SUMMARY OF RESEARCH AND DIAGNOSTIC ACTIVITIES

SUMMARY OF RESEARCH AND DIAGNOSTIC ACTIVITIES 2005 Water Quality ● Production Systems ● Fish Health Thad Cochran NATIONAL WARMWATER AQUACULTURE CENTER Thad Cochran National Warmwater Aquaculture Center 2005 Program Summary Water Quality, Production Systems, and Fish Health This document summarizes current research and diagnostic activities in water quality, production systems, and fish health at the Thad Cochran National Warmwater Aquaculture Center. Projects are listed by specific subject area and include work that was completed or published within the last year and projects that are ongoing in 2005 or planned for 2006. Contents Personnel...................................................................................................................................... 2 Contacts........................................................................................................................................ 4 Publications and Internet Contacts ............................................................................................... 5 Water Quality Off-flavor management ..................................................................................................... 6 Dissolved oxygen and aeration ......................................................................................... 8 Pond ecology and phytoplankton ...................................................................................... 9 Water quality during harvest and transport ..................................................................... 11 Hatchery water quality..................................................................................................... 12 Environmental protection ................................................................................................ 12 Production Systems Fry nursery pond management ....................................................................................... 14 Intensification of pond culture ......................................................................................... 15 Fish Health Diagnostics...................................................................................................................... 16 Epizootiology................................................................................................................... 17 Stress, immunology, and disease development.............................................................. 18 Enteric septicemia of catfish (ESC)................................................................................. 19 Columnaris ...................................................................................................................... 23 Other bacterial diseases ................................................................................................. 25 Channel catfish virus disease (CCVD)............................................................................ 25 Visceral toxicosis of catfish (VTC)................................................................................... 27 Channel catfish anemia (CCA)........................................................................................ 28 Proliferative gill disease (PGD) ....................................................................................... 28 Trematodes ..................................................................................................................... 30 Hatchery-associated disease problems .......................................................................... 31 1 Water Quality, Production Systems, and Fish Health Personnel USDA/ARS Catfish Genetics Research Unit (USDA/ARS-CGRU) Dr. Lanie Bilodeau is a Research Molecular Biologist whose research focuses on improving resistance to ESC in channel catfish and identifying immunological factors associated with disease resistance. She is also working to identify genetic markers useful for marker-assisted selection for disease resistance and to develop non-lethal screening tools for identification of fish with innate resistance to ESC. Dr. Peter Silverstein is a Research Virologist whose research focuses on catfish immune responses to viral pathogens. Some of this research will look at mechanisms of resistance to channel catfish virus, while other projects will focus on the role of specific aspects of the immune system in disease resistance. Dr. Brian Small is a Research Physiologist working in the area of fish stress as it impacts disease susceptibility, growth, and reproduction. This includes research to identify genetic mechanisms that control growth and spawning success. He is also working on methods to improve egg hatching rates. Dr. Les Torrans is a Research Fisheries Biologist working in the area of intensifying catfish production through improvements in the pond environment. He is developing new aeration technology and identifying optimal oxygen requirements for catfish. Dr. Paul Zimba is a Research Microbiologist whose research is focused on optimizing pond water quality. His research involves identification and control of harmful algal species and the use of remote sensing for identification of algal blooms. MSU Mississippi Agricultural and Forestry Experiment Station (MSU-MAFES) Dr. Terry Greenway, Assistant Research Professor. Dr. Greenway has a split research appointment with MAFES and MSU-CVM and conducts research on immunology, vaccine development, and vaccine delivery strategies. Dr. Charles Mischke, Associate Research Professor. Dr. Mischke has a 100% research appointment with MAFES and conducts research on fry and fingerling pond management, general pond ecology, and innovative pond production systems. Dr. Craig Tucker, Research Professor, NWAC Director, and Director of the USDA Southern Regional Aquaculture Center. His appointments are 80% MAFES; 11% CVM; and 9% MSU-ES. In addition to his administrative responsibilities, Dr. Tucker conducts research on off-flavor management, environmental impacts of aquaculture, and general water quality management. Dr. David Wise, Research Professor. His appointments are 63% MAFES; 24% CVM; and 13% MSUES. Dr. Wise conducts research on fish diseases and fish health management, and he is the leader of the applied fish health research program at NWAC. 2 MSU College of Veterinary Medicine (MSU-CVM) Dr. Al Camus, Assistant Professor of Aquatic Animal Health, is a veterinary pathologist and Director of the Aquatic Diagnostic Laboratory at the Thad Cochran National Warmwater Aquaculture Center. In addition to his primary diagnostic role, Dr. Camus participates in a variety of research projects, investigation of new and emerging diseases, and intern training. Dr. Pat Gaunt, Associate Professor of Aquatic Animal Health, is a board-certified veterinary toxicologist. Dr. Gaunt’s time is split evenly between diagnostic responsibilities and research. Dr. Gaunt’s research studies on the efficacy, safety and residue depletion for Aquaflor use in channel catfish were pivotal for its recent FDA approval. In addition, Dr. Gaunt investigates emerging diseases associated with toxicoses and participates in intern training. Dr. Michael Mauel, Assistant Professor of Aquatic Animal Health, is a microbiologist. Dr. Mauel shares in diagnostic responsibilities at the NWAC; however, research in the area of molecular biology is his primary responsibility. Dr. Mauel also participates in intern training and graduate student instruction. 3 Dr. Lanie Bilodeau Research Molecular Biologist National Warmwater Aquaculture Center USDA/ARS Catfish Genetics Research Unit P.O. Box 38, Stoneville, MS 38776-0197 Phone: (662) 686-3591 abilodeau@ars.usda.gov Dr. Al Camus Assistant Professor College of Veterinary Medicine and National Warmwater Aquaculture Center Mississippi State University P.O. Box 197, Stoneville, MS 38776-0197 Phone: (662) 686-3305 camus@cvm.msstate.edu Dr. Patricia Gaunt Associate Professor College of Veterinary Medicine National Warmwater Aquaculture Center P.O. Box 197, Stoneville, MS 38776-0197 Phone: (662) 686-3237 gaunt@cvm.msstate.edu Dr. Terry Greenway Assistant Research Professor National Warmwater Aquaculture Center Mississippi State University P.O. Box 197, Stoneville, MS 38776-0197 Phone: (662) 686-3583 greenway@drec.msstate.edu Dr. Michael Mauel Assistant Professor College of Veterinary Medicine National Warmwater Aquaculture Center Mississippi State University P.O. Box 197, Stoneville, MS 38776-0197 Phone: (662) 686-3303 mauel@cvm.msstate.edu Dr. Charles Mischke Associate Research Professor National Warmwater Aquaculture Center Mississippi State University P.O. Box 197, Stoneville, MS 38776-0197 Phone: (662) 686-3560 cmischke@drec.msstate.edu Dr. Peter Silverstein Research Virologist National Warmwater Aquaculture Center USDA/ARS Catfish Genetics Research Unit P.O. Box 38, Stoneville, MS 38776 Phone: (662) 686-3531 psilverstein@ars.usda.gov Dr. Brian C. Small Research Physiologist National Warmwater Aquaculture Center USDA/ARS Catfish Genetics Research Unit P.O. Box 38, Stoneville, MS 38776 Phone: (662) 686-3586 bsmall@ars.usda.gov Dr. Les Torrans Research Fishery Biologist National Warmwater Aquaculture Center USDA/ARS Catfish Genetics Research Unit P.O. Box 38, Stoneville, MS 38776 Phone: (662) 686-5460 ltorrans@ars.usda.gov Dr. Craig Tucker Director and Research Professor National Warmwater Aquaculture Center Mississippi State University P.O. Box 197, Stoneville, MS 38776-0197 Phone: (662) 686-3286 ctucker@drec.msstate.edu Dr. David J. Wise Research Professor National Warmwater Aquaculture Center Mississippi State University P.O. Box 197, Stoneville, MS 38776-0197 Phone: (662) 686-3239 dwise@drec.msstate.edu Dr. Paul Zimba Research Microbiologist National Warmwater Aquaculture Center USDA/ARS Catfish Genetics Research Unit P.O. Box 38, Stoneville, MS 38776 Phone: (662) 686-3588 pzimba@ars.usda.gov 4 PUBLICATIONS NWAC publications may be viewed by visiting the publications page on the NWAC website: Direct links are also provided below. Research Publications Mississippi State University scientists at NWAC have authored or co-authored more than 750 publications from 1981 to the present. A list of publications by discipline is available at NWAC News The Thad Cochran National Warmwater Aquaculture Center Newsletter is published twice a year. The Newsletter contains research findings, diagnostic summaries, Extension recommendations, industry updates, and notice of upcoming events at the NWAC. The Newsletter is distributed to farmers, researchers, Extension agents, and suppliers in 27 states. The 10 newsletters are available at: MSU College of Veterinary Medicine Fish Diagnostic Laboratory Annual Reports MSU-CVM Fish Diagnostic Laboratory Annual reports are available for 2001 through 2004. These reports may be accessed at: Southern Regional Aquaculture Center Factsheets NWAC scientists and Extension personnel have authored or co-authored 31 factsheets for the Southern Regional Aquaculture Center. The complete collection of SRAC factsheets may be accessed at: MSU-MAFES and MSU-ES Publications Additional MSU publications are available through the Aquaculture Homepage on the MSUCares.com network at: 5 WATER QUALITY Off-Flavor Management Evaluation of sensory versus analytical detection of off-flavors (Casey Grimm, USDA-ARS Southern Regional Research Center, and Paul Zimba) Flavor checkers in processing plants are used to assess fillet quality and often their opinion of quality has been debated. Four processing plants provided fillets that were processed for analysis by flavor checkers; samples for gas chromatography/mass spectrometry were also obtained from each fish. The sensory detection limit for flavor checkers was determined to be 0.1–0.2 ppb for MIB and 0.25–0.5 ppb for geosmin. Results from both analyses were highly correlated; suggesting flavor checkers provided reliable evaluations. The present practice of flavor testing is a reasonable alternative to instrumental detection. Evaluation of an anthroquinone-based algicide for off-flavor management (Kevin Schrader, USDA Natural Products