2012 Annual Report
1a.Objectives (from AD-416):
1. Identify genes affecting variation in production traits in rainbow trout through QTL mapping and functional genomic (i.e. expression based) approaches.
• 1.a. Detect and fine map quantitative trait loci for resistance to bacterial cold water disease in rainbow trout.
• 1.b. Fine map quantitative trait loci affecting response to crowding stress.
• 1.c. Identify genes affecting response to crowding stress through functional genomic approaches.
• 1.d. Evaluate performance of fish differing in stress response phenotypes.
• 1.e. Identify genes affecting carcass quality traits in rainbow trout.
• 1.f. Identify and characterize key oocyte-expressed genes/microRNAs important for folliculogenesis and early development.
2: Develop genomic tools and resources to facilitate the use of state of the art approaches for genetic improvement of rainbow trout.
• 2.a. Facilitate the identification of genes affecting production traits by producing a second generation bacterial artificial chromosome (BAC) map which is anchored to the genetic map.
• 2.b. Develop single nucleotide polymorphic (SNP) markers to enhance fine mapping and enable genomic selection for rainbow trout.
• 2.c. Identification of microRNAs that affect expression of genes controlling production traits in rainbow trout.
3: Develop database to store, and facilitate analysis of genotypic and phenotypic data.
1b.Approach (from AD-416):
The demand for seafood is increasing worldwide while captured fisheries harvest is limited and unsustainable. To meet increasing consumer demand, U.S. aquaculture producers have to achieve improved efficiencies and sustainable practices while maintaining and improving product quality. The application of genomic technologies towards the genetic improvement of aquaculture species is expected to facilitate selective breeding and provide basic information on the biochemical mechanisms controlling traits of interest. In collaboration with U.S. and international scientists, we have developed a suite of genome tools and reagents for rainbow trout to identify and characterize genes affecting aquaculture production traits. Projects concurrent with our previous 5-year project characterized the genetic variation of the National Center for Cool and Cold Water Aquaculture (NCCCWA) broodstock with respect to resistance to Bacterial Cold Water Disease (BCWD) and response to crowding stress. Specific crosses were identified to facilitate the identification of genes affecting these traits through genetic mapping and functional genomic approaches. The current project will improve and utilize genome mapping approaches to identify positional candidate genes affecting these traits. This genetic information will be used for improving our understanding of the genetics of disease resistance and production traits and could be transferred to the US industry through improved germplasm. In addition, possibilities for developing informative crosses and functional genomic approaches which target the identification of genes affecting carcass quality traits will be determined. We will also continue to identify and characterize genes in the oocyte (pre-mature egg) which impact embryonic development and egg quality traits important to breeders.
Progress towards Objective 1 included efforts to identify genes affecting stress response in rainbow trout. Fish response to stress is an important factor in aquaculture production, having significant negative impacts on growth, feed efficiency, immune response, and reproductive characteristics. A total of 316 genes were determined to respond to crowding stress in the liver, 53 of which are linked to specific metabolic pathways.
Also associated with Objective 1 were efforts to identify genes affecting resistance to Bacterial cold water disease (BCWD) which is caused by the bacterium Flavobacterium psychrophilum, a major concern for rainbow trout aquaculture production and managers of recreational fisheries. Genetic markers representing the entire rainbow trout genome were analyzed on a three generation pedigree which revealed a single chromosome region with large effect on this trait. New crosses were made between this population and a commercial populations to further evaluate and validate the effect of this region on disease resistance.
Progress towards Objective 2 includes the development of new genetic markers systems for rainbow trout. A relative new technology termed “Restriction-site associated DNA” (RAD) was used to identify a set of 150,000 single nucleotide polymorphisms (SNPs), a type of genetic marker defined as single base pair differences in DNA sequences between individuals.
Under Objective 3 the NCCCWA database was modified to analyze high volumes of DNA and/or RNA sequence data associated with state of the art gene expression and mapping technologies. Also, the database was updated with fish pedigree and performance information associated with stress response and disease resistance.
