2006 Annual Report 1.What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? Why does it matter? This project is aligned with National Program 106, Aquaculture. With the world population steadily increasing, agricultural research is playing a progressively more critical role in meeting the rising demand for food and at the same time attempting to improve the nutritional value of that food. As the world’s need for renewable sources of protein becomes more important, with declining space and reduced wild animal populations, it will be necessary to raise agricultural livestock in limited space, on readily renewable and relatively inexpensive food sources, and in an ecologically sound manner. If commercial aquaculture producers fail to develop alternative food sources they will be unable to meet future consumer demands and their products will most likely be relegated to a secondary food source and hence, unfeasible as a protein resource for most of the world’s economically underprivileged population. This lack of commercial development could also lead to the loss of billions of dollars in lost revenue, employment opportunities, and exportation products. In addition to presenting the world with a renewable and healthy food source, the development of fish feeds containing cereal grains will provide a sustainable market for cereal grain farmers. Barley and oats represent important commodities for feed, food and malt use in the United States. The continued viability of U.S. industries that depend on these crops will depend on continuing improvements in the efficiency with which they can be grown and used. One goal of this project is to change the type of ingredients used in aquaculture diets from marine sources to sustainable plant sources (particularly small grains). Requirements for plant-derived aquatic feed ingredients are high protein, favorable amino acid levels, decreased phytate levels, increased energy from oils and carbohydrates, presence of omega-3 fatty acids and the improvement of palatability by lowering fiber and antinutritional components. Grain and grain by-products have been components of prepared feeds for fish for over 70 years. Wheat middlings were one of the first feed ingredients used to extend meat mixtures used to rear trout, and remains a key component today. Corn is the main grain component in catfish feeds, primarily because of its geographical proximity to areas where catfish farming is located. Other small grains have been considered as components of aquaculture feeds, but concerns about crude fiber and phytate levels in these grains have limited their use. Recently, genetically enhanced cultivars of barley containing lower levels of phytic acid have been evaluated as components of fish feeds, and found to be superior to normal cultivars in nutrient bioavailability. Development of hulless cultivars of barley and oats further open the prospect of higher utilization of these grains in fish feeds. Historically cultivars of barley and oats were chosen for cultivation on the basis of their agronomic performance and value for livestock feed or as components of human foods. However, there are other germplasm sources available that possess characteristics of interest with respect to fish production. In order to capitalize on the benefits of these genetically diverse cultivars they must first be evaluated for their use in fish feeds. Researchers at the National Small Grains and Potato Germplasm Research Facility are currently screening their vast repository of genetically diverse small grain cultivars for strains that possess traits that might accentuate their value in formulated fish diets. Not only is it necessary to change the diets of commercially produced fish, but it is also necessary to change the fish such that they can efficiently utilize these new plant-based feeds. During the history of aquaculture there has been limited genetic improvement in aquaculture stock. In order for aquaculture to expand in the manner necessary to meet future consumer demands we must follow what has been done historically with other domesticated agricultural livestock. Two of the main elements necessary to increase availability of animal products are the improvement in animal health and nutrition and the means to produce it in a cost efficient manner. In the advancement of all domesticated stock, genetics has played a major role in meeting the two prior mentioned demands. Genetically enhanced stocks are more resistant to disease, reach market size more rapidly, and demonstrate an improved feed conversion ratio. For aquaculture to continue its growth, genetically enhanced domesticated stocks of fish will need to be generated that are specific for the consumption of the most cost efficient and readily available diet. A successful conclusion to this research will enable the total replacement of fishmeal in trout feeds with sustainable plant-derived ingredients, while maintaining optimal production efficiencies 2.List by year the currently approved milestones (indicators of research progress) Year 1 (FY2006) 1. Test effect of replacing fish meal and fish oil in formulated diets on spawning fish. 2. Characterize changes in expression of metabolic genes in fish fed plant based diets. 