Lab, and Craig Tucker) Currently there are only two algicides, copper sulfate and diuron, registered for managing algae-related off-flavors in pond-raised catfish and their long-term status remains unclear. Mississippi State University scientists cooperated with personnel from the USDA-ARS Natural Products Research Unit to identify and evaluate new algicides for use in managing algae-related off-flavors. One promising compound—a modified anthroquinone—identified in laboratory screening was tested in outdoor tanks and ponds, and was found to be selectively algicidal to odor-producing blue-green algae, thereby reducing levels of odorous compounds in water. Evaluation of sodium carbonate peroxyhydrate as an algicide for off-flavor management (Craig Tucker and Kevin Schrader, USDA Natural Products Lab) An alternative algicide for managing blue-green off-flavors has recently been approved for aquaculture use by EPA. This chemical, sodium carbonate peroxyhydrate (SCP), is an oxidizing agent that works by releasing hydrogen peroxide when dissolved in water. The chemical is attractive because it degrades to water, oxygen, and carbon dioxide, leaving no residue in the pond or fish. However, field trials showed that effective dosages varied considerably from pond to pond, but were generally in the range of 5 to 10 ppm. Effects were short-lived and multiple treatments would probably be needed to effectively eliminate odorous blue-green algae and treat off-flavors. The cost and logistics of using SCP do not appear favorable at this time. Evaluation of a bacterial pathogen of blue-green algae as a biocontrol agent for off-flavor (Kevin Schrader, USDA Natural Products Lab, Craig Tucker, and Lynn Walker, Louisiana Tech) Current off-flavor management is based on using chemical algicides to eliminate odorous blue-green algae. All chemical algicides suffer the disadvantages of leaving residues in the pond and fish. Biological control of odorous algae using natural pathogens is an attractive alternative. A bacterium that kills odor-producing blue-green algae has been isolated by Louisiana Tech scientists. The bacterium is strikingly effective under laboratory conditions but results have been highly variable in 2 years of field testing at Stoneville. Additional work is planned to identify the basis for the mixed success when using this organism for algal management. 6 Evaluation of a new herbicide for control of undesirable blue-green algae (Paul Zimba) Selective control of problematic algae in aquaculture ponds has been an elusive goal. Efficacy of various compounds was tested and one compound has been identified that exhibits desirable control at reasonable application rates (0.3 mg/L to kill 50% of algae present). Current works include obtaining an EPA use permit and ARS patenting. This chemical should provide a cost effective alternative to copper compounds and diuron for control of off-flavor and toxin-producing algae. Evaluation of silver carp polyculture to reduce blue-green algal off-flavors (Craig Tucker) At present, using chemical algicides to eradicate odor-producing algae is the most dependable means of preventing off-flavors in catfish. Nevertheless, using algicides to control flavor problems has several disadvantages and there has been interest in biocontrol of plankton communities using planktivorous fishes. In this study, low densities (0, 25, or 100 fish/acre) of silver carp were co-cultured with catfish. Silver carp did not eliminate odor-producing blue-green algae from pond phytoplankton communities and did not reduce the incidence or intensity of off-flavors in catfish. On-farm evaluation of the effectiveness and economics of copper sulfate for off-flavor management (Kevin Schrader, USDA Natural Products Lab, Craig Tucker, and Terry Hanson, MSU Ag Econ Department) This study evaluated the effectiveness of weekly low-dose applications of copper sulfate (0.5 ppm) on the prevalence of off-flavor in catfish on commercial farms. The study was conducted over 3 years in ponds on two farms. Overall prevalence of off-flavor was reduced by 50% based upon fish flavor analysis. Copper sulfate treatment reduced potential harvest delays by nearly half and reduced costs associated with off-flavor by 35%. Based upon these results, weekly low-dose applications of copper sulfate appear to be beneficial in mitigating musty off-flavor problems in commercially produced catfish. Diuron residues in catfish exposed to the chemical over multiple years (Craig Tucker) Diuron has been a useful off-flavor management tool for almost 10 years, yet EPA continues to question the environmental and health impacts of its use in catfish ponds. A study was conducted to determine whether multi-year exposure of channel catfish to diuron resulted in tissue residues exceeding the EPA tolerance level of 2.0 ppm. The study showed that diuron residue levels in fillets remained well below 1 ppm after treatment and there was no carryover of residues in fish from one year to the next. Results show that this algicide can be safely used to manage this important problem in catfish farming. Data from this study were critical in the recent favorable EPA ruling to allow a full registration for this chemical in the catfish industry. Uptake and depuration of off-flavor compounds from catfish (Paul Zimba and Casey Grimm, USDA-ARS Southern Regional Research Center) Off-flavor in catfish fillets is very costly to the industry. We evaluated MIB uptake and removal from channel catfish. Fish tissue rapidly accumulates MIB to ambient levels within 24 hours. In flowing well water, MIB was removed from fish fillet tissue at approximately 50% reduction per day. As “typical offflavor” concentrations in fish exceed 3 ppm, it takes over 3 to 4 days for fish to have below detection levels of off-flavor. In certain farm situations direct purging may be useful for reducing off-flavor. 7 Dissolved Oxygen and Aeration Evaluation of a novel u-tube aerator for catfish ponds (Les Torrans and Craig Tucker) A three-year study was completed using 15, 1-acre ponds at the Delta Western Research Center comparing the performance of internal u-tube aerators (Wilfley Weber Subterranean Stratamix Aerators) with conventional paddlewheel aerators operated at different oxygen set-points. Fish in the u-tube treatment consumed less feed and grew to a smaller size than fish in both paddlewheel treatments. Food conversion was similar, but net production in the u-tube treatment was 493 lbs/acre less than the low oxygen paddlewheel treatment and 1502 lbs/acre less than the high oxygen paddlewheel treatment (P<0.05). However, the incidence of off-flavor was significantly reduced in the u-tube ponds. U-tube aerators also are difficult to install and probably have little potential to replace paddlewheels in conventional ponds. Effects of low oxygen on channel catfish production (Les Torrans) From 2001 through 2004 seven studies were conducted using monitored and controlled aeration in 0.25acre and 1-acre ponds on two different facilities. These studies consistently demonstrated that maintaining the dissolved oxygen (DO) above 3.0 ppm does not improve catfish food consumption, growth or production, but does significantly increase aeration costs. When the DO is allowed to drop to between 2.0 and 3.0 ppm on a nightly basis, production parameters are significantly affected. If the DO is allowed to drop to 1.5 mg/L on a nightly basis, there are major impacts on performance. While food conversion is not significantly affected, average fish weight is reduced by 31%, net production by 54%, and food consumption by 45%. The best trade-off between aeration costs and production appears to occur in the DO range of 2.0 to 3.0 ppm. Effects of paddlewheel placement on fish production (Les Torrans in cooperation with Paul Dees, Dillard and Company, Inc.) Ten, 17-acre commercial ponds were used in a study of aerator placement. Rather than the traditional method of placing paddlewheels adjacent to each other to create a single current which circulates the entire pond, paddlewheels in five ponds were arranged to circulate water in approximately 25% of the pond area. The premise was that for a similar aeration cost, a slightly higher DO concentration could be maintained in the area where the fish are congregated. Based on previous research, this should result in increased feed consumption, growth and production. All ten ponds have been through one single-batch production cycle; four ponds were re-stocked for a second production cycle (the fingerling shortage in 2004 limited the number ponds managed for food fish production during a second cycle). While the differences in most parameters are not statistically significant, the new arrangement resulted in higher morning DO, reduced aerator usage, reduced emergency (tractor) aeration, slightly higher fish production, and improved food conversion. Development of a new aerator: the Power Tube (Les Torrans with John Carradine of “Big John Aerators”) A prototype commercial-scale u-tube aerator has been constructed and installed in a pond at the MSU Delta Branch Experiment Station. The tube was fabricated from a 36-inch diameter corrugated culvert and was buried to a depth of 20 feet below the pond bottom. The motor, gear box, impeller and open loop vector AC drive were all installed as planned. This initial prototype shows promise. With an impeller speed of 150 RPM, the motor draws 12.7 amps (5.36 hp) and has an output of 8300 gpm. 8 Several diffuser types have been tested. Oxygen transfer efficiency tests were conducted using a variety of diffuser types and configurations. The optimum conditions produced an increase in dissolved oxygen of 2.3 ppm (outflow DO – inflow DO) and an SAE of 1.66 pounds O2/HP-hr. These results were encouraging but were less than desired for commercial application. Technical (improving efficiency) and practical (reducing cost and producing a unit that can operate reliably in a commercial environment) recommendations will be incorporated into the Phase-II design for continued testing next year. Comparative oxygen requirements of blue and channel catfish (Les Torrans) It is generally accepted that the blue-channel catfish hybrid is a superior production fish. Unfortunately, reproductive constraints limit the widespread industry adoption of the hybrid. While the blue catfish has primarily been valued only for its role in the production of hybrids, it has several production traits of interest to our industry. Specifically, it is resistant to ESC, produces a larger dressed whole fish, and can be seine-harvested with near-100% efficiency. However, anecdotal information indicates that the species requires a higher dissolved oxygen concentration than channel catfish. A study is currently underway in 15 one-acre ponds comparing the practical oxygen requirements of blue and channel catfish. Further laboratory and pond studies on the oxygen requirements of blue and blue-channel hybrid catfish (compared to channel catfish) are planned for next year. Oxygen requirements of channel catfish eggs and fry (Les Torrans and Jim Steeby) This year a study was conducted to determine the oxygen requirements of channel catfish eggs and fry in commercial hatcheries. It was determined that oxygen stress results in premature hatching and a decreased survival to swim-up stage. Surprisingly, it was determined that a DO saturation as high as 94% (7.4 PPM at 27º C) resulted in premature hatching and 16% poorer survival to swim-up. A survey of 26 commercial hatcheries revealed that only 7 (27%) maintained a DO saturation high enough to prevent premature hatching. It is estimated that fry survival in our 1.8-billion fry industry could be increased by 150-300 million per year by increasing the DO concentration to near-saturation during the last day of egg incubation. Further research on this topic is planned for next spring. Pond Ecology and Phytoplankton Assessment of equipment for detection of blue-green algae in aquaculture ponds (Paul Zimba, Steve Thomson, USDA-ARS, and Anatoly Gitelson, University of Nebraska) Blue-green algae include a number of species known to produce off-flavor and toxic compounds that severely impact fish survival and farm profitability. We assessed the equipment needed for detection of algae during the past several years, including use of spray plane fitted with digital cameras/camcorders, hyperspectral sensors, and hand-held sensor systems. Digital camera/camcorder systems were the most versatile in terms of optimization; however, available filters were not sufficiently precise for identifying specific wavelengths needed to identify blue-green algae. Hyperspectral systems on aircraft provided high resolution pond imagery although effects of cloud cover and expenses associated with processing software and high equipment costs negated routine usage. Handheld spectral radiometers appear to offer the most promise because sensitivity of the hyperspectral systems is maintained, data processing can be conducted using spreadsheets, and startup costs are less than $10,000. Our goal is to identify a simple, affordable technology that will allow assessment of blue-green algal abundance in ponds. 