2005 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? What does it matter? The specific problem area is to improve economically important traits of channel catfish through an applied breeding program that incorporates new biotechnologies and addresses all areas of quantitative and qualitative genetics, reproduction, and molecular and cellular genetics. Genetically improved germplasm will be developed, evaluated, and released to commercial producers. Research areas have been identified through research planning sessions with industry, state and federal research, and cooperative extension representatives. All research focuses on problems of critical importance to the catfish industry that can be solved through genetic improvement. The applied breeding program for catfish genetic improvement focuses on identifying economically important traits, determining the amount and sources of phenotypic and genetic variation for those traits, determining the best approaches to improve the traits through genetic selection, and incorporating molecular genetic and specific breeding aid technologies into the applied breeding program. The project’s overall objective is to develop a breeding program to improve catfish production. Specific objectives are:. 1)characterize variation for important traits;. 2)develop molecular markers and a genetic map, and incorporate molecular markers into the breeding program;. 3)conduct research to develop management protocols for optimizing growth, production and fish health of genetically improved catfish lines released to commercial industry; and. 4)release genetically improved germplasm to commercial producers. The planned research falls under National Program 106, Aquaculture. It addresses the Genetic Improvement, Integrated Aquatic Animal Health Management, and Growth and Development, and Nutrition components of the Action Plan. There has been limited genetic improvement of aquaculture stocks so there are major opportunities for improvement through traditional animal breeding, broodstock development, germplasm preservation, molecular genetics, and allied technologies. Commercial catfish farmers, like all aquaculture producers, essentially utilize fish stocks that are not many generations removed from wild, unselected stocks. Research will address improvement of growth rates, feed efficiency, survival, disease resistance, fecundity, fillet yield and product quality; environmental tolerances; genetic characterization and gene mapping; and conservation and utilization of important aquatic germplasm. Development of catfish lines or germplasm with superior performance for commercially important traits and utilization of these lines in commercial culture will help solve production problems, increase efficiency and profitability, and provide a quality product for consumers. 2.List the milestones (indicators of progress) from your Project Plan. Objective 1: Measure genetic variation in production traits in the USDA103 line and characterize correlations between traits. Sub-objective 1.1: Measure growth and carcass yield in catfish after rearing in ponds. -Determine growth and carcass yield breeding values for Generation 4 (G4) fish. -Determine growth and carcass yield breeding values for Generation 5 (G5) fish. Sub-objective 1.2: Correlate levels of growth hormone, insulin-like growth factors I and II (IGF-I, IGF-II) and IGF binding proteins (IGFBPs) with growth efficiency in catfish families. -Develop a fluoroimmunoassay for IGF2 and real-time PCR assay for IGF binding proteins. -Calculate correlations for growth efficiency and IGF molecular data. Sub-objective 1.3: Quantify family variation in resistance to E. ictaluri in the 4th and 5th generations of the USDA103 line. -Rank Generation 4 families for ESC resistance. -Rank Generation 5 families for ESC resistance. Sub-objective 1.4: Determine levels of gonadotropic hormones in spawning catfish and correlate with individual variation in reproductive efficiency. -Validate GTH-II immunoassay. -Correlate GTH-II response and reproductive success. Objective 2. Develop genomic resources for integrating functional genomics into the catfish applied breeding program. Sub-objective 2.1: Add interspecific conserved genes to the catfish genetic linkage map to facilitate comparative genomic analyses. -Add two hundred expressed sequences to linkage map. -Place additional conserved genes on linkage map. -Develop web-based access to database of molecular markers and linkage map. Sub objective 2.2: Identify chromosomal regions (quantitative trait loci) containing genes controlling carcass yield and resistance to ESC. -Produce F1 carcass yield families. -Produce F1 ESC resistance families. -Produce F2 carcass yield families. -Produce F2 disease resistance families. -Identify QTL’s for carcass yield and ESC resistance. Sub-objective 