3. Select for growth and nutrient utilization in 2nd generation of rainbow trout stock (even year generation) reared on formulated plant based feeds. Year 2 (FY 2007) 1. Genetic analysis on the different families of fish to characterize marker differences related to dietary growth effects. 2. Evaluation of the effects of different methods of ingredient processing for incorporation of plant based material into fish feeds. 3. Select for growth and nutrient utilization in 3rd generation of rainbow trout stock (odd year generation) reared on formulated plant based feeds. Year 3 (FY 2008) 1. Identify molecular markers for traits linked to specific aspects of growth for rainbow trout reared on cereal grain diets. 2. Evaluate newly developed processing methods on different types of plant material in fish feeds. 3. Select for growth and nutrient utilization in 3rd generation of rainbow trout stock (odd year generation) reared on formulated plant based feeds. Year 4 (FY 2009) 1. Determine the potential of biological modification of ingredients, use of microbial or fungal incubation to increase the potential of plant material for incorporation into fish feed. 2. Identify the requirement for specific vitamin inclusion in plant-based feeds. 3. Select for growth and nutrient utilization in 4th generation of rainbow trout stock (odd year generation) reared on formulated plant based feeds. Year 5 (FY 2010) 1. Identify the ideal amino acid profile for alternative fish meal and fish oil fish diets. 2. Performance of nutrient digestibility studies for established diets containing different plant materials, biologically modified plant products, and differentially processed plant material containing established amino acid profiles and identified vitamin inclusion levels. 3. Select for growth and nutrient utilization in 4th generation of rainbow trout stock (even year generation) reared on optimized formulated plant based feeds. 4. Determine formulation for a fish feed with fish meal and fish oil replaced with alternative sources and contains an optimal amino acid profile and vitamin level for achieving equal or enhanced growth levels and carcass processing characteristics in rainbow trout. 5. Farm scale testing of progeny generated from marker selected broodstock for growth on plant based diets on optimized fish meal and fish oil replaced diet. 4a.List the single most significant research accomplishment during FY 2006. Taurine Supplementation of Plant-based Trout feeds. Growth of carnivorous fish fed plant-based diets is usually less than fish fed fish meal based diets, even when all known nutrient requirements are met. A study was conducted that identified the sulfur-containing amino acid taurine to be conditionally indispensable to trout when fed plant-based feeds. This amino acid is abundant in fish meal, but virtually absent in plant-derived ingredients. Supplementation of this amino acid to plant-based feeds increased growth of trout to levels equivalent to trout fed fish-meal based feeds. Addition of this commercially available synthetic amino acid to plant-based trout feeds represents a viable approach to increasing growth rate of rainbow trout fed plant-based diets. These studies address the NP106 components growth, development and nutrition and sustainability and environmental compatibility of aquaculture and were conducted at the Hagerman Fish culture Experiment Station in collaboration with the University of Idaho. 4b.List other significant research accomplishment(s), if any. Genetic Markers for Metabolism in Rainbow Trout Understanding the genetic components in trout involved in diet utilization is critical for US aquaculture industry to remain competitive particularly in the face of higher feed costs, limited protein resources, and stricter environmental standards. Differential expression relating growth, health and metabolism of trout fed diets based on fish meal or containing high levels of barley were studied. Rainbow trout of distinct lineage were separated and reared on either of these two diets for one year after which, tissue samples were taken and the changes in gene expression were evaluated through analysis using quantitative real-time PCR for several genes and microarray hybridization. The findings from this work will now allow researchers more efficient means to evaluate selected broodstock for improved performance and aid in the evaluation of formulated diets. These studies address the NP106 components of genetic improvement, growth, development and nutrition, and sustainability and environmental compatibility of aquaculture and were conducted at the Hagerman Fish culture Experiment Station in collaboration with the University of Idaho. Fractionization and