9 Optimization of remote sensing data analysis methods for aquaculture ponds (Paul Zimba and Anatoly Gitelson, University of Nebraska) Remote sensing has been used in open oceans and clear water lakes to estimate algal biomass for over 50 years. Software to estimate algal biomass has been optimized for these non-turbid systems. Use of these programs in channel catfish ponds historically has resulted in poor temporal modeling capabilities (models generated from one day poorly fit data collected during subsequent sampling periods). Using hand held spectral radiometers, reflectance was measured, and water samples were simultaneously collected for turbidity and biomass determinations from ponds containing varying turbidity (suspended solids) and algal biomass. A conceptual model was applied to these data, and the model was “tuned” to minimize unexplained variance. For total algal biomass the resulting model fit was 14% better than that obtained from commercially available software (accounting for 87% of model variance). The new model was tested using jackknife analyses on 1.5 years of data and model fit to field data exceeded 70%. Current work involves building a conceptual model for blue-green algal biomass estimation and model evaluation. Development of specific relationships found in eutrophic production ponds is necessary to evaluate utility of remote sensing in these systems. Identification of a novel biotoxin produced by Euglena sanguinea (Paul Zimba, Richard Triemer, Michigan State University and Peter Moeller, NOAA) Fish kills have been attributed to an alga previously considered benign. The causative organism is from the division previously not known to produce toxins. Fish kills in MS, AR, NC, TX, and SC have been attributed to blooms of this alga. Cultures of this alga kill fish at densities of 900-1500 cells/mL. Current work includes identification of the toxin structure and culture studies to understand toxin synthesis. The alga is controlled by mixing and copper herbicide treatments. Euglenophycin, a novel biomarker pigment for the division Euglenphyta (Paul Zimba, Richard Triemer, Michigan StateUniversity, and Peter Moeller, NOAA) Rapid methods for assessing populations of algae are essential for pond management of harmful algal species. One method for accomplishing this separation of algal groups is through unique pigments present in different algal taxa. During routine pond analyses, we identified the first unique algal carotenoid specific for the division Euglenophyta. This pigment is found in 10 species of cultured euglenophytes representing all phylogenetic trees within the division. This biomarker will be useful for people using remote sensing methodology and pigment characterization of algal composition in catfish production systems. Pond age-water column trophic relationships in catfish production ponds (Paul Zimba, Charles Mischke) Long-term changes in nutrients, phytoplankton and zooplankton have not been previously investigated in production ponds. We analyzed the relationships of pond age with nutrients, zooplankton and phytoplankton populations, and the incidence of off-flavor occurrence. Nutrients accumulated in ponds during the first 3 years of production, but leveled off after this time. After year 4, the algal composition became dominated by filamentous cyanobacteria, and zooplankton composition was dominated by larger copepods and cladocerans. Younger ponds have lower incidence and intensity of off-flavor relative to older ponds. Results of this study may be used to help predict off-flavor occurrence, and management practices can be altered in older ponds. 10 Annual pond water biotic succession in channel catfish production systems (Paul Zimba, Charles Mischke, and Casey Grimm, USDA-ARS) While it is known that nutrient cycling is controlled by planktonic and benthic pond organisms, little is known regarding seasonality of micro-organisms in ponds. An annual survey has been initiated to follow seasonality of bacteria, fungi, phytoplankton, and zooplankton in seven, 4-acre ponds used for catfish grow-out. One clear preliminary finding is that bacterial densities are low compared to hypereutrophic lakes, suggesting that microzooplankton may play a key role in grazing bacteria populations. Optimization of trophic interactions between algae, bacteria, and zooplankton is necessary to maintain optimal pond function and promote fish health and production. Water Quality During Harvest and Transport Managing dissolved oxygen during harvest with the Sock Saver (Les Torrans, Sam Pilkinton, Bent Oak Fish Farm, and Charlie Hogue, MSU-ES) The Sock Saver was designed and built to hold three 50-gallon LOX Dewars that deliver oxygen to catfish being held in a sock through eight 200 scfh flow meters, each supplying a self-weighted bioweave diffuser. The transfer efficiency of these diffusers was over 15% in water 3.5 feet deep. Measurements showed it increased dissolved oxygen in a sock up to 0.9 mg/L. It was estimated that this system could meet over 80% of the oxygen demand of 50,000 pounds of catfish being held in a sock. The entire system cost less than $8,400. While it was designed to deliver oxygen at full capacity for nearly 11 hours, in practice flows rarely exceeded a third of maximum flow. At a LOX cost of $0.62/gal, it cost less than $3.00/hour to operate. A harvest crew using the Sock Saver felt it made a major difference in their ability to hold fish overnight, with fish having darker color (blanching of the skin is a sign of oxygen stress in channel catfish) and more energy while being loaded after a night in a sock. The Sock Saver has proven to be simple to operate, durable and virtually maintenance-free. It has a small footprint and does not interfere with other harvest equipment. A subsequent model designed and built on an existing trailer to hold two (leased) Dewars costs $2,300. Effects of transport water temperature, aerator type, and oxygen level on catfish fillet quality (Brian Bosworth, Brian Small, Charles Mischke) Changes in transport water quality may have adverse effects on catfish fillet quality. We determined the effects of transport water temperature, aerator type, and oxygen level on other transport water quality parameters, serum cortisol and fillet quality of catfish. Results indicated that increasing transport DO levels improve fillet quality. An economic analysis of the costs and benefits associated with increasing transport oxygen are needed to determine if changing current transport conditions is warranted. Effect of harvest and transport conditions on fish losses and fillet quality during processing (Craig Tucker, Les Torrans, and Brian Bosworth) Fish rejected by the processor (including fish dead on arrival and those with “red” or soft fillets) may range up to several percent of the fish processed during summer months. However, preliminary data collected this summer indicate that under “best management practices” total weighbacks may average only 0.6%. A large field study planned to begin in 2006 will examine the range of causes for reduced product quality, including pre-harvest fish health and condition, harvest techniques and equipment, water quality during harvest/holding, transport conditions/time, and post-delivery handling. 11 Hatchery Water Quality Environmental and physical factors affecting catfish egg hatching success (Brian Small and Terry Bates) These studies examined the effects of temperature, water Ca-hardness, and management techniques on catfish hatching rates. Results showed that developing catfish embryos are more sensitive to handling and changes in temperature and water hardness during the first 24-hours after spawning. The optimal temperature for holding and hatching catfish eggs is 78 and 82oF and Calcium hardness should be maintained above 10 ppm. Water temperatures outside this range or Calcium hardness below 10 ppm during the first 24-hour after spawning were shown to reduce hatching rate by 30 to 70%, respectively. Eggs not tempered, left on the pond bank for 30 minutes or more, or not treated with an anti-fungal therapeutant also exhibited significantly reduced hatching rates. These studies establish means of improving egg and fry survival to hatch. Environmental Protection Effluents from channel catfish hatcheries (Craig Tucker) Effluents from aquaculture facilities have recently come under scrutiny by regulatory agencies. In response to possible regulation, considerable research has been conducted to characterize the water discharged from aquaculture facilities. In this study, I assessed the quality of effluents from five commercial channel catfish hatcheries in the Delta region of Mississippi. Net pollutant loads (effluent concentration minus inflow concentration) were low for all variables. However, four of the hatcheries were supplied with groundwaters containing significant background levels of phosphorus or nitrogen. Nevertheless, concentrations of all potential pollutants are lower than corresponding concentrations in effluent-receiving streams in northwest Mississippi and total effluent volume from catfish hatcheries constitutes less than 0.02% of total annual streamflow in the region. It is therefore highly unlikely that catfish hatchery effluents will have a negative effect on receiving stream water quality. Quality of effluents from catfish ponds during draining (Craig Tucker and John Hargreaves, LSU) Development of effective waste management plans for pond aquaculture depends on characterization of the water discharged from ponds. This study assessed the quality of pond effluents when ponds are drained and characterized the nature of the material discharged. When ponds are drained, the initial flush of water discharged consists of pond water and a slurry of sediment that has accumulated over the screen inside the pond, but the effluent clears in 5 to 30 minutes and all water subsequently discharged is simply pond water. Since only a small proportion (1 to 4%) of the total solids discharged during pond draining was contained in the initial flush, it will be uneconomical to design elaborate treatment facilities to remove the material in that small volume of water. Characterization of solids discharged from catfish ponds (Craig Tucker and John Hargreaves, LSU) Processes that rely on gravity settling are the most economical method of removing solids from the initial flush of water released when ponds are drained. We measured the settling characteristics of solids in catfish pond effluent and used equations to calculate design criteria for settling basins. For average pond discharge rates, removal of 95% of solids requires a basin area ranging from 95 to 125 square meters. 