Project Number: 2050-21310-005-21-I
Project Type: Interagency Reimbursable Agreement
Start Date: Oct 1, 2018
End Date: Aug 31, 2020
The proposed research supports the ARS’ mission of conducting research to improve nutrition, sustainability, and environmental compatibility of aquaculture. For this project, we will work toward improving our understanding of fish-disease dynamics, with the long-term goal of reducing losses to the U.S. rainbow trout industry. For this two-year period, we will address this goal with the following objectives: Objective #1 - To perform a multi-pathogen challenge and measure disease performance responses among trout strains with different genetic backgrounds. Objective #2 - To examine innate gene expression for the identification of genes differing among trout strains with different genetic backgrounds. Objective #3 - To integrate innate gene expression with disease performance traits for the identification of candidate genes underlying non-specific immunity in trout.
After a decade of selective breeding, we have developed a rainbow trout strain that thrives on an all-plant protein diet. From this selection, this strain has also undergone positive selection for non-specific disease resistance. The relationship between selection for alternative feeds and enhanced non-specific immunity has largely been unexplored but has the potential for great impact. The goal of this project is to significantly reduce disease losses in the U.S. rainbow trout industry by identifying key physiological factors linking genetics-diet-immunity and utilizing this information to enhance breeding/selection programs for multi-disease resistance and in the development of more effective health maintenance strategies. We will measure gene expression along with performance traits among three unique rainbow trout strains after multi-pathogen challenge: selected for resistance to IHNV and Flavobacterium columnare; not selected for disease resistance; and selected for dietary utilization and displaying an enhanced non-specific pathogen resistance. Four families consisting of 104 fish from three different strains of rainbow trout (12 families total) will be separately disease challenged at 63 days post-spawning to compare their responses to infectious hematopoietic necrosis virus (IHNV) and bacterial (Flavobacterium columnare) pathogen exposure. Relative percent survival (RPS) and mean number of days to death (MDD) will be calculated for each group. IHNV or F. columnare will be re-isolated from at least 20% of the fish that die each day to confirm the cause of death. For analysis of gene expression assessing disease resistance (or susceptibility) related to innate immune response, 80 fish in total from each strain and disease treatment will be randomly sampled immediately prior to challenge (0 hr) and at four time-points post-infection (4, 12, 24, and 48 hr). The entire intestine will be removed, slit lengthways, the mucus removed by scraping with a scalpel, and then saving a section of the distal intestine. Serum from whole blood and mucus will be used for lysozyme testing, and head kidney, spleen, and distal intestine (described above) will be collected. Total RNA will be isolated from collected tissue samples and will then be made into directional RNA-sequencing libraries for RNA-sequencing. All Oncorhynchus species rRNAs available in GenBank and the rainbow trout transcriptome will be used to align against RNA-seq data from the study. Read counts will be used to calculate gene expression fold-changes. Differentially expressed genes comparisons will be made between time-points and strains within a given treatment. For functional characterization of differentially expressed genes, Gene Ontology (GO) enrichment analyses will be performed. Weighted correlation network analysis (WGCNA) will be performed to determine consensus gene co-expression networks among strains post-infection. Perturbations to identified biological pathways will be shown, and functionally-similar gene families that are co-expressed among strains will be used to compile a candidate gene list of mechanistic genes involved in non-specific innate immunity.