2.3: Develop molecular markers for disease resistance candidate genes based on comparative functionality with other species. -Obtain real-time PCR results for TLR3, TLR5, lysozyme C, lysozyme G -Identify additional Toll-like receptors. -Correlate molecular data obtained above with ESC resistance. Sub-objective 2.4: Identify candidate genes and gene products controlling economically important traits using differential gene expression on oligonucleotide microarrays. -Produce oligonucleotide microarray. -Identify candidate genes for ESC resistance/susceptibility. -Identify candidate genes for incidence of spawning. Objective 3: Increase biological efficiency through selective breeding of catfish and transfer improved catfish germplasm to the U.S. catfish industry. Sub-objective 3.1: Develop and implement an index based on breeding values of traits and their economic values to select individuals with superior composite phenotypes. -Determine economic values for traits. -Develop individual selection index for G4 broodfish. -Develop individual selection index for G5 broodfish. Sub-objective 3.2: Quantify differences in performance for production traits between selected lines and the USDA103 base population. -Obtain data from comparison of G5 with G2 (NWAC103). -Produce fingerlings for release to industry. 4a.What was the single most significant accomplishment this past year? The USDA103 line of channel catfish was developed and evaluated at the Thad Cochran National Warmwater Aquaculture Center and released under the name NWAC103 to commercial producers in 2001. After two generations of selectively breeding USDA103 channel catfish for rapid growth, a new experimental channel catfish line (USDA303) has been developed. Research was conducted to assess growth improvements in the USDA303 line of channel catfish, and showed a marked improvement in USDA303 channel catfish growth. Two generations of selection for increased growth resulted in a 21% increase in USDA303 channel catfish body weight compared to USDA103 catfish at the end of the study. Continued improvements in growth through selective breeding will lead to more efficient production for U.S. catfish farmers. 4b.List other significant accomplishments, if any. The growth hormone-insulin like growth factor (GH-IGF) system plays an important role in growth of mammals, but its function in catfish is poorly understood. Assays were developed that allowed the measurement of GH, GH receptor (GHR), IGF-I, and IGF-II mRNA as well as an assay to measure protein levels of IGF-I. Research that examined regulatory mechanisms of the GH-IGF system in catfish found that IGF-I and IGF-II may play important roles in growth and that GH and GHR may play important roles in regulating the action of IGF-I. Identifying polymorphisms in GH or IGF genes may have potential for use in DNA marker-assisted selection programs. Insulin-like growth factors-I and –II (IGF-I and IGF-II) are potential regulators of catfish growth and may also play roles in immune function. We measured levels of gene expression for IGF-I, IGF-II, and Toll-like receptors 3 and 5 in developing catfish as well as the role of IGF-I in catfish challenged with virulent Edwardsiella ictaluri. Results showed that growth (IGF-I and IGF-II) and immune (TLR3 and TLR5) associated genes could be functional and play important roles during embryogenesis and early development of catfish and that IGF-I decreases as Edwardsiella ictaluri in the blood increases. Understand mechanisms regulating catfish growth and immune function may lead to the ability to genetically select groups of individuals with improved growth and disease resistant phenotypes. Glucocorticoids are known to impede somatic growth in a number of vertebrate species. In order to better understand the mechanisms through which they may act in channel catfish, we examined the effects of feeding cortisol on the growth hormone (GH)/insulin-like growth factor-I (IGF-I)/IGF-binding protein (IGFBP) system. Similar studies were conducted in collaboration with the Harry K. Dupree Stuttgart National Aquaculture Research Center in palmetto and sunshine bass. The catfish results showed that cortisol administration increased a 20-kDa IGFBP in circulation and decreased plasma IGF-I levels. This research identified a 20 kDa IGFBP as a possible important regulator of growth. One mechanism through which cortisol may impede growth of catfish is through an increase in a low molecular weight IGFBP which may lead to inhibitory effects on the action of IGF-I. Lower molecular IGFBPs may be used as a marker of growth or stress. Molecular genetic markers have the potential to improve the selection for desired traits, such as growth and feed efficiency, in catfish by serving as predictors of genetic merit. Two genes known to regulate growth and