12 We also assessed the capacity of common drainage ditches to remove solids and nutrients from pond effluents. At typical pond effluent discharge rates, more than 95% of the solids in initial pond draining effluent was removed after the effluent traveled 120 to 220 meters downstream in a 1-meter wide ditch. This was consistent with the results of basin-design modeling and show that it is unnecessary to build elaborate facilities to improve catfish pond effluent quality because considerable improvement in quality occurs as effluent flows down the simple drainage ditches that are common features of most commercial catfish farms. Effect of supplemental feed phosphorus on catfish pond effluent quality (Craig Tucker) Unused feed phosphorus is excreted into the culture water where it may be discharged and degrade the quality of effluent-receiving streams. Regulatory agencies sometimes promote modification of feed formulations as an approach to reducing phosphorus discharge from aquaculture facilities. To assess the effect of dietary phosphorus on potential waste discharge from ponds, phosphorus and organic matter were measured in catfish ponds during three feeding trials. Phosphorus modifications in the diets included using different levels of dicalcium phosphate, using defluorinated rock phosphate, and using phytase to liberate plant phosphorus in the feed ingredients. Modification of dietary phosphorus did not affect pond phosphorus or organic matter concentrations. Therefore, the source and level of dietary phosphorus in channel catfish feeds should be based primarily on nutritional and economic considerations rather than potential environmental impact. Evaluation of an environmental best management practices for catfish ponds (Craig Tucker) Improving the environmental performance of catfish can best be accomplished by using simple management practices rather than by using expensive “end-of-pipe” treatment technologies. A set of these “best management practices” or BMPs to conserve water and reduce pollutant discharge from ponds has been evaluated for 4 years. One of the practices, water-level management, was implemented to reduce effluent volume. The other three practices were implemented to reduce the concentration of substances in effluents. Those practices were 1) limiting daily feed inputs to 100 pounds/acre per day, 2) using a 28% protein feed, and 3) maintaining a maximum fish density of 7,500 fish/acre. Using these practices reduced pollutant discharge by over 50% and groundwater use by 60% without reduction in fish production. 13 PRODUCTION SYSTEMS Fry Nursery Pond Management Fertilization of catfish nursery ponds (Charles Mischke and Paul Zimba) Commonly used nursery pond fertilization practices were developed over 40 years ago and had not been studied in Mississippi ponds. We evaluated responses to several combinations of organic and inorganic fertilizer. Ponds responded more to nitrogen additions than phosphorus additions, and organic fertilizer did not produce a response. Increasing nitrogen application stimulated the algal bloom more quickly and significantly increased preferred zooplankton densities. We are now evaluating different nitrogen application levels. Changing the previous fertilization methods can increase densities of zooplankton and provide a healthy algal bloom which should lead to improved fry growth, survival and health. Zooplankton size and taxonomic selectivity of catfish fry (Charles Mischke, David Wise and Ryan Lane) Zooplankton consumption by catfish fry had previously not been well documented—types and sizes of preferred zooplankton were not known. We determined the zooplankton size and taxa selectivity of catfish fry. We found that catfish fry show a preference for the larger cladocerans and copepods. We concluded that fry stocking decisions should focus on copepod and cladoceran densities and sizes rather than total zooplankton abundance. Zooplankton sizes should be 450-700 micrometers for cladocerans and >650 micrometers for copepods. Stocking fry based on the abundance of preferred zooplankton may improve survival, growth and general fry health. Zooplankton nutritional value (Charles Mischke, Menghe Li, and Paul Zimba) Large zooplankton are readily consumed by catfish fry, but the nutritional contribution of these zooplankton was not known. We determined the nutritional value of zooplankton from catfish nursery ponds. The zooplankton consumed meet or exceed all nutritional requirements of channel catfish fry. These zooplankton are high in protein (65%), contain essential amino and fatty acids, and are excellent sources of vitamins and minerals. Because of the high nutritional value of zooplankton present in catfish nursery ponds, the importance of maintaining high densities of zooplankton is confirmed. Effects of stocking age on fry survival (Charles Mischke and David Wise) Generally, fry are held in hatcheries for ‘feed training’ before being stocked into ponds. Based on our zooplankton work, these fry may be missing out on the benefits of natural food organisms. We are currently comparing production characteristics of fry stocked into nursery ponds at different ages. The results of this study should help determine the most efficient length of time to hold fry in the hatchery. Importance of pH tempering for catfish fry (Charles Mischke and David Wise) Pond fertilization is necessary for proper algal blooms and zooplankton populations, but fertilization can cause a large shift in the pond pH. Most producers acclimate fry to temperature changes, but pH is rarely measured before stocking. We are determining fry tolerance to pH changes and the tempering times required for large differences in pH between the hatchery water and the pond water. Results from these studies will identify a possible mortality sink for fry, and modification of stocking practices will lead to improved fry survival. 14 On-farm studies to verify the effectiveness of fry pond management practices on a commercial scale (David Wise and Charles Mischke) Results from experimental trials are not always applicable on a commercial scale and need to be evaluated under commercial conditions before definite recommendations can be made. Results of fry pond management studies are being applied on a commercial scale with a cooperating farm. These new recommendations will be compared against previously used management practices to verify commercial application. Intensification of Pond Aquaculture Intensification of foodfish culture (Craig Tucker and Les Torrans) Some catfish farmers, particularly in the Black Belt of west Alabama and east Mississippi, report routine fish production that is considerably higher than the industry-wide average. The basis for increased production is not clear, but several aspects of pond management in west Alabama differ from the traditional approach used in the Mississippi Delta. Next year, we will compare two sets of production practices in 9-acre ponds at Stoneville. A set of ponds managed with traditional “Delta” practices will be compared with a set of “Alabama” ponds that are deeper, with more aeration, and intensely managed to prevent excessive algal abundance. The study will be conducted over multiple years and data will be analyzed to determine the economic benefits of intensified pond management. Effect of pond depth on water quality and fish production (Craig Tucker) Some farmers believe that deeper ponds are partly responsible for the high yields reported from Alabama catfish ponds. To investigate this possibility, ponds were maintained with average water depths of either 3.5 feet or 6 feet over the 2004 production season. All ponds were stocked with 6,000 fingerling channel catfish (40 pounds/1000 fish) per acre on May 20, 2004 and fed to satiation daily through fall. There were no treatment-related differences among concentrations of total ammonia, nitrite, or carbon dioxide. Net fish production was 4,582 pounds/acre in deep ponds and 4,029 pounds/acre in shallow ponds. The study is being repeated in 2005. Evaluation of modified partitioned aquaculture system (Craig Tucker and Charles Mischke) Catfish ponds should be capable of producing 15,000 pounds of fish per acre per year, yet average fish production across the industry falls far short of that potential. Engineers at Clemson University have developed a culture system that allows routine fish production at the limits of the potential for culture systems with no water exchange. The partitioned aquaculture system (PAS) as currently configured at Clemson consists of an extensive, shallow “waste-treatment” basin representing about 95% of the total system area and an intensive fish-confinement area in which fish are crowded at about 20-40 times the density of traditional ponds. A low-speed, energy-efficient paddlewheel circulates water between the fish-holding section and the waste-treatment section. A modified PAS system has been constructed at Stoneville that confines fish at a much lower density than the Clemson system. The overall concept is to take advantage of the fish confinement benefits of the PAS (facilitation of inventory, harvest, health management, and protection against predation) while avoiding the need for intensive system management. In 2005, net fish production has been approximately 12,000 pounds/acre and environmental conditions have remained excellent, indicating that higher loading rates are possible. Work is also planned at Stoneville in 2006 and 2007 using the true “Clemson” PAS system. 15 FISH HEALTH Diagnostics The Aquatic Diagnostic Laboratory (ADL), administered by the College of Veterinary Medicine, offers a comprehensive disease diagnostic service to Mississippi catfish producers. In 2004, the ADL received a total of 1,770 fish case submissions, 778 diagnostic and 992 research. Cases were received from 96 farms, or approximately 25% of the Mississippi industry. In addition, 978 water quality samples from 60 farms were analyzed. The laboratory is fully accredited by the American Association of Veterinary Laboratory Diagnosticians, which sets rigorous standards for quality assurance and supports guidelines outlined by the World Organization for Animal Health (OIE). Routine services performed include a visual examination, plus testing for the presence of bacterial, viral, fungal, and parasitic diseases. Every effort is made to provide farmers with a summary of preliminary findings on the day services are rendered, updates as needed, and a final written report when a case is completed. Whenever possible, treatment suggestions are also offered. In conjunction with its primary diagnostic role, the ADL works closely with MAFES fish health professionals to offer treatment recommendations to producers, monitor disease trends, provide surveillance for and investigation into the causes of new and emerging diseases, provide field service investigation, serve as an archive for bacterial isolates, and maintain a database for epidemiologic information relating to diseases of catfish. A unique opportunity is offered to student preceptors, interns, and graduate students seeking advanced training in fish disease diagnostics and research. The ADL supports the research efforts and diagnostic needs of other NWAC units, including MAFES, MSU Extension Service, MSU College of Veterinary Medicine and USDA-ARS Catfish Genetics Research Unit. Furthermore, the laboratory provides an outlet for the dissemination of information gained from research efforts back to producers. Disease Diagnoses as a Percent of Total Case Submissions 1997-2004 Disease Columnaris Disease Enteric septicemia Proliferative gill disease Winter fungus Channel Catfish virus Catfish Anemia Bolbophorus trematodes Visceral toxicosis Ichthyophthirius No pathogens identified Number of cases Avg 44.1 37.2 21.7 8.6 4.6 4.0 2.9 2.7 1.3 15.6 1452 2004 40.9 30.7 10.7 3.7 10.8 2.1 2.6 3.2 5.0 20.8 778 2003 44.7 34.7 10.8 5.3 8.9 5.2 1.1 3.7 0.5 18.3 832 2002 44.5 39.8 16.3 10.1 5.8 5.3 2.0 2.0 2.2 16.2 1057 2001 37.2 36.4 20.1 10.4 7.3 5.0 4.4 2.5 1.8 19.2 1602 2000 42.6 33.5 29.8 10.5 2.3 4.9 5.6 2.7 15.0 2189 1999 45.5 41.2 30.0 8.7 1.8 2.8 1.5 0.7 15.2 2007 1998 44.8 41.2 16.3 8.6 3.1 3.0 0.5 11.4 1647 1997 49.1 33.6 28.6 6.4 3.0 1.7 0.8 13.6 831 Epizootiology 16 The epidemiology of bacterial diseases in food-size channel catfish (David Wise, David Hardin, MSU-CVM, and Al Camus) The epidemiology of bacterial diseases was evaluated as part of the USDA National