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2008 Tracy Research Study Plan
Holding-duration-associated Impacts of Salvaged Fishes at South Delta Fish Collection Facilities
Principle Investigator
Donald E. Portz, Fisheries Biologist, Fisheries & Wildlife Resources Group, Bureau of Reclamation, dportz@do.usbr.gov
Problem Statement
Population declines of several fish species throughout the west may threaten to disrupt water delivery systems. Water diversions are suspected of being one of the primary causes for the losses of fishes through entrainment (Brown and Moyle 1993, Bennett and Moyle 1996). California water officials halted water exports from the Sacramento-San Joaquin Delta in the spring of 2007 as a direct result of a lawsuit after rising numbers of endangered fish were thought to be killed. These federally and State-listed threatened and endangered fish species, and those of concern because of an economically viable sport fishery or listed as a POD species (Pelagic Organism Decline), are frequently recovered during salvage operations. Water is vital to the growth and prosperity of the American West and recent lawsuits from the California Department Fish & Game and environmental groups have dealt major blows to Delta water export. Water resource agencies will have to comply to tougher environmental standards in the near future and will have to lessen their impact on fish and wildlife, while providing water for agriculture and municipalities.
Operations of Reclamation facilities at water diversions and dams require proper screening, passage, capture, and handling of fish. These functions are of major importance for the survival of fishes impacted by Reclamation; however handling stress associated with entrainment may inadvertently harm the fish that Reclamation is attempting to save. Exposure of fishes to environmental stressors, such as capture and handling, can be a great concern to fisheries biologists, in that extreme or prolonged stressors may plague fish performance and overall health, adversely affecting population size and sustainability. Measuring the physiological stress and potential direct and indirect mortality experienced by fishes during the different components of Reclamation operations is vital to understanding its negative impacts. Results of this proposed study and one we are currently finishing will provide information on methods to reduce the harmful effects of entrainment and handling, leading to a reduction in the incidental take of fishes and a reduction of direct and indirect mortality from sublethal stressors. The literature on the physiological effects of capture and handling stress on entrained or salvaged fish species through daily operations at Reclamation and other water resource agencies’ facilities is scant and needs to receive more research attention.
Evaluations and improvements of both the State and Federal fish salvage facilities have been ongoing for a number of years, though emphasis has been more on the facilities themselves rather than the operations for handling, transporting, and release of fishes (Liston et al. 2000). Efforts by California Department of Fish and Game have demonstrated problems with survival of salvaged fish after transport to the release site and Reclamation researchers have contributed to the understanding of survivorship and injury of fishes associated with the collecting/holding tanks at TFCF (Raquel 1989, Karp and Lyons 2007). These studies, and ongoing technological advances in concepts related to fish holding, indicate an important need for accelerating and expanding studies at the two salvage facilities. Research and monitoring plans have been instituted to investigate the consequences that these water diversions are inflicting on Sacramento-San Joaquin River Delta fish fauna and ways to minimize their negative impacts.
Study Summary
Over the past three years, we have been studying the deleterious effects of the fish salvage process at the Tracy Fish Collection Facility (TFCF; Tracy, California) and in Reclamation’s Hydraulics Laboratory (Denver, Colorado). We have examined holding tank design and fish removal and transfer methods, and how they relate to acute physiological stresses and potential direct and indirect mortality experienced by salvaged fishes (Portz et al. 2006). The next logical step and aim of our proposed study for FY 2008 is to examine the changes in the stress physiology of salvaged fishes during (and throughout) a normal holding period before they are removed to be transported back to the river for release. This will be accomplished by comparing changes in plasma constituents attributable to capture and handling stress of striped bass (Morone saxatilis) and green sturgeon (Acipenser medirostris) and how the concentrations of the plasma constituents changes over a holding duration. The research question being asked is how does the holding duration physiologically affect striped bass and green sturgeon (ca. 300 mm TL)?
Taking blood specimens from fish during physiological studies typically involves netting, handling, inducing anesthesia, and withdrawing a blood sample by syringe. Although simple and rapid, this procedure can result in a stress response by the fish and would limit repetitive blood sampling from the same individual, and removal of one animal can induce a stress response in other individuals in the same tank. Pickering et al. (1982) found that the sequence in which several fish from one tank were sampled had a significant influence on plasma cortisol levels. In addition, single measurements of plasma cortisol are merely instantaneous snapshots of a much larger dynamic process over time. The use of indwelling catheters is recommended to minimize disturbances during sampling (Heisler 1984) and gain insight to the physiological changes throughout fishes’ holding durations.Objectives
The objectives of this study are: 1) to determine if stress exhibited in held fish is affected by the duration detained, 2) to evaluate dorsal aorta cannulation as a viable alternative to handling, netting, and anesthetizing fish for blood sampling during salvage processes, and 3) to evaluate serial blood sampling as a technique to determine physiological stress responses in fish over time.
