Rainbow trout is one of the most commonly farmed salmonid species across the world. Infectious diseases represent a significant threat to the sustainable development of fish farming. Understanding the basis of fish natural resistance to pathogens is important for genetic selection. This project was developed in the frame of “ Advancing European Aquaculture by Genome Functional Annotation” (AQUAFAANG). The objective was to achieve a detailed functional annotation analysis of two rainbow trout isogenic lines chosen based on their resistance/susceptibility status to two pathogens, a bacterium (Flavobacterium psychrophilum , Fp) and a virus (Viral Hemorrhagic Septicemia Virus, VHSv). The first isogenic line is resistant to Fp infection by injection and susceptible to VHSv infection by immersion; the second is resistant to VHSv and susceptible to Fp.
Experimental disease challenges were performed by intramuscular injection for standardisation of the stimulation. Two days post-infection, the head kidneys of 32 fish (2 lines; 4 experimental conditions: control Fp, injected Fp, control VHSv and injected VHSv; 4 fish per line and per condition) were sampled for RNAseq and ATACseq libraries construction. The 32 RNAseq and 32 ATACseq libraries were mapped on the Ensembl rainbow trout genome assembly (strain Arlee), and count tables were generated using bioinformatics pipeline nf-core/RNAseq (3.14) and nf-core/atacseq (2.12), respectively. To reduce the false discovery rate during read count, due to the high level of duplication in salmonids and multiple transcript isoforms, we retained uniquely mapped reads and only kept the longest transcript isoform for each gene in the reference database for annotation . Differential gene expression analysis and differential chromatin accessibility analysis were performed with DESeq2 for each disease separately . The integration of RNAseq and ATACseq data was performed by Regularized Generalized Canonical Correlation Analysis (RGCCA), Multi-omic Factor Analysis (MOFA), Weighted Gene Co-expression Network Analysis (WGCNA) and Similarity Network Fusion (SNF) , retaining only intersection of the mechanisms detected by the different methods.
Divergent transcriptional responses between the resistant and susceptible lines were revealed at the basal level and following infection. In particular, after VHSv infection, higher induction rates of interferon stimulatory genes were detected in the resistant line. Also, markers of inflammatory response were up regulated in the two lines following Fp infection. In addition, d ifferences in chromatin accessibility were observed between infected and control fish for VHSv but not for Fp challenges. The integration between RNAseq and ATACseq data will lead to a better understanding of molecular mechanisms involved in the contrasted response of the two isogenic lines to both pathogens.