significantly alter the transcriptome of Atlantic salmon cells, however cells from 48h and 96h samples clustered clearly apart whether they were infected or not, suggesting paracrine signalling. At 48 and 96h infected cells significantly up-regulated several genes related to ubiquitination. To further investigate this process we performed immunoprecipitation of ubiquitinated proteins in SHK-1 cells, finding an increase of ubiquitination at 24 and 48h after ISAV infection. Mass spectrometry of the ubiquitin-proteome revealed key host immune mechanisms regulated by ubiquitination in response to ISAV. These results improve our understanding of host response to ISAV in salmon and highlight potential target to improve host resistance.