Hypothesis
We hypothesize that the cylindrical collecting/holding tanks at salvage facilities in the south Delta significantly alter stress-related blood constituents. Stress exhibited in held fish is affected by the duration detained; fish collected and detained for extended periods of time (e.g., >6 h, but <48 h) will show plasma constituents returning to basal levels during the holding period. Arguably, it might be beneficial to hold fishes for longer periods of time during salvage practices to provide them with an opportunity to reduce the effects of stress after being louvered and diverted into holding tanks and before loading them in tanker trucks for transport to release sites. In order to hold fish longer the TFCF would have to 1) remove stressors from the holding tanks, and 2) obtain a new Biological Opinion (Fish and Wildlife Service with concurrence from California State Department of Fish and Game) to allow a holding period longer than 8 hours for delta smelt and 12 hours for Chinook salmon. Results from this study and those from prior research focusing on holding-associated stress of Chinook salmon (Oncorhynchus tshawytscha) may be useful for extending holding periods for other species of interest.
Materials and Methods
Source and Care of Fish:
Striped bass (Morone saxatilis) and white catfish (Ameiurus catus; ca. 300 mm TL) will be obtained from the Tracy Fish Collection Facility (TFCF; Tracy, California). Fish will be cannulated and allowed 24 hours to recover from the handling stress associated with inserting the cannulae prior to experiments. Striped bass and white cat fish will be used in this study because Chinook salmon (Oncorhynchus tshawytscha) and delta smelt (Hypomesus transpacificus) entrained at the TFCF are too small for cannulation and serial blood sampling. Studies in Denver will require fish to be transported and held in the Fish and Wildlife Resources Group Aquaculture Facility. Experimental fish will be maintained in 757-l circular tanks equipped with an aerated, partial recirculating water system to deliver water continuously along with dechlorinated, air-equilibrated municipal water. Water temperatures will be maintained at 18˚C. Fish will be held under a natural photoperiod (38° N latitude) and supplemented with halogen lights, and were fed BioOregon (BioOregon Inc., Longview, Washington) semi-moist pellets at 1.5-2% body weight per day or live minnows.
Cannulation Procedure:
Striped bass and channel catfish will be cannulated in the dorsal aorta. Fish will be anesthetized with a 200 mg/l tricaine methanesulfonate solution (MS-222, Argent Chemical Laboratories Inc., Redmount, WA) and placed on a V-shape table. Anesthesia is maintained and gills continuously irrigated with a MS-222 solution (100 mg/l) during the operation at a rate of 1,500 ml/min using a submersible pump. A 75-mm long, 17 gauge, hypodermic needle is inserted through the skin and musculature to the hemal canal and a 200-cm-long cannula (Clay Adams PE-50 tubing, Division of Becton Dickinson, Parsippany, NJ) will be inserted through the needle into the dorsal aorta (non-occlusive cannulation). The cannula tubing will be filled with heparinized saline and sutured (Ethicon 000 silk) to the skin at the point of entry at the lateral aspect of the body wall. A three-way stopcock is attached to the distal end of the cannula, which enables undiluted blood samples to be collected by syringe. The cannula is refilled with heparinized saline after each blood sample. Surgical placement of the cannula and recovery from anesthesia will be completed within 10 min. Rapid (<5 min) recovery will be initiated by switching artificial ventilation to aerated, anesthetic free water. Once ventilatory activity returns, the fish will be placed into a 757-l recovery tank receiving a continuous flow of aerated, dechlorinated water. Fish will be allowed 24 hours to recover and allow stress-related plasma constituents to return to pre-handling levels.
The Experiment: Effects of holding duration on the stress response salvaged fishes
Tests are to be performed at both the TFCF and the laboratory simulation TFCF tank in Denver). Three cannulated fish are released in the collecting/holding tank. A float is attached to the distal end of the cannula with the stopcock. The float is hooked and slowly maneuvered where a blood sample may be taken without disturbing the fish. Blood samples (0.5 ml) will be collected via the cannula at intervals of 0, 1, 2, 4, 8, 12, 24, and 48h. A control fish remaining in the 757-l tank is bled (0.5 ml) at 0, 24, and 48h. Not all samples are drawn from every fish. No more than three blood samples are withdrawn from any one fish. This protocol ensures blood hemoglobin concentration which is directly proportional to intracellular buffer capacity (Davenport 1974, Crocker and Cech 1998).
