Progress 06/01/07 to 05/31/12
Outputs
Progress Report Objectives (from AD-416): The objective of this cooperative research project is to conduct aquaculture research addressing fish health problems limiting the production of channel catfish, Ictalurus punctatus. Approach (from AD-416): All research conducted under this agreement will address practical, production oriented fish health or disease problems facing the farm- raised catfish industry in the southeastern United States and will address components of the USDA-ARS action plan for National Program 106- Aquaculture. Research projects will be conducted using pond and laboratory facilities at the Delta Research and Extension Center (National Warmwater Aquaculture Center, Stoneville, MS). These facilities are under the direction of the Mississippi Agricultural and Forestry Experiment Station (MAFES) and Mississippi State University College of Veterinary Medicine (CVM). Research will address problems in the following broad subject areas: fish health management, fish diagnostics, epidemiology, and risk assessment modeling. On-farm demonstration projects, scientific articles, technical bulletins and educational workshops will be used to disseminate information to stakeholders. Catfish health management uses diagnostics, field services, in addition to applied and basic research to address fish health concerns in warmwater aquaculture. Diagnostic services and research projects were conducted with the objective of improving the production efficiencies and profitability of catfish farming. The service component of the program centers around the Aquatic Research & Diagnostic Lab (ARDL) which provides catfish farmers with a comprehensive disease diagnostic service. The ARDL also provides a mechanism to monitor changing disease trends and identify emerging disease problems. Real-time polymerase chain reaction (PCR) tests have been developed and validated for the detection of significant catfish pathogens in pond environments and fish tissues. Assays have currently been validated for Bolbophorus damnificus, Edwardsiella (E.) ictaluri, Henneguya (H.) ictaluri, and Aeromonas hydrophila and are partially completed for Edwardsiella tarda and Flavobacterium columnare. Validated procedures have been integrated into disease surveillance and diagnostic programs and have been used to develop risk assessment models for proliferative gill disease and trematode infections. A digenetic trematode, Drepanocephalus spathans, previously thought inconsequential to catfish health, has been demonstrated as infective to juvenile catfish fingerlings that can result in mortality, further supporting the need for adequate snail control programs on commercial operations. Smallmouth buffalo were shown to reduce populations of Dero digitata in experimental pond trials and may serve as a biological control measure of proliferative gill disease. Field trials investigating the impact of smallmouth buffalo on Dero digitata on commercial farms has been initiated. Laboratory and field studies led to FDA approval of a new drug, florfenicol (FFC), to treat E. ictaluri and Flavobacterium columnare infections. An orally delivered enteric septicemia of catfish (ESC) vaccine has been developed and shown highly effective in preventing E. ictaluri infections in experimental pond trials resulting in an estimated increase in gross sales by $1, 400/acre. Improvements in delivery platforms have greatly improved the efficacy of vaccination against E. ictaluri infection and the integrated use of vaccines, restricted feeding schedules and florenicol medicated feed is anticipated to eliminate most bacterial infections associated with the production of fingerling catfish. In laboratory feed trials, amprolium was shown to be efficacious against developmental stages of the causative agent of proliferative gill disease (PGD). Visceral toxicosis of catfish was shown to be caused by botulinum neurotoxin-type E. Environmental sampling and disease surveillance programs (utilizing molecular, immunological and analytical methodologies) have been initiated to determine BotulinumE (BoNT/E) exposure at the population level and identify possible point sources of contamination and routes of intoxication. Catfish suffering from idiopathic anemia, a major cause of losses, were shown responsive to iron supplementation and may serve as supportive therapy.