Characterization of Barley Many nutrients are concentrated in the outer layers of grains. A major problem that hinders cereal research is lack of simple and small-scale methods in decorticating grains. We have developed a lab-scale method to meet this need. Using this newly developed grain pearling method, we have been able to study effects of low phytic acid (LPA) mutation on contents and distribution patterns for different types of phosphorous and minerals within a barley seed. We observed that there is no difference in mineral concentrations and distribution patterns between conventional barley variety and 4 LPA mutants. This implies that there is no direct role of localization of phytic acid synthesis in mineral distribution within a barley seed and that LPA breeding does not lead to reduction in mineral contents in barley seeds. The studies address the NP106 components of growth, development and nutrition, and sustainability and environmental compatibility of aquaculture. New Approach for DNA Marker Development in Oat The SSR (Simple Sequence Repeat) as new generation of DNA markers is essential molecular tool for genetic studies of nutritional traits and efficient selection of desired traits in crop breeding. Oat, an important potential resource of plant-based ingredients for fish diet, lacks of this type of markers because of limited genomic database. To search for alternatives of SSR sources, we systemically surveyed SSR markers from closely related species of oat. Results confirmed that abundant SSRs from barley and wheat genomes are good alternatives for oat. Discovery from this study will greatly contribute to the development of oat SSR markers and genetic researches related to improvement of nutritional traits in oat. The studies address the NP106 components of growth, development and nutrition, and sustainability and environmental compatibility of aquaculture. Biological Enhancement of Soybean/Barley mixture for Rainbow Trout. To be an effective replacement for fish meal in trout diets, soybeans and barley contain too little protein and too many anti-nutrients or poorly digestible nutrients. To increase the value of these ingredients for trout a fermentation using selected, human food grade, fungal strains was conducted. Products contained high levels of protein (over 60%) and low levels of carbohydrate, and no phytic acid was detectable after fermentation, and the digestibility of protein, energy, dry matter, organic matter and phosphorus were greatly improved. Unfortunately, feeding studies indicated poor palatability limiting inclusion rate of this ingredient into the formulation. This project is being conducted in collaboration with Montana Microbial Products through a CRDA and in collaboration with the U.S. Fish and Wildlife Service and meets the NP106 components of growth, development and nutrition, and sustainability and environmental compatibility of aquaculture. Successful development of these products will provide the fish feed manufacturing industry with another ingredient that meets the criteria for nutrient availability, sustainability, environmental compatibility and cost effectiveness. 4c.List significant activities that support special target populations. None. 4d.Progress report. None. 5.Describe the major accomplishments to date and their predicted or actual impact. This project is aligned with National Program 106, Aquaculture. The Action Plan identifies the need to determine the bioavailability of nutrients in traditional and novel feed ingredients and to develop complete feeds based on this data. The Action Plan also identifies a goal to “Identify phenotypic and molecular differences between stocks and species, which will be used to characterize and develop markers for unique traits or characteristics identified or developed within the U.S.” This project is a collaboration between the Small Grains and Potato Germplasm Facility in Aberdeen, Idaho and the Hagerman Fish Culture Experiment Station, University of Idaho. With this being a new and unique area of research within the Agricultural Research Service, there has been rapid progress in this novel area of looking to enhance cereal grains for trout diets and in improving trout strains for growth and the utilization of these new diets. During the span of this project we plan to develop cereal grains with accentuated positive traits for incorporation into fish feeds. Traits such as reduced phytic acid will aid in reducing phosphorus excretion into the environment, while increasing oil and protein levels and inserting them into cultivars possessing strong agronomic traits will aid in the production and marketing of a finished product. We also plan to have delineated a number of molecular markers that are closely linked to genes that allow rainbow trout to grow well and efficiently utilize a fish diet containing cereal grains in place of a large portion of the typically used fish meal and fish oil. We are also identifying compounds that improve growth and feed efficiency that are present in fish meal but not in plant-products, developing improved processing technology for ingredients and complete feeds, and reevaluating vitamin and trace mineral supplements for plant based feeds. Development of enhanced stocks of cereal grains for utilization in fish foods, formulated cereal grain fish diets, and improved rainbow trout stocks should have a major effect in the future production of food finfish in regards to volume and price of fish produced and its impact on the environment. 