feeding in other animals are growth hormone receptor and ghrelin. Research was conducted to identify these genes in catfish, determine their roles in catfish growth, and identify differences in sequence and gene expression patterns. Both genes were identified in catfish, found to be potent regulators of growth hormone, and to play central roles in the regulation of catfish growth. This new insight into the regulation of catfish growth together with observed differences in sequence and expression patterns of the two genes may prove to be useful tools for identifying superior families of catfish. Reproductive efficiency is important for the sustained production of channel catfish; however, the hormonal and genetic regulation of catfish reproduction is not understood. Ongoing research has identified several genes with the potential to regulate catfish reproductive efficiency. One gene in particular, steroidogenic factor-1 (SF-1), was identified in reproductive tissues of mature channel catfish and has the potential to control the production of steroids important for successful reproduction. Identification of factors regulating reproductive development will allow for the genetic selection of superior broodfish and provide information toward the development of tools for spawning induction. Development of ESC resistant line. ESC is the most prevalent disease affecting commercial catfish farms. 100 families (full-sib groups) were screened and ranked for susceptibility to ESC infection (with virulent Edwardsiella ictaluri) in three separate challenge experiments. Survivors from the 10 most resistant families were pit-tagged and released into ponds. In 2 years, these fish will be used for breeding and heritability of resistance to ESC will be measured. This project addresses component 1.3 of the project plan and meets the milestone for ranking G4 families for ESC resistance. This project also addresses component 2.2 and its milestone of completion in yr2 in that the fish that will be used to produce the F1 resistant families have been selected, pit-tagged, and released into the ponds. TLR expression in resistant catfish. Blue x channel catfish hybrids show increased resistance to ESC infection. Lysozyme activity and expression of two toll-like receptor (TLR) genes, which show association with immune response to ESC, were measured in back-cross hybrid (channel x blue/channel) fish. Both lysozyme activity and expression of both TLR genes increased after exposure to virulent Edwardsiella ictaluri, however the patterns of expression differed from what was previously found for channel catfish. This research will improve efforts towards selectively breeding catfish for disease resistance. Detection of CCV carriers with genetic detection assay. Channel catfish virus (CCV) may be transmitted both vertically and horizontally. A quantitative genetic assay was optimized for detection of CCV in asymptomatic fish. Both broodfish and fingerlings from both commercial farms and research populations were determined to be carriers of CCV. The incidence of CCV carriers was on average 10–50% of fish sampled. This information will be used for screening and assessment of CCV susceptibility in future projects. Channel catfish virus (CCV) accounts for approximately 5% of the annual losses to disease on commercial farms. However, localized CCV outbreaks can produce large losses of fingerlings on individual farms. In cooperation with investigators from Mississippi State University, an extensive series of viral challenges was performed to determine the relative susceptibility of various catfish lines to CCV, especially the NWAC103 line that was developed at the Catfish Genetics Research Unit. NWAC103 performance was average for the catfish strains tested, with no significant increase or decrease in susceptibility to CCV compared with other strains. These experiments provided initial data that will be used to examine the genetic control of resistance to CCV in channel catfish. Stressors in production systems reduce the efficiency of catfish production. Research was designed to develop a tool for characterizing levels of physiological stress in catfish. The proopiomelanocortin (POMC) gene was cloned which provided DNA sequence useful for studying factors which regulate POMC gene activity. This molecular tool will allow us to measure variation in stress responses between fish, and correlate the responses of catfish with resistance to diseases and efficient growth. Genes encoding the Major Histocompatability Complex (MHC) class I and II molecules have been identified in a number of fish species, including the channel catfish, but there is still a dearth of knowledge concerning their functional roles in fish immune responses. Animals from two families were identified that were matched or mismatched for MHCI and