Plasma Analysis:
Blood is immediately centrifuged (Clay-Adams Autocrit Ultra3, Becton Dickinson Diagnostics, Sparks, Maryland) for 4 min at 12,000 x g to separate the plasma from the packed cells. Plasma obtained from each fish is transferred into plastic cryogenic freezing vials and temporarily stored in a 10-l liquid-nitrogen dewar flask (-196˚C). These samples are then transferred to a -80ºC freezer for storage and later analyses. Plasma is later thawed for plasma cortisol, lactate, and glucose measurements. Plasma cortisol concentrations are measured using a modified enzyme immunoassay (ELISA; Munro and Stabenfeldt 1984, Munro and Stabenfeldt 1985, Barry et al. 1993). Plasma lactate and glucose are measured with a polarographic analyzer (YSI 2700 Select, Yellow Springs Incorporated, Yellow Springs, Ohio).
Data Analyses:
Statistical analyses will be performed using SAS 9.1 (SAS Institute Inc., Cary, North Carolina) and Sigmastat 3.0 (Jandel Scientific, San Rafael, California) statistical software packages. Differences between treatments and controls are tested using an unbalanced 2x9 factorial design random complete block design (RCBD) analysis of variance (ANOVA) with each group of the 9 durations constituting a block, blocks nested within hours with hours fixed (Steel et al. 1997). Plasma constituent data will be analyzed either by using RCBD factorial analysis of variance, one-way analysis of variance, or two-way analysis of variance with durations as a factor (Zar 1984, Steel et al. 1997). The Tukey’s test will be used for all pair-wise multiple comparisons for parametric data. The Shapiro-Wilk’s test for normality and the Levene’s test for homogeneity of variances are used to determine ANOVA assumptions. Data that does not meet the assumptions of ANOVA and were unable to be power or log transformed will be compared using a Kruskal-Wallis non-parametric analysis of variance on ranks with the Dunn’s test for pairwise multiple comparisons (Zar 1984, Steel et al. 1997). Differences were considered significant at P < 0.05.
Assumptions/Study Limitations
Access to collecting/holding tanks on-site at TFCF poses a significant obstacle during final stages of construction. In addiction, maintaining a swirl for up to 48 hours may be difficult to accomplish due to logistical limitations, presence of species of special concern, and incoming high fish densities. The logical alternative might be the Denver Hydraulics Laboratory and is a well-equipped research facility. However laboratory experiments are meant to simulate field conditions as closely as possible while maintaining additional control of environmental variables. Needless to say, it is impossible to completely duplicate all conditions that are experienced on-site.
Significance
Water diversions have been implicated as one of the primary causes of decline in California’s fish fauna (Stevens et al. 1985, Bennet and Moyle 1996). Current information on the effects of the capturing and holding processes on the survival of threatened and endangered species is needed to improve their survival during this process. Results of this study will provide information on methods to reduce the harmful effects of entrainment and holding, leading to a reduction in the incidental take of fishes and a reduction of indirect mortality from sublethal stressors. Loss of fishes due to entrainment into water diversions is a problem throughout the world. Refined fish screening methods and improved holding practices implemented at the TFCF will lead fisheries biologists and engineers to make substantial advances in the reduction of entrainment and successful fish salvage at other water diversions. Our study will provide valuable information to refine fish salvage methods in a cost effective manner, while providing methods to lower O&M costs.
Coordination and Collaboration
This study is a continuation of a collaborative effort between US Bureau of Reclamation, California Department of Fish and Game, and California Department of Water Resources. We would like to link these experiments to research currently underway by the Collection, Handling, Transport, and Release (CHTR) studies. This project lends an excellent opportunity for enhancing cooperative interagency efforts, leading towards minimizing salvage-facilities-associated fish mortalities, in the future. Regularly scheduled meetings (TTAT and CHTR), at least once a month, will continue to be held to facilitate interagency coordination of research activities. This study will be coordinated with the Fisheries and Wildlife Resources Group, Denver Water Resources Laboratory, TFCF Staff, California Fish and Game, California Department of Water Resources, other researchers, and the interagency Tracy Technical Advisory Team.
Resources and Capabilities
The Fisheries and Wildlife Resources Group provides a diverse range of expertise that includes staff with knowledge of fish biology, physiology, and aquaculture. The Denver Federal Center has excellent research facilities and space needed to study physiological stress associated with holding duration in collecting/holding tank. The Denver Hydraulics Laboratory also contains a replica TFCF collecting/holding tank available for experiments formerly used by other fish physiological stress studies.
