1: Define phenotypic measures, estimate genetic and phenotypic parameters, and develop a selection index in Atlantic salmon for commercially important traits such as carcass weight, cold tolerance, fillet color, fat content, and sea lice resistance. 1A. Define phenotypic measures and estimate genetic parameters for sea lice resistance and fillet fatty acid levels in Atlantic salmon. 1B. Develop a multi-trait selection index in Atlantic salmon germplasm for carcass weight, fillet fatty acid levels, and sea lice resistance. 2: Evaluate and validate the usefulness of incorporating genotypic information into salmon selective breeding program. 3: Establish links between disease susceptible/resistant phenotypes and genotype for the Eastern Oyster, Crassostrea virginica. 3A: Define disease susceptible and disease resistant phenotypes in selectively-bred C. virginica families through disease challenges and transcriptome analysis. 3B: Discover polymorphisms in candidate genes for disease susceptibility and resistance in C. virginica and develop single nucleotide polymorphism (SNP) markers that can be genotyped in a high-throughput assay. 3C: Identify Single Nucleotide Polymorphisms (SNPs) associated with disease-susceptible and disease-resistant phenotypes in C. virginica.
The National Cold Water Marine Aquaculture Center (NCWMAC) addresses the coldwater marine aquaculture industry’s highest priority research needs. Coldwater aquaculture production has great potential for expansion, and both Atlantic salmon and Eastern oysters are widely accepted as seafood by American consumers. Commercial salmon and oyster producers predominantly utilize stocks that are not many generations removed from wild, unselected stocks. Salmon producers are legally required to culture certified stocks of North American salmon, and the NCWMAC is the only program supporting the US coldwater marine aquaculture industry and developing genetically improved salmon. Aquaculture of the Eastern oyster is a large segment of shellfish aquaculture in the US, and minimal selective breeding has been accomplished in this species. In both species, there is a need to improve the performance of existing stocks. This project plan proposes to meet this need through the following objectives: 1) define phenotypes, estimate genetic and phenotypic parameters, and develop a selection index in Atlantic salmon for important traits such as carcass weight, cold tolerance, fillet color, fat content, and sea lice resistance; 2) evaluate and validate the usefulness of incorporating genomic information into a salmon breeding program; and 3) establish links between disease susceptible and resistant phenotypes and genotype for the Eastern Oyster. Research accomplished during this project will result in the development of genetically improved Atlantic salmon for release to U.S. producers and consumers. Identification of genes associated with oyster disease will provide markers that can be used to enhance and accelerate the development of high-performing oyster lines through selective breeding and will support the East Coast shellfish aquaculture industry.
Pedigreed families were produced by spawning broodstock selected for improved carcass weight from 2013-2014 year class (YC13-14) salmon in the National Cold Water Marine Aquaculture Center (NCWMAC) salmon breeding program. Fish from YC14-15 were cultured in marine net pens in collaboration with industry, and growth, fillet color, and fillet fat data will be analyzed to obtain estimated breeding values on broodfish to be spawned as a line selected for increased carcass weight, fillet color, and omega-3 fatty acids. The long-term goal of the sea lice project is to utilize both phenotypic and genotypic metrics of sea lice resistance to improve the overall resistance to sea lice infection of the Atlantic salmon strains being propagated at the NCWMAC. The primary focus over the past year has been the evaluation and screening of the YC15-16 families for both phenotypic resistance to sea lice infection and collection of tissue samples for analysis once the revised SNP chip work is completed. To this end, 1366 fish from 94 different YC15-16 families were challenged with sea lice. Samples were also sequenced from a total of 88 fish selected from 6 populations maintained at the NCWMAC. These sequences are currently in the pipeline for SNP discovery to create a 30-60k North American salmon SNP chip. Crude lipid and fatty acid analysis are being conducted on the fish used for harvest evaluation. The data demonstrated that the largest fish had the highest fillet color scores because fillet color is a function of feed intake and time. We also found that more of the carotenoid, astaxanthin, is deposited in fillets of fish that consume more feed. Omega-3 fatty acid concentrations, Eicosapentaenoic acid (EPA) and Docosahexaenoic acid (DHA) are being