MHC II types, and genetic linkage and cell culture studies were performed. These studies demonstrated that MHC class I and II genes do not exist on the same chromosome in catfish as they do in mammals. Preliminary functional studies also indicated spontaneous non-specific cytotoxic responses between cells of different fish are likely mediated by differences in the MHC class I, but not class II, region molecules. This research provides insights into the control of catfish immune responses. In mammals, CD45 plays an important role in T and B lymphocyte cell receptor and cytokine signaling, however very little is known about its role in fish. We cloned the catfish CD45 gene and determined the DNA sequence. The catfish genome contains one functional CD45 gene, which can be differentially expressed depending of the cell type and the level of activation of the cell. The catfish gene makes use of alternative exon splicing to produce variants of the molecule. This research provided basic knowledge of CD45 structure for future experiments that will discern the role of this molecule in the response of the catfish immune system to pathogens. Little is known about the role of repetitive elements in the channel catfish genome. A previously identified class of repetitive DNA sequences, termed XbaI elements, were visualized on channel catfish chromosomes using fluorescence in situ hybridization. The XbaI elements were located primarily at the centromere of every catfish chromosome. This information will assist researchers in the assembly of catfish genetic maps, the integration of genetic and physical genome maps, and the identification of chromosomal regions that control important production traits in catfish. The role of the natural killer cell is not well understood in the catfish immune system. The catfish gene encoding the Natural Killer Cell Enhancing Factor (NKEF) was identified and sequenced. The gene was found to be active in all major tissues, and NKEF gene expression slightly increased after exposure of fish to bacterial cell components. NKEF gene variation was used to place it on the catfish genetic map. This research provided tools to allow scientists to understand the function of natural killer cells and to explore whether DNA sequence variants in the NKEF gene are linked to disease resistance. Estimation of phenotypic and genetic (co)variances for economically important traits are required for development of a breeding program to produce superior catfish germplasm for release to catfish producers. Data is being collected for full and half-sib USDA 103 strain channel catfish families for growth, meat yield, and disease resistance to allow estimation of phenotypic and genetic (co)variances. This information is being used to develop superior germplasm for release to catfish producers which will benefit producers, processors, and consumers. Information on the relative economic value of various production traits is needed to determine the amount of emphasis that should be applied to selection for improvement in different traits. Research is currently underway to estimate the relative economic value of traits important in farmed catfish (growth, feed conversion, meat yield, disease resistance). This information will be used in combination with information on heritability of traits to more efficiently produce catfish germplasm. Meat yield, an economically important trait for farm-raised catfish, is affected by catfish strain/species and season of the year. A project comparing effect of season on meat yield of blue catfish, channel catfish and channel x blue catfish hybrids was completed. Hybrid catfish had higher whole carcass and fillet yield than blue or channel catfish during both the spring and fall. Blue catfish had higher carcass yield than channel catfish during both the spring and fall. Channel catfish had higher fillet yield than blue catfish in the fall, but blue catfish had higher fillet yield than channel catfish in the spring. Blue catfish had higher nugget yield (lower valued rib-meat) than channel or hybrid catfish. Catfish processors could increase processing yields by processing different genetic groups of catfish at certain times of the year. Economic losses associated with diseases are an increasing problem for farm-raised catfish producers. A study to compare fingerling growth and survival of 4 genetic groups of catfish was initiated, groups included: USDA 103 strain channel catfish, USDA 102 strain channel catfish, USDA 103 x USDA 102 crossbred, and USDA 103 strain channel catfish x blue catfish hybrids. Survival, growth, and disease resistance are being evaluated in these groups. The study should provide insight into disease resistance and growth that will be useful to farmers looking for groups of fish with improved disease resistance and to researchers