Genetic mapping of handling stress response in rainbow trout. Fish response to stress is an important factor in aquaculture production, having impacts on growth, feed efficiency, immune response, and reproductive characteristics. Efforts to identify the genes responsible for these effects include measurements of the hormone cortisol in the blood as fish respond to a crowded environment. Through genetic mapping approaches ARS researchers at the National Center for Cool and Cold Water Aquaculture (NCCCWA) at Leetown, West Virginia identified regions of 8 chromosomes harboring genes affecting this trait in NCCCWA broodstock and therefore quantifying sources of genetic variation for this trait within and between populations that may independently affect overall performance.
Generation of a new resource for analyzing stress response in rainbow trout. Identifying and understanding the genes affecting stress response will facilitate the development of breeding schemes and management practices aimed at mitigating negative effects on production traits. To this end RNA samples from gill, brain, liver, spleen, kidney and muscle representing stressors common to aquaculture production environments (high and low temperatures, high salinity, crowding, low dissolved oxygen/high carbon dioxide) were sequenced by ARS researchers at the National Center for Cool and Cold Water Aquaculture at Leetown, West Virginia to produce 3,160,306 gene sequences known as a “transcriptome.“ This resource significantly increases the amount of publicly available rainbow trout gene sequence data and is specifically designed to aid in the identification of metabolic pathways and genes involved in stress responses in rainbow trout.
Miller, M.R., Brunelli, J.P., Wheeler, P.A., Liu, S., Rexroad III, C.E., Palti, Y., Doe, C.Q., Thorgaard, G.H. 2012. Next-generation sequencing reveals a conserved haplotype controlling parallel adaptation in geographically distant rainbow trout (Oncorhynchus mykiss) populations. Molecular Ecology. 21(2):237-249.
Sanchez, C., Weber, G.M., Gao, G., Cleveland, B.M., Yao, J., Rexroad III, C.E. 2011. Generation of a reference transcriptome for evaluating rainbow trout responses to various stressors. Biomed Central (BMC) Genomics. 12:626.
Palti, Y., Genet, C., Gao, G., Hu, Y., You, F., Boussaha, M., Rexroad III, C.E., Luo, M. 2012. A second generation integrated map of the rainbow trout (Oncorhynchus mykiss) genome: analysis of synteny with model fish genomes. Marine Biotechnology. 14:343-357.
Salem, M., Vallejo, R.L., Leeds, T.D., Palti, Y., Liu, S., Sabbagh, A., Rexroad III, C.E., Yao, J. 2012. RNA-Seq identifies SNP markers for growth traits in rainbow trout. PLoS One. 7(5):e36264.
Palti, Y. 2012. Toll-like receptors in bony fish: from genomics to function. Developmental and Comparative Immunology. 35(12):1263-1272.
Overturf, K.E., Vallejo, R.L., Palti, Y., Barrows, F., Parson, J. 2012. Microarray analysis of differential utilization of plant-based diets by rainbow trout. Aquaculture International. 20:213–232.
Ma, H., Hostuttler, M.A., Wei, H., Rexroad III, C.E., Yao, J. 2012. Characterization of the rainbow trout oocyte microRNA transcriptome. PLoS One. 7(6):1-8.
Liu, S., Rexroad III, C.E., Couch, C.R., Cordes, J.R., Reece, K.S., Sullivan, C.V. 2011. A microsatellite linkage map of striped bass (Morone saxatilis) reveals conserved synteny with the hree-spined stickleback (Gasterosteus aculeatus). Marine Biotechnology. 14(2):237-244.
Burki, R., Krasnov, A., Bettage, K., Rexroad III, C.E., Afanasyev, S., Antikainen, M., Burkhardt-Holm, P., Wahli, T., Segner, H. 2012. Molecular crosstalk between a chemical and a biological stressor and consequences on disease manifestation in rainbow trout. Aquatic Toxicology. 127:2-8.
Burki, R., Krasnov, A., Bettage, K., Rexroad III, C.E., Afanasyev, S., Antikainen, M., Burkhardt-Holm, P., Wahli, T., Segner, H. 2012. Pathogenic infection confounds induction of the estrogenic biomarker vitellogenin in rainbow trout. Journal of Society of Environmental Toxicology and Chemistry. 31(10):2318-2323.