evaluated for the current year class. Fish that have higher monounsaturated fatty acid levels have lower long chain Omega-3 fatty acids. This data will be used to determine if fatty acid profiles and color can be added as traits to the breeding program. Dermo disease is one of the greatest threats to production of the eastern oyster. We characterized eastern oyster phenotypes resistant to Dermo disease and identified potential mechanisms contributing to resistance. Three oyster families developed at the Aquaculture Genetics and Breeding Technology Center were exposed to the Dermo-causing parasite in the laboratory and monitored for survival and changes in parasite density. Oysters from the same three families were subsequently subjected to feeding experiments in the presence and absence of the parasite to determine whether behavioral avoidance contributes to the observed variation in resistance. Results suggested that reduced feeding behavior is a component of the Dermo-resistant phenotype in eastern oysters. These findings have important implications for oyster breeding strategies and industry practices. Research to define resistant and susceptible phenotypes associated with Dermo disease in the eastern oyster continued. Sequenced reads from resistant and susceptible oysters exposed to the Dermo-causing parasite in 2015 and 2016 were mapped to the newly released eastern oyster reference genome and differentially expressed genes (DEG) between exposed and control treatments at 36h, 7d, and 28d post-exposure were detected using multiple statistical algorithms. Across analytical methods, Dermo-resistant and -susceptible families exhibited distinct transcriptome responses to the parasite. In resistant families, transcriptomic response to the parasite was immediate and DEG were associated with negative regulation of peptidase activity and oxidation reduction processes. By 28d post exposure, no DEG were observed between exposed and control oysters in the resistant families. In contrast, susceptible families did not respond to the parasite until 28d post exposure and the vast majority of DEG were associated with downregulation of metabolic processes. Only one DEG overlapped between resistant and susceptible groups across all time points. Potentially informative SNPs identified in DEG were tested for their utility in a Fluidigm genotyping assay. High Resolution Melting (HRM) genotyping assays were designed for 104 SNPs and individuals from resistant and susceptible families as well as wild and selected lines were screened with each assay. In total, only 15% (21) of the SNPs tested generated suitable melting curves and reliable genotypes. Thirty-seven assays yielded poor amplification, 12 were monomorphic, and more than three distinct genotypes were detected in 34. Subsequent sequencing of PCR products with more than three genotypes revealed the presence of additional SNPs in the amplicon sequence. Thus, the high polymorphism rate across the eastern oyster genome precluded the development of a high throughput genotyping assay. To circumvent the challenges encountered with the current approach to SNP marker development, the ARS shellfish genetics lab collaborated with the eastern oyster genome consortium to conduct whole genome resequencing of 92 oysters collected from four geographic regions and two ecotypes within each region across the species range; selected oyster lines were also re-sequenced at 30X coverage. SNP variants are being called from the resequencing dataset and the new plan is to develop a high-density (56K or 200K) SNP array for genotyping laboratory and field challenged oysters. NCWMAC’s optimized Dermo challenge protocols are again being applied in an experiment to further measure the extent of variation in Dermo resistance among selectively-bred oyster families and to tease apart resistance and tolerance traits. Twelve families (n = 250 oysters per family) from the Aquaculture Genetics and Breeding Technology Center’s (ABC VIMS) oyster breeding program are included in this challenge experiment. The challenge will run through August 2018.
1. Sea lice resistance in Atlantic salmon. Sea lice are the most economically damaging pest to the global and U.S. salmon farming industry. Sea lice are exhibiting resistance to most of the drugs used globally to manage this parasite. ARS researchers at the National Cold Water Marine Aquaculture Center in Orono, Maine, have combined selection for resistance to sea lice with selection for improved growth in their St. John River strain of Atlantic salmon. Eggs from the improved strain have been provided to industry stakeholders for integration and propagation on commercial farms. The release of improved genetics for resistance to sea lice represent a new management option for domestic salmon growers in controlling this pest.
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