attempting to understand genetics of disease resistance in catfish. It is important to understand the effects of dietary protein and feeding frequency on important traits such as meat yield of catfish (the percentage of whole fish weight that is saleable meat) do determine the most profitable combination of dietary protein and feeding frequency. Effects of dietary protein level and frequency of feeding on meat yield of channel catfish were determined. Meat yield increased as dietary protein level and feeding frequency increased. Catfish farmers need to consider the impact of dietary protein level and feeding frequency on fish growth and meat yield in determining the optimal protein level and feeding frequency. However, the optimal combination of dietary protein level and feeding frequency changes with the price of feed and the price of fish. 4c.List any significant activities that support special target populations. Development of catfish lines with superior performance for commercially important traits and utilization of these lines in commercial culture will help solve production problems, increase efficiency and profitability for both small and large catfish farmers, and provide a quality product for consumers. Most catfish producers with limited acreage buy fingerlings from large breeders that are very likely to utilize improved brood stocks, and the development and use of improved catfish lines can quickly affect the profits of small producers. Because small farms do not enjoy the same economies of scale experienced by larger operations, breeding fish with improved production traits will be highly beneficial to small farmers. The USDA Census of Aquaculture conducted in 2000 classified 84% of catfish farms as small businesses, with annual sales of less than $500,000. Of the 1,370 catfish farms in the United States, 38% (515) reported annual revenues of less than $25,000. 4d.Progress report. Under a Specific Cooperative Agreement 6402-31000-008-02S, “Candidate Genes for Catfish Disease Resistance,” with Mississippi State University. The problem of Enteric Septicemia of catfish (ESC), caused by the gram negative, facultative intracellular bacterium Edwadsiella ictaluri is the most economically important infectious disease to channel catfish aquaculture. We addressed the basis for pathogen resistance in mammals are often due to genes whose products function in the early stages of infection. Yet, little is known about the early response of channel catfish to E. ictaluri infection. Our objective was to observe the expression profile of select genes during the first 96 hours of infection by E. ictaluri. This involved the development of real-time rtPCR assays for transcripts of genes encoding an acute phase proteins (transferrin, serum amyloid P), macrophage responsive protein (Nramp), cell stress responsive protein (heat shock 70-HSP70), neutrophil proteins (beta integrin and E1B)and 18s ribosomal RNA. Then the expression of these genes were evaluated in two trial in which Channel catfish that were injected intraperitoneally with E. ictaluri or medium (controls) and then sampled. In the first trial the fish were sampled (n=7) at 6, 12, 24, 48, and 96 h after injection. The spleen, liver, anterior kidney (AK), and gut were sampled for RNA extraction. The posterior kidneys (PK) were collected for bacterial counts. In the second trial whole blood, liver and spleen were sampled for RNA expression analysis and tumor necrosis factor (TNF) was added to the gene expression profile. Also blood smears were taken for whole blood counts and samples of the liver and spleen were embedded in OCH medium and quick frozen for cryostat sectioning. The first trial demonstrated that most transcripts were found universally but, Serum Alkaline Phosphatase (SAP) and transferrin were liver-specific. Expression of EB1 (AK, spleen and liver), Hsp70 (AK and spleen), 1-integrin (liver) and Nramp (spleen and gut) significantly increased by 48 hpi. Transferrin was strongly up-regulated and SAP was down-regulated by 72 hpi, suggestive of positive and negative acute phase reactants. The second ESC challenge experiment, EB1, Hsp70, 1-integrin, TNF and Nramp expression were demonstrated in the peripheral blood. SAP and transferrin expression were not detected. TNF was upregulated at 24h in response to infection. The spleen also demonstrated a 24h TNF induction and this was followed by the EB1, Hsp70, 1-integrin, and Nramp induction at 48h and 72h. Cryostat sections were used for histochemical analysis of leukocyte population and for laser capture micro dissection (LCM) to evaluate tissue specific gene expression within the spleen. Analysis of the 72 hr samples demonstrate a marked increase in neutrophils throughout the spleen in infected fish. Neutrophils were focused around the periarterial lymphoid sheaths (PALS) in non-infected fish. LCM sampling of both tissue types from infected and non-infected fish and real-time quantitative PCR on the RNA extracts supports the data indicating differential distribution of neutrophils. EB-1 was non-detectable in parenchymal tissue, and expressed at a relatively low level in the PALS of non-infected tissue compare to high levels of expression throughout both tissues types in infected fish. As a comparison, HSP70 was expressed in both tissues in both groups but expression was highest in the infected parenchymal tissue. By evaluating expression of genes involved in the inducible portion of the innate defenses during the early stages of ESC we were be able to identify the critical time points that determine ESC outcome. The initial response occurs by 24 hours and secondary responses were apparent by 48 hours post-infection. This will allow researchers to focus on this critical window to identify the genetic basis of ESC resistance and factors that enhance or suppress the protective response. This report documents research conducted under Specific Cooperative Agreement 6402-31000-008-03S, “Channel Catfish Molecular Markers,” with Auburn University. The very first objective was to identify DNA markers within genes of known functions, particularly those already mapped on the human and zebrafish linkage maps, are valuable for mapping the catfish genome because they provide landmarks for comparative genome mapping. The second was to characterize certain innate immune-related genes, in particular the CXCL8 gene that appear to have multiple versions of cDNAs that may be exploited as a marker for mapping of this gene. Third, after initial development of genetic linkage maps using various resource families, it is important to integrate the maps using the same resource DNA. For the first objective, we took the approach of “intron tagging”. By amplification and sequence analysis of introns of known genes, polymorphic markers can be identified because intron sequences are highly variable and often contain microsatellites. On the basis of progress made in previous years, in the last year, we have completed the objective. We designed primers for 50 genes to amplify their introns. Of the 50 pairs of primers designed, 42 produced products. These PCR segments were sequenced. Sequence analysis of the 42 introns indicated that 8 (19%) included microsatellite sequences in them (Table 1). The sequences have been deposited to GenBank available for public use and their accession numbers are included in the publication. The information was transferred to the Catfish Genetic Research Unit whose personnel conducted mapping analysis of the polymorphic markers. This work was published in Animal Genetics (Serapion et al., 2004). Interleukin-8 is a CXC type chemokine produced in response to stimulation by pro-inflammatory cytokines or bacterial lipopolysaccharides. We cloned and characterized interleukin-8 cDNAs and its genomic segments containing all the exons and introns from channel catfish (Ictalurus punctatus). Multiple interleukin-8 cDNA clones were identified during analysis of expressed sequence tags (ESTs). Sequence analysis indicated presence of four types of alternatively spliced and two types of alternatively polyadenylated transcripts. Analysis of the genomic DNA of the locus confirmed that all four types of transcripts were likely derived from partial splicing. Channel catfish interleukin-8 gene has four exons and three introns with highly conserved splice sites as compared to interleukin-8 genes of other organisms. In spite of the structural conservation through evolution, the piscine interleukin-8 genes showed a much greater sequence divergence than their counterparts among mammals. RT-PCR indicated that two of the four splicing forms were expressed at high levels whereas the other two aberrant splicing forms were not detectable. Expression of interleukin-8 gene was up-regulated in channel catfish and blue catfish after infection with pathogenic bacteria Edwardsiella ictaluri. This work was published in Developmental and Comparative Immunology (Chen et al., 2005). We initiated an expanded objective: Integration of various linkage maps by using the same resource markers. In the last several months, experiments have been initiated to genotype microsatellite markers that were mapped using the channel catfish resource families. In this case, genotyping was conducted using PCR with channel catfish x blue catfish interspecific resource DNA. Several points are important for this objective:. 