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Progress 10/01/10 to 09/30/11
Outputs
Progress Report Objectives (from AD-416) The objective of this cooperative research project is to conduct aquaculture research addressing fish health problems limiting the production of channel catfish, Ictalurus punctatus. Approach (from AD-416) All research conducted under this agreement will address practical, production oriented fish health or disease problems facing the farm- raised catfish industry in the southeastern United States and will address components of the USDA-ARS action plan for National Program 106- Aquaculture. Research projects will be conducted using pond and laboratory facilities at the Delta Research and Extension Center (National Warmwater Aquaculture Center, Stoneville, MS). These facilities are under the direction of the Mississippi Agricultural and Forestry Experiment Station (MAFES) and Mississippi State University College of Veterinary Medicine (CVM). Research will address problems in the following broad subject areas: fish health management, fish diagnostics, epidemiology, and risk assessment modeling. On-farm demonstration projects, scientific articles, technical bulletins and educational workshops will be used to disseminate information to stakeholders. Methods to Protect Catfish with Vaccines: 1) Bactericidal antibiotics were shown more effective in treating infection than bacteriostatic drugs but could interfere with development of acquired immunity. 2) Trials with anticoccidial drugs, amprolium and salinomycin, in the feed during and after exposure to Henneguya (H.) ictaluri actinospores indicated that fish readily consumed diets containing 100 mg/kg amprolium but did not consume diets containing 60 mg/kg salinomycin. Smallmouth buffalo (SMB) may be effective for biological control of benthic oligochaetes, which transmit myxozoan parasites 3) see Objective 2. 4) Primer and probe combinations have been developed for specific amplification for B. damnificus, and the assay has been validated for field use. 5) IgM antibodies from blue, hybrid and channel catfish were affinity purified from their respective sera. Preliminary results indicate that 9E1 binds equally with the IgM antibodies of blue and hybrid catfish. With the currently available tools, immunoassays assessing humoral immunity in blues and hybrids have been developed to key catfish pathogens. Monitoring and surveillance of biotoxin, pathogen, pathogen/host interactions in aquatic environments 1) Primer and probe combinations of real-time polymerase chain reaction assays have been developed for specific amplification for Edwardsiella (E.) ictaluri, Flavobacterium (F.) columnare, Bolbophorus (B.) and H. ictaluri. Assays for H. ictaluri, B. damnificus and E. ictaluri have been validated. 2) Polyclonal antibodies for E. ictaluri were ordered and received and are being used to detect E. ictaluri. 3) The 2010 annual report summarizing the diagnostic case submissions, frequency of reported diseases and identification of emerging diseases was published and posted on the NWAC website. Highlights included the lack of any antibiotic resistance for Columnaris and Enteric Septicemia of Catfish. Diseases of hybrid and blue catfish were also separated out again this year to provide additional information of hybrids. 4) A toxin-free enzyme-linked immunosorbent assay (ELISA) procedure has been developed to measure the level of anti- Botulinum neurotoxin type E (BoNT/E) antibody in catfish sera. A peptide library has been constructed based on the known primary structure of the heavy chain subunit of BoNT/E. Archived channel catfish sera collected from fish exposed to BoNT/E has been screened for antibody binding with a panel of 69 synthetic peptides composed of 18 amino acid residues. Chemical and mycotoxin feed contaminants affect growth and disease resistance of catfish. 1) Studies have been initiated to evaluate the effects of mycotoxin contaminated feed on growth and disease resistance of catfish. Corn naturally contaminated with alfatoxin was used to formulate diets to contain graded toxin concentrations ranging from 0 to 160 ug/kg feed. Fish were fed and evaluated for growth and disease resistance. No differences in mean weight gain or mortality following disease challenge were observed. 2) Evaluate effects of chemical feed contaminants and adulterations on growth and disease resistance of channel catfish. Monitored by email and site visits.
