Location: Dairy Forage Research2012 Annual Report
1a. Objectives (from AD-416):
Current ARS, and ARS partners’, efforts are directed at advancing aquaculture in the Great Lakes region. The Great Lakes region is home to approximately 32% of the U.S. population but produces less than 3% of the total U.S. aquaculture production. The Great Lakes region consumes 1.5 billion pounds of seafood annually (valued at $23 billion, 2004 dollars). Yellow perch are a high-value food fish ($16.00-$22.00/lb retail) and current ARS (and ARS partners) efforts are aimed at developing improved yellow perch broodstocks, and rearing/production practices, to advance aquaculture in this region. The emergence of viral hemorrhagic septicemia virus (VHSv) in the Great Lakes region of North America has led to multi-species epidemics in wild populations. This disease poses a serious threat to aquaculture production and profitability in this region and thus is the impetus for the scientific efforts of the ARS program. To address the needs of the Great Lakes region, we shall undertake the following objectives: 1) Continue genotype x environment trials on F3 progeny to evaluate growth and surival in commercial settings, 2) Identify F3 selects from on-site performance trial, PIT tag and photoperiod and temperature cycle to induce maturation, 3) Investigate use of specialty micro-diets and live-diets for production of high-quality perch fingerlings and initiate evaluation of diets for efficient grow-out of perch fingerlings, 4) Complete development of the STaRT-PCR assay for rapid detection of the VHS pathogen, and 5) Characterize innate immune response and pathogen recognition and response pathway(s) to VHSv (and subunits) in fish cell lines.
1b. Approach (from AD-416):
1) Continue genotype x environment (GxE) studies on F3 progeny to evaluate growth and survival in commercial settings (Binkowski, UWM-SFS). Maintain and complete ongoing Genotype x Environment studies, which are essential to evaluating growth and survival of our genetically-improved (F3) progeny at commercial indoor farms. The information gained from the GxE studies will be used to improve genetic gain for growth and identify sources of genetic variation for survival and growth in our broodstock strains. 2) Identify F3 selects from on-site performance trial, PIT tag and photoperiod and temperature cycle to induce maturation (Binkowski, UWM-SFS). Select animals from the F3 generation will be identified, individually tagged, genotyped and photoperiod/temperature cycled to bring these animals into spawning condition to enable future crosses and development/testing of early life-history and grow-out diets. 3) Investigate use of specialty-micro-diets (SMD) and live-diets for production of high-quality perch fingerlings and initiate evaluation of diets for efficient grow-out of perch fingerlings (Binkowski, UWM-SFS). The early life-history dietary work will focus on the use of commercially-available larval diets to improve yellow perch larval and fingerling survival. The aim is to identify a substitution to live diets, which will have a direct impact on continued broodstock improvement efforts and to the yellow perch aquaculture industry. This work will be extended to a production setting by evaluating dietary nutrient requirements needed to sustain efficient grow-out of high-quality fingerlings. This work will also support activities of a new ARS post-doc who will be conducting nutritional research on yellow perch. 4) Complete development of the STaRT-PCR assay for rapid detection of the VHS pathogen (Stepien, UT; Willey, UT and Leaman, UT). Continued work is essential to completion of the STaRT-PCR assay. The final phases of this work are focused on validating platform specificity for the VHSv pathogen. When completed, this platform will be used as a means to verify and validate a TaqMan-based real-time PCR assay format that will make this detection technology more widely available and cost-effective. Cooperators will also disclose this technology to APHIS/USDA in an effort to obtain further direction/input that will enable this testing platform to meet requirements for Federal approval. 5) Characterize how VHSv structural and non-structural genes evade the host innate immune capacity to detect and respond to the VHS pathogen (Leaman, UT and Stepien, UT). In vitro studies, using newly developed reagents and fish cell-lines, will be aimed at understanding the molecular pathways involved with the viral recognition and response pathways in fish. Additionally, this work will result in a better understanding of how various VHSv genes influence virulence and pathogenicity in these cell-lines. This work is critical to developing new means to detect the pathogen and in identifying novel therapeutic approaches to interfere with the pathogen lifecycle as a means to prevent and/or treat this disease.
3. Progress Report:
This project is related to the following objectives of the parent project: Objective 1) Develop yellow perch brood-stock, define growth and VHS resistant phenotypes, characterize genetic diversity, and evaluate genotype x environment interaction for growth; Objective 2) Characterize critical pathways involved in growth and VHSv resistance in yellow perch through gene expression and physiological studies; Objective 3) Improve early survival and methods for producing feed trained fingerlings; Objective 4) Develop and evaluate challenge assays, detection tools and vaccines for protecting yellow perch and other Great Lakes region species of fish from VHS. As part of an ongoing research effort to increase yellow perch fingerling production cost-effectively, we repeated an early life stage husbandry trial in January 2012, using an egg ribbon density of nine egg strands/tank. Fingerling production was 57,500 per tank and survival was 28%, which represents a 40% increase over previous trials. Confirmation of this finding results in a direct increase in reproductive efficiency and availability of fingerlings for the aquaculture industry. To expand the use of yellow perch produced under intensive aquaculture conditions, we are evaluating yellow perch performance within integrated urban aquaponic systems. This represents a divergence away from large-scale commercial recirculating aquaculture systems. We are evaluating the growth and survival of yellow perch fingerlings in urban aquaponics systems in cooperation with Growing Power and Sweet Water Organics, both located in Milwaukee, WI. To further develop genetically-improved, faster-growing, yellow perch broodstocks, third generation (F3) progeny were produced from 45 pairwise crosses of Choptank and Perquimans River parental (F2) strains. Performance of the F3 progeny was evaluated on-site and off-site for growth, survival, and susceptibility to the viral hemorrhagic septicemia virus (VHSv). In the spring of 2012, the top ~37% performers (largest) of the F3 generation for each of our two strains (Perquimans and Choptank) were identified following a 1-year on-site performance trial and individually tagged. These fish are presently being photothermally cycled to induce gonadal maturation for spawning in 2013. Genotype x environment studies to evaluate performance of F3 progeny were completed at two commercial recirculating aquaculture producers in the Midwest U.S. Genetic material and data on growth and survival have been collected. Disease challenge trials were conducted on F3 progeny to evaluate survival following a standardized exposure to the VHSv pathogen. Results show strain-specific differences in survival to the VHSv pathogen. Studies using fish cell lines, that stably express major VHSv type IVb proteins, have shown that the M and V genes negatively influence fish cell function, allowing the VHSv pathogen to survive and replicate within the host cell. By contrast, expression of another VHSv gene was found to stimulate the host (cell) innate immune response, which suggests that this gene/protein may be used as a vaccine adjuvant to boost the immune response to vaccination. The standardized reverse transcriptase polymerase chain reaction (StaRT-PCR) method for detecting the VHSv pathogen in aquaculture species was developed. The StaRT-PCR assay is specific to the VHSv pathogen, is rapid (only takes days to accomplish versus several weeks) and displays the highest known detection range and accuracy. The higher accuracy results in significantly lower false positive and false negative rates seen with other methods.