1)integration of existing genetic maps will enhance the resolution of the linkage maps; and. 2)use of the same set of microsatellite primers in different resource families will allow comparison of the mapping populations and their impact on genetic distances, particularly between the intraspecific and interspecific families. To date, almost 50 microsatellite primer pairs have been tested and genotyped with 64 fish from an interspecific family. Under a Specific Cooperative Agreement 6402-31000-008-04S, “Channel Catfish Breeding for Disease Resistance,” with the University of Arkansas, Pine Bluff. Research was conducted to improve selective breeding of channel catfish resistant to ESC. Channel catfish families were selected based on ESC resistance and challenged with Edwardsiella ictaluri at the Catfish Genetics Research Unit and blood samples collected before and after challenge were tested for humoral immune function in the laboratory of Aquaculture/Fisheries Center, University of Arkansas at Pine Bluff. Serum complement activity is constitutively higher in ESC resistant fish, it remains high after E. ictaluri challenge and there is no clear relationship between serum lysozyme activity and ESC resistance. Selection of fish based on humoral immune function could produce catfish resistant to ESC and it may be adventitious to select for catfish with high serum complement activity. This research was terminated in FY 2004 when the graduate student supported by ARS funding left the university. Under a Specific Cooperative Agreement 6402-31000-008-05R, “Sequence Characterization of the Channel Catfish Immunoglobin Heavy Chain Locus,” with University of Mississippi. Research was conducted to identify DNA clones containing the catfish immunoglobin heavy chain locus, sequence these clones, and establish the gene structure in this locus. Two large DNA clones were identified and fragmented to make DNA sub-libraries. The sub-libraries were sequenced using the high-throughput capacity of the USDA, ARS, Mid South Area Genomics Laboratory. Two additional DNA clones have been identified that overlap the ends of the initial clones and sequencing is ongoing. Final completion of the sequencing is scheduled for FY 2005. This information will be useful in determining the structure of this important immunity-related molecule from a primitive vertebrate to help determine how these molecules have developed in more advanced species. It will also assist researchers in understanding the functionality of the catfish immune system. 5.Describe the major accomplishments over the life of the project, including their predicted or actual impact. This project was initiated on 12/01/2004. Major accomplishments expected over the life of the project are: . 1)Continued improvement in performance of catfish line USDA103;. 2)development and publication of a second-generation catfish genetic linkage map containing conserved genes useful for comparative mapping; and. 3)continued development of a multi-trait selection program utilizing the USDA103 and other catfish germplasm to develop additional improved lines for future release to commercial producers. Action Plan components: Genetic Improvement sections b and c. 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? This current research project was initiated in November 2004. To date, the main transfer of science and technology has been scientific publication in peer-reviewed journals, popular press, discussions with catfish producers, and a joint ARS-MAFES extension workshop to educate catfish producers in the methods for producing hybrid catfish. The most significant transfer of technology under the previous project from the Catfish Genetics Research Unit was the development of the USDA103 catfish line, which demonstrated superior growth performance. Broodfish from this line, named NWAC103, were made available to commercial producers through a joint release by ARS and MAFES beginning February 1, 2001 and completed in February 2002. Another germplasm release is planned for the end of this project, but this will depend on acceptance and approval by partners and stakeholders. 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). Tucker C., Silverstein P, Camus A, Bilodeau L, Wise D, Waldbieser G. 2005. Channel catfish virus disease and NWAC103 channel catfish. The Catfish Journal. January 2005. p.8. Bilodeau, A.L., Small, B.C., Wolters,W.R., Wise, D.J. 2005. Early Host Response Improves Disease Resistance In Channel Catfish. Global Aquaculture Advocate. 2005. 