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Progress 10/01/09 to 09/30/10
Outputs
Progress Report Objectives (from AD-416) The objective of this cooperative research project is to conduct aquaculture research addressing fish health problems limiting the production of channel catfish, Ictalurus punctatus. Approach (from AD-416) All research conducted under this agreement will address practical, production oriented fish health or disease problems facing the farm- raised catfish industry in the southeastern United States and will address components of the USDA-ARS action plan for National Program 106- Aquaculture. Research projects will be conducted using pond and laboratory facilities at the Delta Research and Extension Center (National Warmwater Aquaculture Center, Stoneville, MS). These facilities are under the direction of the Mississippi Agricultural and Forestry Experiment Station (MAFES) and Mississippi State University College of Veterinary Medicine (CVM). Research will address problems in the following broad subject areas: fish health management, fish diagnostics, epidemiology, and risk assessment modeling. On-farm demonstration projects, scientific articles, technical bulletins and educational workshops will be used to disseminate information to stakeholders. Efforts in fish health management research and diagnostic services have lead to the development of diagnostic tools for the identification of new emerging diseases, risk assessment models to predict the occurrence of disease related losses, and fish health management practices that have increased production efficiency. Of primary concern to commercial producers of fingerling catfish are the development of management practices to control bacterial infections, Edwardsiella (E.) ictaluri and Flavobacterium (F.) columnare, through integrated methods of prevention and treatment. Prevention focuses on effective vaccination schedules and restricted feeding practices with treatment relying primarily on the use of medicated feeds. Laboratory and field studies led to FDA approval of a new drug, florfenicol (FFC), against E. ictaluri infections and work is currently underway to obtain FDA approval of FFC against Flavobacterium columnare infections. Improvements in delivery platforms have greatly improved the efficacy of vaccination against E. ictaluri infection. Vaccines combined with restricted feeding schedules and FFC medicated feed is anticipated to eliminate most bacterial infections associated with the production of fingerling catfish leading to vastly improved yields and production efficiencies. Quantitative polymenose chain reaction (QPCR) tests have been developed and validated for the detection of disease agents in pond water and have been integrated into disease surveillance programs. Risk assessment models have been constructed for proliferative gill disease (PGD) and trematode infestations. These programs can eliminate PGD related mortalities associated with newly stocked fish and when integrated with an effective treatment program can effectively control the deleterious effects caused by trematode infestations. Visceral toxicosis of catfish (VTC) was shown to be associated with botulinum type E toxin (BoNT/E) Environmental sampling and disease surveillance programs (utilizing molecular, immunological and analytical methodologies) have been initiated to determine BoNT/E exposure at the population level and identify possible point sources of contamination and routes of intoxication. The involvement of other naturally occurring neurotoxins in VTC is also being investigated. Participants are in the same location and often converse through face to face meetings or phone conversation.
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Progress 10/01/08 to 09/30/09
Outputs
Progress Report Objectives (from AD-416) The objective of this cooperative research project is to conduct aquaculture research addressing fish health problems limiting the production of channel catfish, Ictalurus punctatus. Approach (from AD-416) All research conducted under this agreement will address practical, production oriented fish health or disease problems facing the farm- raised catfish industry in the southeastern United States and will address components of the USDA-ARS action plan for National Program 106- Aquaculture. Research projects will be conducted using pond and laboratory facilities at the Delta Research and Extension Center (National Warmwater Aquaculture Center, Stoneville, MS). These facilities are under the direction of the Mississippi Agricultural and Forestry Experiment Station (MAFES) and Mississippi State University College of Veterinary Medicine (CVM). Research will address problems in the following broad subject areas: fish health management, fish diagnostics, epidemiology, and risk assessment modeling. On-farm demonstration projects, scientific articles, technical bulletins and educational workshops will be used to disseminate information to stakeholders. Significant Activities that Support Special Target Populations Efforts in fish health management research and diagnostic services have lead to the development of diagnostic tools for the identification of new emerging diseases, risk assessment models to predict the occurrence of disease related losses, and fish health management practices that have increased production efficiency. Of primary concern to commercial producers of fingerling catfish are the development of management practices to control bacterial infections, Edwardsiella ictaluri and Flavobacterium columnare, through integrated methods of prevention and treatment. Prevention focuses on effective vaccination schedules and restricted feeding practices with treatment relying primarily on the use of medicated feeds. Laboratory and field studies led to FDA approval of a new drug, florfenicol (FFC), against E. ictaluri infections and work is currently underway to obtain FDA approval of FFC against Flavobacterium columnare infections. Improvements in delivery platforms have greatly improved the efficacy of vaccination against E. ictaluri infection. Vaccines combined with restricted feeding schedules and FFC medicated feed is anticipated to eliminate most bacterial infections associated with the production of fingerling catfish leading to vastly improved yields and production efficiencies. Quantitative Polymerase Chain Reaction (QPCR) tests have been developed and validated for the detection of disease agents in pond water and have been integrated into disease surveillance programs. Risk assessment models have been constructed for proliferative gill disease (PGD) and trematode infestations. These programs can eliminate PGD related mortalities associated with newly stocked fish and when integrated with an effective treatment program can effectively control the deleterious effects caused by trematode infestations. Visceral toxicosis of catfish (VTC) was shown to be associated with botulinum type E toxin (BoNT/E) Environmental sampling and disease surveillance programs (utilizing molecular, immunological and analytical methodologies) have been initiated to determine BoNT/E exposure at the population level and identify possible point sources of contamination and routes of intoxication. The involvement of other naturally occurring neurotoxins in VTC is also being investigated. This project was monitored through telecoms, e-mails and site visits.