Animal Health Monitoring System which is tasked with continual surveillance of animal health in the US. A logistic regression model identified four risk factors associated with increased reporting of bacterial diseases: operation size, pond draining interval, and source and size of fish. This study was conducted to generate hypothesis describing possible managerial and environmental interactions that represent significant risks to production. Follow up studies are in progress to demonstrate these relationships on commercial catfish farms. Identifying the cause of these relationships will allow for the development of management strategies to decrease the incidence of disease related losses and increase production efficiency. Establishment of a computerized data-collection system to study production efficiency and disease losses (Al Camus, David Wise, Pat Gaunt, David Hardin and Fred Cunningham, MSU-CVM) The goal of this project is to develop a data collection and analysis system that will allow catfish producers to maintain a competitive edge in production and profitability. The College of Veterinary Medicine and MAFES have initiated a program to develop a computerized database that will allow producers to make management decisions, based on their own unique production records. Production parameters will ultimately be integrated with financial records to give producers an overall picture of the financial health of an operation. Diagnostic information integrated into the database will permit the analysis of relationships such as the cost of disease related losses and control programs, and correlations between disease incidence and specific production practices. At present the basic structure of the database has been constructed, although it is expected that it will continue to undergo modifications over time. In a pilot program, approximately 24 months of “back” records on hundreds of ponds has been entered into the database. The data includes stocking and harvest information, feed consumption, water quality, mortality estimates, and where available disease diagnostic information. In particular, the project will focus on determining risk factors associated with the development of visceral toxicosis of catfish (VTC) and channel catfish anemia (CCA). Stress, immunology, and models of disease development Development of techniques to increase vaccine efficacy using adjuvants and transcutaneous methods of delivery (Terry Greenway and David Wise) This project investigates the use of adjuvants (substances which can augment immune responses to vaccine) and vaccine delivery strategies to identify factors contributing to enhanced immune function and increased disease resistance. Certain microbial products are often used in conjunction with vaccines to heighten responses to vaccination. Moreover we are examining the potential use of substances which may temporarily enhance vaccine penetration through the skin. Additional mechanical means such as electroporation and ultrasound will be tested in conjunction with the above mentioned for potentiating vaccine delivery, and the subsequent immune response. 17 Interactions between stress and disease (Brian Small, Lanie Bilodeau, and Pete Silverstein) This research aims to identify correlations between catfish stress and disease susceptibility, and develop means (genetic or otherwise) to reduce stress-induced disease outbreaks. To date, our results demonstrate that there are differences in the stress response between genetic groups of catfish, and clearly show that stress conditions increase ESC susceptibility. In one study, crowding stress increased ESC-related mortalities by 26%. In an effort to determine what aspect of the stress response is responsible for increased disease susceptibility, we have shown that cortisol (the primary stress hormone in fish) is not singly responsible for increased ESC susceptibility. This research is ongoing and will be expanded to include studies with CCV. It is the goal of these studies to develop breeding and management strategies for improving catfish stress tolerance and disease resistance. Development of experimental models to induce bacterial, viral, parasitic, and fungal infections in channel catfish (David Wise and Terry Greenway) The ability to investigate the pathogenesis, treatment, and management of disease is dependent on the ability to induce and reproduce the pathological aspects of the disease. Developing disease challenge models is a priority research area and has led to procedures that can be used to study the pathogenesis and treatment of bacterial, parasitic, viral, and fungal infections in the field and laboratory. Interaction between nutrition, feeding, and disease resistance (David Wise, Ed Robinson, Terry Greenway, Menghe Li, and Bruce Manning) The objectives are to determine the impact of dietary components, mycotoxins, and feeding practices on disease resistance. This research has been used to help assess the impact of various feed formulations on fish health. Dietary supplements of trace minerals, vitamins, and menhaden oil were not shown to improve resistance of fish against E. ictaluri infection. This data suggests that dietary supplements will not dramatically improve disease resistance and that other strategies for improving fish health should be pursued. Development of immunobead-capture assays to study Edwardsiella ictaluri, Flavobacterium columnare, and Henneguya ictaluri, the causative agents of enteric septicemia of catfish (ESC), columnaris, and proliferative gill disease (PGD) respectively (Michael Mauel) An “immuno-capture” system to extract the respective disease agents from pond water is being developed. In this assay, antibody coated magnetic beads are used to extract the target organism from the water. The organisms can then be quantified by plating on microbiological media or by using molecular techniques (real-time PCR). The assays will be used to study how numbers of these disease organisms fluctuate with temperature throughout the year and if there are minimum numbers required to initiate outbreaks in ponds. It may then be possible to avoid outbreaks by maintaining numbers below this threshold by chemical or other means. To date the ESC and columnaris assays have been successfully used to measure the number of bacteria in filtered pond water to which known numbers of bacteria have been added. The assays are currently being tested using field samples of raw pond water. Investigation of components of the immune system in channel catfish (Lanie Bilodeau, Geoff Waldbieser) This study examined the response of two genes (toll-like receptors) associated with the immune system after fish were exposed to virulent Edwardsiella ictaluri. The amount of the genes present in tissue samples increased after exposure, indicating that they are important components of the immune system in 18 channel catfish. This information will be used in future studies investigating the components of the immune system that contribute to increased survivorship after ESC challenge. Measurement of toll-like receptor gene expression in backcross hybrid (channel x blue x channel) catfish (Lanie Bilodeau, Brian Peterson, Brian Bosworth) Blue x channel catfish hybrids show reduced susceptibility 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 increased survivorship after exposure to ESC. Enteric Septicemia of Catfish (ESC) Efficacy of Aquaflor® (Schering-Plough Animal Health) against Edwardsiella ictaluri in channel catfish (Pat Gaunt) An effective and palatable antibiotic in floating feed is needed in catfish to combat ESC. MSU-CVM conducted three tank efficacy trials, one pond efficacy trial, and one tolerance trial to assess palatability and safety, and one residue depletion study in support of FDA approval of Aquaflor® a new antibiotic for fish. Results showed that Aquaflor® is palatable, effective, and safe for use in catfish. Based on the residue-depletion study, withdrawal time of Aquaflor® in catfish was set at 12 days. This work was submitted to FDA and accepted within 89 days of submission. In October 2005, Aquaflor® became the first antibiotic approved in 20 years by FDA for use in catfish. Aquaflor® (Schering-Plough Animal Health) use in channel catfish: A pharmacokinetic study (Pat Gaunt) Based on the efficacy studies conducted at MSU-CVM, Aquaflor® at a dose rate of 10 mg active ingredient/kg of body weight was approved by the U.S. FDA for control of mortality associated with E. ictaluri. This study will clearly define how quickly the blood levels of Aquaflor® are achieved, how long they are sustained, and the rate at which the drug is cleared from the catfish bloodstream. Pharmacokinectic data is needed for this drug to substantiate the current recommended dose rate for ESC in catfish. Evaluation of AQUAVAC-ESC vaccinated fish fed fry diets supplemented natural pond biota using single- and continuous-dose challenge models (David Wise, Charles Mischke and Terry Greenway) Experiments were conducted to develop laboratory protocols that better reflect the growth characteristics of pond-raised fish and pathogenesis of natural E. ictaluri infection. Channel catfish fry were fed commercial diets supplemented with natural pond biota and vaccinated 10-12 days of age post-hatch. Following vaccination resistance of fish to E. ictaluri infection was evaluated using standard single- and continuous-dose challenge models. Mortality differences were not detected between vaccinated and nonvaccinated fish or among dietary treatments using a standard single-dose challenge model. In contrast, vaccination and diet significantly affected mortality when disease resistance was evaluated using the continuous-dose model. Using this method mortality was lowest in vaccinated fry fed supplemented 19 diets. This data also suggests that diet is critical in the growth and development of fry and that the methods used to induce disease in the laboratory can influence the outcome of disease challenge trials. Production efficiency of AQUAVAC-ESC/Columnare vaccinated fry on commercial fingerling operations (David Wise, Terry Greenway, and Charles Mischke) Experimental tests show that a commercially available vaccine (AQUAV-ESC, Intervet, Inc.) is effective in reducing ESC associated mortality. Commercial field trials showed significant increases in fish size, feed conversion, and net production of vaccinated fish. Vaccination increased net revenue by $500 to $1,000 per acre. Field and laboratory tests are also being conducted to evaluate the efficacy AQUAVACESC used in combination with AQUAVAC-Columnare in 7 to10-day old post-hatch channel catfish fry. Use of restricted feeding practices and medicated feed to control bacterial infections in channel catfish production systems (David Wise and Terry Greenway) Manipulation of feeding strategies has been shown to be very effective in controlling ESC and F. columnare infections in channel catfish fingerlings. Through restricted feeding practices, losses associated with bacterial infections can be reduced by 50 to 80%. The mechanism responsible for this observed response appears to be related to the ingestion of bacteria in the environment that inadvertently occurs during feeding. The program is developing management strategies that will maximize both size and survivorship and decrease the use of medicated feeds. This information is also being used to develop strategies for controlling disease related losses in growout ponds. Current projects focus on the strategic use of antibiotics to clear subclinical infections and enhance the development of acquired immunity before the onset of losses. Susceptibility of genetic strains and families of channel catfish to Edwardsiella ictaluri and PGD (David Wise, Terry Greenway, Brian Bosworth, and Lanie Bilodeau) Studies have been conducted to evaluate the response of genetic lines of channel catfish to bacterial and parasitic infections. Data from these studies demonstrated variation in susceptibility among lines and families of channel catfish to ESC and PGD, indicating considerable potential of increasing disease resistance through genetic selection. Evaluation of chemical treatments (chloramine-T, potassium permanganate, copper sulfate, and sodium carbonate peroxyhydrate) for controlling bacterial pathogens in pond water (David Wise, Terry Greenway, and Michael Mauel) The initial phase of the project will screen various chemicals for bactericidal properties against Flexibacter columnare (columnaris disease) and Edwardsiella ictaluri (ESC). The level of chemical that is shown to reduce pathogen levels in pond water will be evaluated for toxicity to fish to determine its potential as a disease treatment or as a disinfectant. The second phase will be conducted in experimental and commercial ponds to evaluate select bactericidal chemicals as a potential disease treatment. Procedures for isolating low concentrations of F. columnare from pond water have been developed and are currently being used to evaluate pathogen loading rates during columnaris outbreaks in experimental ponds. Immunological tests are being developed to detect Edwardsiella ictaluri in pond water. These procedures will allow for a more accurate evaluation of disease management strategies. 