8(3):84-85. Peterson, B.C., Small, B.C. 2005. The use of recombinant bovine growth hormone (rbGH; Posilac) to characterized the GH-IGF axis in channel catfish. [Abstract] Aquaculture 2005 Conference. New Orleans, LA. Jan 17-20, 2005. p. 323. Peterson, B.C., Bilodeau, A.L. 2005. Development of real time PCR assays to measure levels of somatostatin-14 and somatostatin-22 mRNA in channel catfish. Abstract, 15th International Congress of Comparative Endocrinology Meeting. Boston, MA. May 22-27, 2005. p. 110. Small, B. C, Bilodeau, A.L. 2004. Cortisol, stress, and pathogen susceptibility in channel catfish (Ictalurus punctatus). Proceedings of the 5th International Symposium on Fish Endocrinology. p. O57. Small, B.C., Peterson, B.C. 2005. Establishment of a time-resolved fluoroimmunoassay for measuring plasma insulin-like growth factor I (IGF-I) in fish: effect of fasting on plasma concentrations and tissue mRNA expression of IGF-I and growth hormone (GH) in channel catfish. Proceedings of Aquaculture America 2005. p. 424. Small, B.C., Chatakondi, N. 2005. AQUI-STM reduces routine-handling stress in mature channel catfish. Proceedings of Aquaculture America 2005. p. 424. Small, B.C., Chatakondi, N. Optimizing chemotherapeutic treatment of hybrid catfish eggs for maximal hatching success. Proceedings of Aquaculture America 2005. p. 425. Peterson B.C., Small, B.C. 2005. Effects of recombinant bovine growth hormone on growth and the GH/IGF axis in channel catfish. Proceedings of Aquaculture America 2005. p. 323. Barrero, M., Small, B.C., D’Abramo, L.R., Kelly, A.M., Hanson, L.A. 2005. Plasma steroid, cathpsin activity and egg size and protein content during in vivo oocyte maturation in four strains of channel catfish broodstock. Proceedings of Aquaculture America 2005. p. 23. Welker, T.L., Klesius, P.H., Arias, C.R., Small, B.C. 2005. Effect of hypoxia on stress and heat shock protein expression in channel catfish (Ictalurus punctatus Rafinesque). Proceedings of Aquaculture America 2005. p. 487. Small, B.C., Waldbieser, G.C., Murdock, C.A., Peterson, B.C. 2005. Molecular cloning, genomic organization and functional characterization of channel catfish growth hormone receptor: Changes in hepatic GHR mRNA expression following fasting and feeding exogenous cortisol. Proceedings of the XV International Conference of Comparative Endocrinology. Boston, MA. p.120. Murdock, C.A., Small, B.C., Waldbieser, G.C. 2005. Molecular cloning of channel catfish steroidogenic factor-1 and the effects of exogenous LHRH treatment on expression. Proceedings of the XV International Conference of Comparative Endocrinology. Boston, MA. p.101. Review Publications Bilodeau, A.L., Peterson, B.C., Bosworth, B.G. 2005. Response of toll-like receptors, lysozyme, and igf-i in back-cross hybrid (f1 male (blue x channel) x female channel) catfish challenged with virulent edwardsiella ictaluri. Fish and Shellfish Immunology 20:29-39. Bilodeau, A.L., Waldbieser, G.C. 2005. Activation of tlr3 and tlr5 in channel catfish exposed to virulent edwardsiella ictaluri. Developmental and Comparative Immunology 29:713-721. Karsi, A., Waldbieser, G.C., Small, B.C., Wolters, W.R. 2005. Genomic structure of the proopiomelanocortin gene and expression during temporal stress in channel catfish, Ictalurus punctatus. General and Comparative Endocrinology. Peterson, B.C., Small, B.C. 2005. Effects of exogenous cortisol on IGFBPs and mRNA expression levels of IGF-I and GH in channel catfish. Domestic Animal Endocrinology 28:391-404. Peterson, B.C., Bosworth, B.G., Bilodeau, A.L. 2005. Differential expression of IGF-I, IGF-II, and toll-like receptors 3 and 5 mRNA during embryogenesis in hybrid (channel x blue) and channel catfish. Comparative Biochemistry and Physiology 141:42-47. Peterson, B.C., Waldbieser, G.C., Bilodeau, A.L. 2004. IGF-I and IGF-II mRNA expression in slow and fast growing families of USDA103 channel catfish (Ictalurus punctatus). Comparative Biochemistry and Physiology 139:317-323. Peterson, B.C., Waldbieser, G.C., Bilodeau, A.L. 2005. Effects of recombinant bovine somatotropin on growth and abundance of mRNA for IGF-I and IGF-II in channel catfish (Ictalurus punctatus). Journal of Animal Science 83:816-824. Quiniou, S.M., Wilson, M., Bengten, E., Waldbieser, G.C., Clem, L.W., Miller, N.W. 2004. Mhc rflp analyses in channel catfish full-sibling families: identification of the role of mhc molecules in spontaneous allogeneic cytotoxic responses. Developmental and Comparative Immunology. Small, B.C., Chatakondi, N. 2005. Routine measures of stress are reduced in mature channel catfish during and following Aqui-S anesthesia and recovery. North American Journal of Aquaculture 67:72-78. Small, B.C., Bilodeau, A.L. 2005. Effects of cortisol and stress on channel catfish (Ictalurus punctatus) pathogen susceptibility and lysozyme activity following exposure to Edwardsiella ictaluri. General and Comparative Endocrinology 142:255-261. Weber, T.E., Small, B.C., Bosworth, B.G. 2005. Lipopolysaccharide regulates myostatin and myod independently of an increase in plasma cortisol in channel catfish (Ictalurus punctatus). Domestic Animal Endocrinology 28(1):64-73. Weber, T.E., Bosworth, B.G. 2005. Effects of 28 day exposure to cold temperature or feed restriction on growth, body composition, and expression of genes related to muscle growth and metabolism in channel catfish. Aquaculture 246:438-492.