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Progress 10/01/07 to 09/30/08
Outputs
Progress Report Objectives (from AD-416) The objective of this cooperative research project is to conduct aquaculture research addressing fish health problems limiting the production of channel catfish, Ictalurus punctatus. Approach (from AD-416) All research conducted under this agreement will address practical, production oriented fish health or disease problems facing the farm- raised catfish industry in the southeastern United States and will address components of the USDA-ARS action plan for National Program 106- Aquaculture. Research projects will be conducted using pond and laboratory facilities at the Delta Research and Extension Center (National Warmwater Aquaculture Center, Stoneville, MS). These facilities are under the direction of the Mississippi Agricultural and Forestry Experiment Station (MAFES) and Mississippi State University College of Veterinary Medicine (CVM). Research will address problems in the following broad subject areas: fish health management, fish diagnostics, epidemiology, and risk assessment modeling. On-farm demonstration projects, scientific articles, technical bulletins and educational workshops will be used to disseminate information to stakeholders. Significant Activities that Support Special Target Populations Proliferative Gill Disease (PGD), caused by the myxozoan parasite Henneguya ictaluri, is one of the most devastating parasitic infections affecting commercial channel catfish (Ictalurus punctatus) aquaculture. The current diagnostic techniques of gross examination of gill clip wet mounts and histopathology are subject to false negatives during the early stages of infection, and the quantifiable nature of end-point polymerase chain reaction (PCR) is subjective. To evaluate the pathogenesis of infection and predict potential losses associated with this disease, a real-time polymerase chain reaction (QPCR) assay was developed using published H. ictaluri specific end point PCR primers and protocols. The sensitivity of the assay was the equivalent of a single H. ictaluri actinospore, and in a pond challenge study, quantitative real-time PCR proved to be more sensitive than gross examination, microscopic examination of gill clip wet mounts, and histopathologic examination of gill tissue sections in detection of H. ictaluri. Also, utilizing the QPCR assay, two methods of determining the PGD status of a pond were evaluated. Method 1 quantifies the organism in gill tissue of sentinel fish held in a pond for 24 hours. Previous research has determined the infective window of the organism to be 24-48 hours; therefore, the parasite level in fish tissue exposed to a pond for 24 hours indirectly represents the rate at which the actinospore stage is being released by the oligochaete host. Method 2 involves quantification of H. ictaluri actinospores directly from pond water samples. Both methods identified parasite levels which indicate a high risk for losing fish stocked into the system and levels which represent a low risk of losing fish if water quality parameters are optimal. Future research will involve validation of these two procedures, disease monitoring studies to evaluate the prevalence and severity of PGD in commercial catfish production ponds and determine the relationship between PGD severity and environmental parameters and the evaluation of potential chemotherapeutic and biological treatments for this disease. An �immuno-capture� system to extract E. ictaluri or F. columnare cells from pond water has been developed and successful in laboratory settings but has been unsuccessful in field applications. A water filtration method using the real time PCR assay has been developed in substitution of the �immuno-capture� method. The procedure is being optimized for field applications with limited success. Publications: Silverstein P., B. Bosworth, P. Gaunt. 2008. Differential susceptibility of blue catfish, channel catfish, and blue x channel catfish hybrids to channel catfish virus. Journal of Fish Diseases. 31:77- 79. Mauel M.J., C. Ware, P. Smith. 2008. Growth of Piscirickettsia salmonis on enriched bloor agar. Journal of Veterinary Diagnostic Investigations. 20(2):213-214. Soto E, M. Lawrence, M.J. Mauel. 2008. Improved pulsed-field gel electrophoresis procedure for the analysis of F. columnare isolates previously affected by DNA degradation. Veterinary Microbiology. 128:207�212.