Endangered Species Concerns
This study will not involve handling or capturing endangered or threatened species. Species used in this study will include fishes commonly entrained and of special concern to the TFCF: striped bass and white catfish. Neither fish is listed as threatened or endangered pursuant to both the Federal Endangered Species Act and California Endangered Species Act. Applicable State and Federal permits will be (have been) obtained to conduct research with these species (California Scientific Collecting Permit #801159-05, Colorado Importation License and Aquaculture Permit # 051MPT014). No fish will be sacrificed, intentionally, during this study, and the fish used in the experiments may be released after such use if allowable by law.
Dissemination of Results (Deliverables and Outcomes)
The primary deliverable from this proposed study will be articles published in both the Tracy Volume Series and a peer-reviewed scientific journal. Technical updates may also be provided at TTAT/TFRED and CHTR meetings, along with posters and oral presentations given at scientific forums in late summer/fall 2008. Additionally, information will be gained on the successes and limitations of the fish collection and salvage process at the TFCF. This knowledge will help guide future improvements in the fish collection, holding, and transport process.
Literature Cited
Barry, T.P., Lapp, A.F., Kayes, T.B., and Malison, J.A. (1993). Validation of a microtitre plate ELISA for measuring cortisol in fish and comparison of stress responses of rainbow trout (Oncorhynchus mykiss) and lake trout (Salvelinus namaycush). Aquaculture, 117, 351-363.
Bennett, W.A. and Moyle, P.B. (1996). Where have all the fishes gone? Interactive factors producing fish declines in the Sacramento-San Joaquin Estuary. In J.T. Hollibaugh (ed), San Francisco Bay: The Ecosystem (pp. 519-542). San Francisco: American Association for the Advancement of Science, Pacific Division.
Brown, L.R. and Moyle, P.B. (1993). Distribution, ecology, and status of the fishes of the San Joaquin River Drainage, California. Calif. Fish Game, 79, 96-114.
Crocker, C.E. and Cech, Jr., J.J. (1998). Effects of hypercapnia on blood-gas and acid-base status in the white sturgeon, Acipenser transmontanus. J. Comp. Physiol., B, 168, 50-60.
Davenport, H.W. (1974). The ABC of acid-base chemistry. (6th ed.) Chicago: The University of Chicago Press.
Heisler, N. (1984). Acid-base regulation in fishes. In W.S. Hoar and D.J. Randall (eds.), Fish Physiology, Vol. 10 (pp. 315-401). London: Academic Press.
Karp, C. and Lyons, J. (2007). Fish Holding Evaluation at the Tracy Fish Collection Facility. Tracy Fish Collection Facility Studies, Vol. 37 (17 pp). U.S. Bureau of Reclamation, Mid-Pacific Region and Denver Technical Service Center.
Liston, C.R., Christensen, R., Mefford, B., Glickman, A., and Johnson, P. (2000). Feasibility Report Number 2: A proposed technology development facility to support improvement and/or replacement of fish salvage facilities at Tracy and other large fish screening sites in the Sacramento-San Joaquin Delta, California. Bureau of Reclamation, Denver Technical Service Center and Mid-Pacific Region, Denver, CO, and Sacramento, CA.
Munro, C. and Stabenfeldt, G. (1984). Development of a microtiter plate enzyme immunoassay for the determination of progesterone. J. Endocr. 101, 41-49.
Munro, C. and Stabenfeldt, G. (1985). Development of a cortisol enzyme immunoassay in plasma. Clin. Chem. 31, 956.
Pickering, A.D., Pottinger, T.G., and Christie, P. (1982). Recovery of the brown trout, Salmo trutta L., from acute handling stress: a time-course study. J. Fish Biol. 20, 229-244.
Portz, D.E., Woodley, C.M., and Cech, Jr., J.J. (2006). Stress-Associated Impacts of Short-Term Holding on Fishes. Rev. Fish Biol. Fish. 16, 125-170.
Raquel, P. (1989). Effects of handling and trucking on Chinook salmon, striped bass, American shad, steelhead trout, threadfin shad, and white catfish salvaged at the John E. Skinner Delta Fish Protection Facility. California, Department of Fish and Game, Interagency Ecological Study Program for the Sacramento-San Joaquin Estuary, Technical Report 19.
Steel, R.G.D., Torrie, J.H., and Dickey, D.A. (1997). Principles and procedures of statistics: a biometrical approach. (3rd ed.). New York: McGraw-Hill.
Stevens, D.E., Kohlhorst, D.W., Miller, L.W., and Kelley, D.W. (1985). The decline of striped bass in the Sacramento-San Joaquin Estuary, California. Trans. Am. Fish. Soc. 114, 12-30.
Zar, J.H. (1984). Biostatistical Analysis. Toronto: Prentice-Hall.Updated December 16, 2007