20 Investigation of early stages of gut invasion by the bacterium Edwardsiella ictaluri, the causative agent of enteric septicemia of catfish (Michael Mauel) Early stages in the infection process of ESC will be investigated by examining proteins produced by Edwardsiella ictaluri exposed to channel catfish intestinal mucus and in chambers placed in the abdomens of catfish. Disease processes result from precise interactions between the disease agent and its host. Edwardsiella ictaluri gains entry to catfish through the intestinal wall by the production of specific proteins following ingestion and contact with intestinal mucus and other catfish proteins. The goal of this project is to identify proteins responsible for gut invasion by E. ictaluri and examine the ability of these proteins to promote a protective antibody response. If protection is generated by injection with these proteins, they can then be incorporated into improved ESC vaccines that block early steps in the infection process. At this time, the procedures to identify the proteins are being developed using media grown bacteria. Kinetics of antibody response and exposure rates to bacterial pathogens in populations of commercially raised channel catfish to predict the occurrence disease epizootics (Terry Greenway and David Wise) This project investigates kinetics of antibody responses following nature exposure to E. ictaluri in the pond environment. Edwardsiella ictaluri levels in the natural environment will be monitored by a novel antibody base technique which can potentially raise detection signals 1000 to 10000 fold over conventional antibody based detection systems (currently under development). The number of fish which seroconvert along with the magnitude of the immune response generated, will be examined in conjunction with E. ictaluri concentrations in the pond. Attempts will be made to establish threshold antibody values resulting in protective immunity and establishing a predictive of number of bacteria that results in disease outbreaks. If the various parameters have predictive value as when a pond may break with ESC, production patterns could be altered to minimize losses. Preliminary trials conducted on a commercial fingerling farm in 2005 have demonstrated a strong correlation between seropositive populations of fish and increased disease resistance. This information was used to make recommendations on feeding practices to maximize growth in resistant populations of fish and minimize losses in susceptible populations of fish. Identification of high temperature tolerant strains of Edwardsiella ictaluri with implications to current farm management practices targeted for controlling disease outbreaks (Terry Greenway and David Wise) We have isolated several temperature tolerant E. ictaluri strains from diseased fish which differ markedly in their ability to confer protection and induce death upon lethal challenge. Unlike other field isolates, bacterial growth can occur at higher temperatures considered non-permissive to the virulent field isolates. Serologically, antibody responses to these temperature tolerant Edwardsiella strains exhibit little or no cross reactivity to virulent field isolates after primary immunization. We are currently investigating whether prior exposure to these temperature tolerant strains alters the kinetics and magnitude of anti-E. ictaluri immune responses as well as protection from challenge with virulent E. ictaluri. Differences in antigenic determinants will be determined to identify factors responsible for the induction of protective immunity (or lack of) and the development of improved vaccines. Development of a strain of catfish with reduced susceptibility to ESC (Lanie Bilodeau) One hundred 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 21 resistant families were pit-tagged and released into ponds. In 2006 years, these fish will be used for breeding and heritability of resistance to ESC will be measured. These fish will also be crossed with the “improved growth” line of USDA303’s that Brian Peterson has developed. Susceptibility to ESC and growth characteristics will be measured in all F1 offspring of both the “Resistant” line and the “Resistant” x “Growth” crosses. Assessing improvement in survival during ESC challenge (Lanie Bilodeau, Brian Bosworth) Multiple families of USDA103 and USDA303 channel catfish are currently being tested for survival during ESC challenge. Survival of families in each group will be compared to determine if improvements in survivorship have been made in the last 2 generations of selection of the USDA103 line (i.e., USDA303). This information will utilized in the selective breeding program to determine heritability levels of susceptibility to ESC and the effectiveness of multi-trait selection on ESC survivorship. Development of a genetic detection assay for Edwardsiella ictaluri (Lanie Bilodeau, Geoff Waldbieser, Bill Wolters, David Wise) A patent was awarded in 2005 for a genetically-based pathogen detection assay for Edwardsiella ictaluri, the causative agent of ESC. The assay is designed to detect genetic material of bacterial cells in small amounts of blood or other tissue samples. This assay is capable of detecting the equivalent of as few as 2.5 cells of E. ictaluri and is specific only to E. ictaluri. Current and future applications include use for monitoring bacterial levels in fish during ESC challenges, development of non-lethal screening tools for immunological assays, and use by diagnostic laboratories. Early host response reduces susceptibility to ESC in channel catfish (Lanie Bilodeau, Brian Small, Bill Wolters, David Wise) Bacterial levels and immune response were measured in three ESC-resistant and three ESC-susceptible families of channel catfish. Bacterial levels were suppressed in the resistant families and the non-specific immune response (lysozyme) was activated 24 hours earlier in resistant families than in susceptible families. The delay in immune response may be related to the development of acute infection in susceptible fish, resulting in increased levels of mortality. This information will be utilized for developing a protocol for screening families in innate resistance to ESC and for the identification of genes associated with innate immune function and disease resistance for marker-assisted selection. Determination of the response of different strains of catfish to ESC challenge (Lanie Bilodeau, Brian Peterson, Brian Bosworth) In an ongoing study, five strains of catfish were subjected to ESC challenge and bacterial levels, immune response, and growth are being measured. The goal of this project is to determine if differences in survivorship are related to differences in immune response. From previous studies, we know that strain differences exist regarding susceptibility to ESC, patterns of lysozyme activity, and timing of increases in expression of specific toll-like receptor genes. Data on survivorship and immune response will provide the framework for future studies regarding susceptibility to bacterial challenge. Development of a non-lethal screening assay for pathogen levels and immune response (Lanie Bilodeau) Responses to virulent and non-virulent E. ictaluri in resistant and susceptible families will be measured and correlated to develop a non-lethal test for disease resistance. The top and bottom 3% of families from 22 previous challenges will be selected and challenged with an attenuated live E. ictaluri vaccine (AQUAVAC-ESC, Intervet Inc., Millsboro, DE). Bacterial levels, mortality rates (for the virulent bacterial challenge), and immune response will be compared for the two treatments. If the results of the virulent/non-virulent ESC challenge indicate that the pathogen detection assay will successfully screen for survival and immune response during challenge, this test will be utilized in future studies. Columnaris Efficacy of Aquaflor® (Schering-Plough Animal Health) against Flavobacterium columnare in channel catfish: an in vitro study (Pat Gaunt) Columnaris (Flavobacterium columnare) is a serious bacterial disease affecting farm-reared catfish in the US. There are no currently approved antimicrobials for catfish to combat columnaris. There are reports of lack of efficacy of the extra-label use of Romet® and Terramycin® for treating this disease. The efficacy of Aquaflor® was assessed against field insolates of F. columnare using microbiology plates. The results show that Aquaflor® is efficacious against F. columnare in vitro, and future studies will look at its use in infected catfish. The efficacy of Aquaflor® (Schering-Plough Animal Health) against Flavobacterium columnare in channel catfish: a tank study (Pat Gaunt) Columnaris (Flavobacterium columnare) is a serious bacterial disease affecting farm-reared catfish in the US. There are no currently approved antimicrobials approved for use in catfish for combating columnaris. Aquaflor® is approved for use in catfish to control mortality associated with ESC. Extralabel use of Aquaflor® for other bacterial infections is prohibited. In vitro work indicates that F. columnare is sensitve to Aquaflor®, and tank studies are underway to test the drug’s efficacy against this bacterium. Results could support FDA approval of Aquaflor® efficacy against columnaris in catfish. Production efficiency of AQUAVAC-ESC/Columnare vaccinated fry on commercial fingerling operations (David Wise, Terry Greenway, and Charles Mischke) Experimental tests show that a commercially available vaccine (AQUAV-ESC, Intervet, Inc) is effective in reducing ESC associated mortality. Commercial field trials showed significant increases in fish size, feed conversion, and net production of vaccinated fish. Vaccination increased net revenue by $500 to $1,000 per acre. Field and laboratory tests are also being conducted to evaluate the efficacy AQUAVACESC used in combination with AQUAVAC-Columnare in 7 to10-day old post-hatch channel catfish fry. Investigation of disease important proteins of Flavobacterium columnare, the causative agent of columnaris (Michael Mauel) The pathogenesis of F. columnare will be investigated by comparing the differences in proteins produced when the bacteria are grown in two different environments, pond water and in chambers placed in the abdomens of catfish. The proteins produced only in the catfish abdomen will be examined for their ability to promote a protective antibody response. If protection is generated by injection with these proteins, they can then be tested in a vaccine. Procedures have been developed to produce sufficient bacteria to measure the variation in protein production using advanced proteomic technology. 