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Progress 10/01/06 to 09/30/07
Outputs
Progress Report Objectives (from AD-416) The objective of this cooperative research project is to conduct aquaculture research addressing fish health problems limiting the production of channel catfish, Ictalurus punctatus. Approach (from AD-416) All research conducted under this agreement will address practical, production oriented fish health or disease problems facing the farm- raised catfish industry in the southeastern United States and will address components of the USDA-ARS action plan for National Program 106- Aquaculture. Research projects will be conducted using pond and laboratory facilities at the Delta Research and Extension Center (National Warmwater Aquaculture Center, Stoneville, MS). These facilities are under the direction of the Mississippi Agricultural and Forestry Experiment Station (MAFES) and Mississippi State University College of Veterinary Medicine (CVM). Research will address problems in the following broad subject areas: fish health management, fish diagnostics, epidemiology, and risk assessment modeling. On-farm demonstration projects, scientific articles, technical bulletins and educational workshops will be used to disseminate information to stakeholders. Significant Activities that Support Special Target Populations This report documents research conducted under a Specific Cooperative Agreement between MS Agricultural & Forestry Experiment Station (MAFES) and ARS. Additional details for this subordinate project may be found in the in-house project 6402-31630-001-00D, �Improving Production Efficiency of Pond-Raised Channel Catfish.� Objective 1: Develop diagnostic tools to detect and monitor diseases in commercially raised channel catfish and determine virulence factors associated with those diseases. Hypothesis 1: A real-time PCR assay can be developed to study the epidemiology of proliferative gill disease. Proliferative Gill Disease (PGD), caused by the myxozoan parasite Henneguya ictaluri, is one of the most devastating parasitic infections affecting commercial channel catfish (Ictalurus punctatus) aquaculture. To evaluate the pathogenesis of infection and predict potential losses associated with this disease a real-time polymerase chain reaction (QPCR) assay was developed using published H. ictaluri specific end point PCR primers and protocols. The sensitivity threshold of the assay was less than 1 H. ictaluri actinospore and there was no subsequent amplification of the genomic DNA of several closely related myxozoans commonly found in channel catfish ponds. Disease monitoring studies are being conducted to evaluate the prevalence and severity of PGD in commercial catfish production ponds. Further application of this assay will involve establishing a critical H. ictaluri actinospore concentration to identify the threshold at which ponds are safe to stock and evaluating potential chemotherapeutic and biological treatments Hypothesis 2: A real-time immuno-PCR assay can be developed to study the epidemiology of enteric septicemia of catfish. An �immuno-capture� system to extract E. ictaluri or F. columnare cells from pond water has been developed and used successfully in laboratory settings, but has been unsuccessful in field applications. A water filtration method using the real time PCR assay has been developed in substitution of the �immuno-capture� method. The procedure is being optimized for field applications. Hypothesis 3: Early stages in the pathogenesis of enteric septicemia can be elucidated by examining differential protein expression by Edwardsiella ictaluri exposed to channel catfish intestinal mucus. We have found that in pond water and when exposed to catfish tissues and mucus E. ictaluri immediately adheres to larger particles. Isolating the bacteria from the host tissues and mucus without changing the protein expression of the bacteria has proven difficult. An �immuno-capture� system to extract E. ictaluri cells from pond water and catfish tissues has been developed, and is being optimized to isolate the bacteria with little or no host materials. Once a method of isolating the bacteria from the catfish tissue and mucus components has been optimized the characterization of the bacterial proteins expressed in vivo and in pond water will proceed. Project was monitored by regular discussions and interactions between the MAFES staff and the CGRU. This project continues work of 6402-31630-001- 02S that terminated in May 2007.
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