6.What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end-user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? Individual families generated from our broodstock selection program have been transferred to an industry cooperator through a CRADA and are being incorporated into their production broodstock. 7.List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: List your peer reviewed publications below). Barrows, F.T. 2006, Fish Meal Replacement; effect of alternate feed ingredients and nutrient density on growth efficiency of rainbow trout Oncorhynchus mykiss”. Book of Abstracts, Soy in Aquaculture Special Symposium; World Aquaculture Society Annual meeting, Las Vegas, NV. Log Number: Barrows, F.T. 2006. Recent results with a plant-based trout feed and review of work on novel protein sources for trout. Book of Abstracts, U.S. Trout Farmers Meeting, World Aquaculture Society Annual meeting, Las Vegas, NV. Log Number: Sealey, W.M., Barrows, F.T., Hang, A., Johansen, K.A., Overturf, K.E., LaPatra, S., and Hardy, R.W. 2006. Evaluation of the ability of barley varities containing different amounts of B-glucan to alter growth, immune function, and disease resistance of rainbow trout (Onchorynchus mykiss) . Book of Abstracts, World Aquaculture Society Annual meeting, Las Vegas, NV. Log Number: 0000190738 Gaylord, T.G., Teague, A.M., and Barrows, F.T. 2006. Taurine supplementation of all-plant protein diets for rainbow trout (Onchorynchus mykiss). Book of Abstracts, World Aquaculture Society Annual meeting, Las Vegas, NV. Log Number: 0000187041 Snyder, S.G., Gaylord, T.G., Teaque, A.M., and Barrows, F.T. 2006. Apparent nutrient digestibilities of alternative ingredients in extruded rainbow trout (Onchorynchus mykiss) diet formulations. Book of Abstracts, World Aquaculture Society Annual meeting, Las Vegas, NV. Log Number: 0000187042 Liu, K., and Bonman, J.M. 2006 Small grains: Old crops with new uses. Book of Abstracts, Annual meeting of the American Oil Chemists’ Society. Log Number: 0000191317 Rawles, S.D., Snyder, S.G., Gaylord, T.G., Mcentrire, M.E., and Freeman, D.W. 2006. The influence of dietary protein and energy on the performance of hybrid striped bass reared at extreme temperature. Book of Abstracts, World Aquaculture Society Annual meeting, Las Vegas, NV. Log Number: 0000187146 Rawles, S.D., Richie, M.A., Webb, J., Gaylord, T.G., Freeman, D.W., Davis, M. 2006. Evaluation of Poultry by-product meal in commercial diets for hybrid striped bass in recirculated tank production. Book of Abstracts, World Aquaculture Society Annual meeting, Las Vegas, NV. Log Number: 0000187141 Rawles, S.D., Richie, M.A., Webb, J., Gaylord, T.G., Freeman, D.W., Davis, M. 2005. Evaluation of Poultry by-product meal in commercial diets for hybrid striped bass in recirculated tank production. Book of Abstracts, International Sustainable marine Fish Culture Conference and Workshop.. Log Number: 0000187141 Review Publications Stone, D.A., Hardy, R.W., Barrows, F. 2005. Effects of extrusion on nutritional value of diets containing corn gluten meal and corn distiller¿s dried grain for rainbow trout, onchorhynchus myskiss.. Journal of Applied Aquaculture. 17(3):1-20. Overturf, K.E., Lapatra, S. 2006. Quantitative expression of immunological factors in rainbow trout (oncorhynchus mykiss) after infection with either flavobacterium psychrophilum, aeromonas salmonicida, or infectious hematopietic necrosis virus (ihnv). Journal of Fish Diseases. 29:215-224. Hu, G., Jackson, E.W., Bonman, J.M. 2006. Enlargement of pcr-based marker resources in oat by surveying genomic-derived ssrs from barley and wheat. American Oat Workers Conference Proceedings. 52 Barrows, F., Lellis, W.A. 2006. Effect of diet processing method and ingredient substitution on feed characteristics and survival of larval walleye (stizostedion vitreum) journal equivalent: journal of the world aquaculture society.. Journal of the World Aquaculture Society. 37(2):154-160. Overturf, K.E., and Johansen, K.A. 2006 Alterations in expression of genes associated with muscle metabolism and growth during nutritional restriction and refeeding in rainbow trout. Comparative Biochemistry and Physiology. 144:119-127. Rawles, S.D., Riche, M.A., Gaylord, T.G., Webb, J., Freeman, D.W., Davis, M. 2006. Evaluation of poultry by-product meal in commercial diets for hybrid striped bass (Morone chrysops x M. saxatilis) in recirculated tank production. Aquaculture. 259:377-389.