23 Identification of genetic variation among isolates of Edwardsiella ictaluri and Flavobacterium columnare, and their role in virulence and epidemiology of disease (Michael Mauel) Archived E. ictaluri and F. columnare isolates will be examined for genetic variation using a variety of genomic “fingerprinting” techniques and a database established for use in future epidemiologic investigations. Applying these genetic tools will determine if variation exists among E. ictaluri and F. columnare isolates and will facilitate rapid identification of new bacterial strains within the catfish industry. It is anticipated that isolates of the two bacteria will fall into specific groups of varying ability to cause mortalities. Ultimately, these studies will identify the mechanisms used by highly lethal strains to cause disease that can be applied to improved vaccine technology in the future. Graduate student Esteban Soto is currently working on identifying variation within the approximately 70 isolates of F. columnaris from the collection of Dr. Mark Lawrence, College of Veterinary Medicine, Mississippi State University. Isolation and characterization of bacteriophages (viruses) that infect Flavobacterium columnaris (Michael Mauel) Bacteriophages are viruses that infect and destroy bacteria. Earlier researchers have reported bacteriophages that infect F. columnaris. The goal of this research is to isolate some of these bacteriophages and evaluate their potential as a treatment or preventative for columnaris disease. Recent studies with other bacterial fish diseases have shown the potential of greater than 50% reduction in mortality when bacteriophages have been incorporated in the diet of diseased fish. Other Bacterial Diseases Characterization of a new streptococcal disease causing losses of brood-sized channel catfish (Al Camus and Patricia Shewmaker, Centers for Disease Control) A previously unknown streptococcal bacterial disease has been diagnosed in brood-sized channel catfish on three Delta farms. The disease is characterized by severe weight loss, arching of the back due to progressive destruction of the vertebral column, and draining sores located primarily at the angle of the jaw and fin bases. Challenge trials have been conducted to fulfill Koch’s postulates demonstrating the organism in question is a true pathogen. Complete characterization of the bacterium is being conducted in collaboration with the Centers for Disease Control in Atlanta. Findings to date indicate this is a previously unknown species of bacteria. At present the significance of this emerging disease is unknown, but monitoring efforts are ongoing to determine its prevalence in the industry. Efficacy of Aquaflor® (Schering-Plough Animal Health) against Streptococcus iniae in nile tilapia: in vivo studies (Pat Gaunt and Al Camus) There are no approved antibiotics for use in most warmwater fish in the U.S. By “crop grouping” (i.e. selecting 2-3 representative species of warmwater fish to represent warm water fishes), the FDA will grant approval for all warmwater fish if efficacy and safety can be demonstrated in each of two to three representative species. MSU-CVM conducted a dose titration and a dose confirmation study in tilapia to assess the efficacy of Aquaflor® against Streptococcus iniae. Results showed that Aquaflor® is effective against S. iniae in tilapia. This work will be submitted to FDA in support of an approval package for use in all warmwater fish in the U.S. 24 Channel Catfish Virus Disease (CCVD) The effect of channel catfish virus upon non-specific immune mechanisms (Peter Silverstein) White blood cells (leukocytes) are one of the main targets of CCV infection; the same cells are responsible for the immune response in the fish. One function of these cells is to engulf microbes and then produce toxic molecules that kill microbes that are engulfed. In this study, three separate pathways that leukocytes utilize to produce these toxic molecules are being investigated. Experiments are in progress to determine the effect of CCV infection on each of these pathways. Data has been obtained that demonstrates that the production of a least one of these toxic molecules is inhibited by CCV infection. This suggests that fish that are carriers of the virus may be more susceptible to other infections. The role and potential application of a catfish antiviral gene in conferring protection against channel catfish virus (Peter Silverstein) In addition to the immune responses such as the generation of antibody, fish have genes that have antiviral properties. One such gene has been shown to confer some level of protection against CCV infection. Studies that are underway indicate that the expression of this gene in leukocytes can be dramatically enhanced by various stimulants. The goal of this project is to determine whether a similar enhancement can be obtained in fingerlings and whether this would confer resistance to CCV infection. Should protection be conferred the treatment could be incorporated into a fingerling production program in order to reduce CCV-associated mortality. The determination of the sensitivity of different catfish strains to channel catfish virus (Peter Silverstein and Al Camus) Although direct mortality due to CCV has not been thought to account for more than 5% of the annual losses in fingerlings outbreaks on individual farms can cause substantial losses. This project focuses on determining the relative mortality rates of different strains of channel catfish with regard to CCV infection. We hope to find catfish strains that exhibit significant resistance to CCV infection. These strains could be utilized in our breeding program to improve disease resistance. The development of an assay to determine the frequency of latent CCV infection in channel catfish broodstock (Peter Silverstein) At present the best available assay for CCV latency utilizes PCR analysis of a fin clip. Although the assay demonstrates that the fish was exposed to CCV at some time, it does not demonstrate that the fish that was sampled has a latent infection. For example, the viral DNA that was detected may not be capable of replication, the fin may be the only site of infection, etc. It should be noted that cells from fish fins are commonly used to grow and detect viruses. The goal of this project is to develop an assay using leukocytes that will enable a determination as to whether the animal in question has a latent infection that could be transmitted to offspring. Such an assay would be a valuable tool in developing CCV-free broodstock and thus lowering the incidence of vertical transmission of this virus. Detection of CCV carriers with a genetic detection assay (Lanie Bilodeau, Peter Silverstein, Larry Hanson, Michael Mauel, Al Camus) 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 25 both commercial farms and research populations were determined to be carriers of CCV. The incidence of CCV carriers was on average 10 to 50% of fish sampled. This information will be used for screening and assessment of CCV susceptibility in future projects. Investigation of a possible role for bullfrog tadpoles in the transmission of channel catfish virus (Al Camus, Peter Silverstein, and Lanie Bilodeau) For years, fish health researchers have postulated the existence of reservoirs for CCV outside the channel catfish. Additionally, rumors have persisted about a potential role for bullfrog tadpoles in the transmission of the virus to catfish fry and fingerlings. During a recent natural outbreak of CCV, tadpoles were noted to be dying in large numbers and CCV was isolated from one of the diseased animals. A series of challenge trials is currently being conducted to investigate whether twdpoles are susceptible to infection or are capable of transmitting the virus. Visceral Toxicosis of Catfish (VTC) Field investigations and epidemiology of visceral toxicosis of catfish and channel catfish anemia (Pat Gaunt, David Wise, and Al Camus) Field and laboratory studies have been conducted to determine the cause of two diseases with unknown etiology. Visceral toxicosis of catfish (VTC) disease is caused by an unknown toxin present in the blood of infected fish. The source of the toxin is unknown. Transmission studies indicate that VTC is noninfectious and most evidence suggests that the toxin is not water-borne. The epidemiology of this disease is currently being investigated to help identify commonality in management and environmental factors among farms that have experienced outbreaks. CCA is another disease of unknown etiology. The disease cannot be induced by the transfer of diseased tissues to healthy fish, co-habitation, or exposure of healthy fish to pond water during an epizootic. Naturally produced toxins are being investigated as a possible cause. Investigation of the etiology of Visceral Toxicosis of Catfish (Pat Gaunt, David Wise, and Al Camus) VTC is a disease seen primarily in market size and brooder fish. A possible cause is a potent, naturally produced toxin either ingested from fish feed, natural foods, or pond sediment. Experimental trials in tanks are being conducted to duplicate the disease and treat it. Ultimately, these tank studies will be used to define the cause of the disease in catfish ponds and investigate preventative techniques. Investigation of the cause of visceral toxicosis of catfish (VTC) and development of a rapid diagnostic test (Michael Mauel) Molecular and immunological techniques are being used to isolate and characterize the toxin responsible for VTC. Once the toxin has been identified, investigations will focus on determining its source. To date it has been found using immunological methods that catfish respond with antibodies to a specific component or antigen associated with VTC disease. An antibody ELISA test will be developed to provide rapid diagnosis of the disease and detect the presence of the toxin in pond water. Protective immune responses to sublethal toxin exposure will be evaluated in challenge trials. Ultimately, it is hoped that these studies will establish methods for preventing losses from VTC. 26 Identification of the organism(s) responsible for VTC syndrome (Paul Zimba, Al Camus, and Pat Gaunt) During cool-water periods, fish kills have routinely occurred in brood-fish and grow-out ponds. A common feature in fish gut contents is the presence of a filamentous blue-green alga called Planktothrix aghardii. Toxin analyses of fish gut contents and concentrated water and sediment samples have identified occasional spikes in anatoxin-a[s]. Continued efforts will be made to culture this alga and then dose fish to confirm preliminary results. Control and/or management recommendations for this benthic alga will then be possible assuming toxicity is demonstrated. Channel Catfish Anemia (CCA) Investigation of the role of iron deficiency in the pathogenesis of channel catfish anemia (Al Camus) On-going studies will confirm the results of preliminary trials that indicate fish affected with channel catfish anemia (CCA), also “white lip” and “no blood,” suffer from iron deficiency and recover rapidly following iron injection. The lifespan of the channel catfish erythrocyte is currently being determined to estimate the length of time required to deplete body iron stores and allow for removal of aged erythrocytes from circulation. Food-size catfish will then be fed an iron-deficient diet and monitored for signs of the disease. Should iron deficiency be proven to play role in the pathogenesis of CCA, investigations will focus on potential causes, including dietary inhibitors of iron uptake from feeds. Ultimately, these studies will establish means of avoiding losses from CCA. Investigation of the role of hepcidin in the development of channel catfish anemia. (Al Camus and Jishu Shi, Auburn University) Collaborative studies with researchers at Auburn University have shown that hepcidin, a key regulator of iron uptake from the intestine, is present in catfish and responsive to levels of stored iron in the body. Hepcidin levels were found to be decreased in catfish affected by CCA, indicating a normal physiologic response to anemia. These findings eliminate the possibility of a role for hepcidin in development of the disease. Feed-related anemia in channel catfish (Bruce Manning, David Wise, and Al Camus) Limited quantities of catfish grow-out feeds were purchased through commercial sources to conduct experiments on ways to improve the health status of juvenile catfish through nutritional intervention. During the course of the initial aquaria study, early recurring morbidity and mortality were encountered. Examination of dead and impaired fish revealed the presence of anemia symptoms as characterized by pale gills, white-lip syndrome, and depressed hematocrits. In many specimens, hematocrit values were less than 10% with some individuals as low as 2-5%. Summary of hematocrits from one experiment showed that approximately 14% of fish sampled had hematocrit values of 10% or less compared to none for catfish fed a standard reference diet. This condition is experimentally repeatable using the same feeds. The condition appears to mimic the characteristics of feed-related anemia that has been observed in pond-raised channel catfish. Research is continuing to determine the cause of the condition and ways to prevent it from occurring in pond-raised channel catfish. 27 Proliferative Gill Disease (PGD) Development of a risk-assessment model for proliferative gill disease (PGD) in channel catfish (David Wise, Al Camus, Terry Greenway, and Linda Pote) Studies were conducted to gain a better understanding of the prevalence, seasonal occurrence and severity of PGD in catfish production ponds and determine environmental factors that contribute to severe epizootics. Infection rates were monitored in commercial production systems by quantitatively grading the severity of gill pathology occurring in sentinel fish. Monitoring efforts indicated that fish in essentially all intensively managed production systems develop some degree of PGD and that the most severe manifestations of the disease occur during the spring when fish are initially stocked into the ponds. Laboratory and field studies indicate that severe manifestations of the disease are dependent on the rate spores are shed into the water and the susceptibility of the fish to infection. This information is being used to modify current management practices in efforts to reduce the impact of PGD. The protocol used in these studies appears to be an accurate method to quantitatively monitor infection rates and can be used to evaluate potential disease treatments and predict the occurrence of PGD related losses. It has been demonstrated on commercial operations that the use of these procedures can effectively eliminate PGD related losses in fish that are stocked in production ponds during the spring. Evaluation of biological control of Dero digitata, the oligochaete host of the parasite that causes PGD (David Wise and Charles Mischke) Investigations are currently underway to evaluate the use of fathead minnows and smallmouth buffalo as a biological control of Dero digitata worms, an oligochaete that is the intermediate host for the parasite that causes PGD. Monitoring PGD infection rates in commercial and experimental production ponds indicates that fathead minnows are not effective in controlling this disease. The effectiveness of smallmouth buffalo is currently being evaluated as a possible biological control species on three commercial catfish farms. Evaluation of chemical treatments to control Dero digitata, the oligochaete host of the parasite that causes PGD (David Wise and Charles Mischke) Approved chemical treatments commonly used in aquaculture have been evaluated as a method of controlling PGD by targeting Dero digitata worm populations or inactivating infectious spores, but appear to have limited application. The LC50 of most of the chemicals evaluated exceeds the rates that can be safely applied to ponds without having serious consequences to fish health. In laboratory trials the use of formalin and potassium permanganate caused a slight reduction in the pathology associated with PGD but were not shown to be therapeutic. The application of chemicals at rates that are lethal to fish can greatly reduce Dero digitata worm populations and may have some value as a preventative on farms that raise fish as single cohorts and completely harvest the pond before restocking. Development of molecular-based assays to study the proliferative gill disease (PGD) organism Henneguya ictaluri (Al Camus, Michael Mauel, and Linda Pote, MSU-CVM) A molecular-based “PCR” assay capable of quantifying numbers of infective PGD spores in pond water has been validated for specificity. An antibody-based capture system to extract spores from pond water is currently being developed. When completed, the assay will be applied in field trials to study the epidemiology of PGD, monitor disease activity in ponds, assess the efficacy of potential treatments, and identify life stages of the parasite in fish and the environment. Another technique, known as in-situ 28 hybridization will be used to determine where the parasite gains entry to the fish and how it migrates to the gills. This project is made possible by a graduate student stipend from the College of Veterinary Medicine to support the PhD studies of Matthew Griffin. Susceptibility of genetic strains and families of channel catfish to Edwardsiella ictaluri and PGD (David Wise, Terry Greenway, Brian Bosworth, and Lanie Bilodeau) Studies have been conducted to evaluate the response of genetic lines of channel catfish to bacterial and parasitic infections. Data from these studies demonstrated variation in susceptibility among lines and families of channel catfish to ESC and PGD, indicating considerable potential of increasing disease resistance through genetic selection. Trematodes Epidemiology and economic impact of the trematode Bolbophorus confusus in commercially raised channel catfish (David Wise, Carla Taylor, MSU-CVM, Terry Hanson, MSU, Ag Econ, and Charles Mishcke) Sampling protocols were established to determine presence and severity of trematode infections on commercial catfish farms. The number of ponds containing infected fish and the overall severity of the infection was evaluated and used to assess the potential impact of the disease. To date, of the 861 ponds sampled throughout 8 counties in the Mississippi Delta, 269 ponds contained trematode infected fish. The severity of infection ranged from mild to severe with most of the farms sampled testing positive for this disease. Farms that were unable to implement an effective bird management program and were within close proximity to pelican loafing sites were much more likely to suffer from severe infections. Trematode infections have resulted in severe losses as a result of mortality and decreased feed consumption and conversion. Economic analysis of infested farms has demonstrated that light to moderate infections decreased production by 27.1 percent. This decrease in production resulted in cash returns that were less than the variable cost of production. Evaluation of hydrated lime and copper sulfate to control snails in commercial catfish ponds (David Wise, Linda Pote, MSU-CVM, and Charles Mischke) Hydrated lime and copper sulfate shoreline treatments were shown effective in reducing snail populations that are present in the aquatic vegetation around the perimeter of the ponds. While effective, these treatments have little impact on snails that are not within close proximity to the pond bank. Uniform application of copper sulfate was evaluated as treatment against snails in catfish production ponds. This work demonstrated that copper sulfate applied as a solution across the pond surface to deliver between 2.5 and 5.0 ppm was effective in killing snails throughout the pond environment. On average pond treatments killed greater than 95% of the test snails, while mortality of control snails was less than 8.5%. Treatments were also shown to reduce the natural populations of snails along the pond margin by 98.3%. Development of an integrated pest management program to control snails in commercial catfish ponds (David Wise, Linda Pote, MSU-CVM, and Charles Mischke) There are currently two chemicals being used to control rams horn snails in commercial catfish ponds: hydrated lime and copper sulfate. This research project will focus on the development of other treatments to control snail populations. Economic comparisons of treatment cost for efficacious 29 treatments will be evaluated. In addition, this project will monitor how the treatments affect different sizes of snails within a population and recruitment (reproduction). The data will then be used to develop a practical pest management program, which can also serve as a model for snail control in other aquaculture systems. Evaluation of health, disease resistance, and growth potential of channel catfish infected with Bolbophorus confusus (David Wise, Terry Greenway, and Al Camus) Trials were conducted in experimental ponds to determine impact of trematode infections on catfish performance characteristics. Light trematode infections reduced feed consumption and overall production by approximately a 30%. These experimental results are comparable decreases in production observed on experimental farms. Aquaria studies demonstrated that once fish are removed from the source of infection that trematode infected fish have equal growth characteristics compared to non-infected fish. Data indicates that breaking the life cycle of the trematode is an effective cure for this disease and that the long term implication of the disease are negligible once the source of infection has been removed. Effect of sublethal exposure to the trematode Bolbophorus sp. on the severity of ESC in channel catfish (David Wise, Al Camus, and Terry Greenway) This study illustrated an experimental model describing the induction of low-level Bolbophorus sp. infection in channel catfish reproducing the pathological aspects of the disease. Experimental investigation of the interaction of E. ictaluri and Bolbophorus sp. infection demonstrated that concurrent exposure to both pathogens significantly increased mortality associated with ESC. However, no significant increase in mortality was observed when exposure to Bolbophorus sp. cercarieae occurred 28 days prior to E. ictaluri exposure. This data supports field observations and experimental growth data that the presence of the mature parasite in fish alone does not have a negative impact on fish health. Hatchery-Associated Disease Problems Optimization of hydrogen peroxide treatment of catfish eggs (Brian Small and Terry Bates) These studies were conducted in support of the approval of hydrogen peroxide by the US-FDA as a new animal drug for treating fish eggs, with the expectation that the label will include the treatment of catfish eggs. Results of this research showed that the effectiveness of hydrogen peroxide may be impacted by temperature. When hatchery water temperature was 78oF, a daily, 15 minute bath in a solution of 250 ppm active hydrogen peroxide (715 ppm of 35% hydrogen peroxide) was as effective as formalin at disinfecting eggs and improving hatch rates. Twice as much hydrogen peroxide at this temperature was found to be toxic to the eggs. At cooler temperatures, toxicity was reduced and higher concentrations of hydrogen peroxide were more effective. These studies established static bath and continuous-flow treatment recommendations for managing catfish egg diseases with hydrogen peroxide. Developing egg treatment recommendations for hybrid catfish eggs (Brian Small) The strip-spawning process required to make blue-channel catfish hybrids often results in a large number of unfertilized and dead eggs, which are prime targets for bacterial and fungal infections. These studies were conducted to identify the chemotherapeutant, respective concentration, and treatment methodology that yield the greatest hybrid catfish hatching success. Hatching success was highest among eggs treated 30 daily with a 15-minute bath of 100 ppm formalin when compared to similar treatments with copper sulfate, hydrogen peroxide, and povidone iodine. The optimal frequency of formalin treatments was determined to be three times daily. Together, these results identify a chemotherapeutic treatment regime for improving hybrid catfish hatching success by as much as 40%. 31