Containment of Transgenic and Exotic Fishes at the University of Minnesota Fisheries and Aquaculture Laboratory
July 11, 1995
Fish Sizes Two fish sizes are distinguished for the purposes of containment. Effluents from the facility can take one of two routes, the sanitary sewer or the storm sewer system. The largest screen preventing fish from leaving the facility via the storm sewer is a wedge-wire screen with a 1 mm opening. Therefore, only fish with a minimum head diameter of 2 mm will be held in the part of the system connected to the storm sewer. Drainage from tanks containing smaller fish will be directed to the sanitary sewer after passing through filters whose mesh size is as small as is needed to contain the fish.
Personnel Access Facility doors are locked when authorized personnel are not present. Authorization is given only by the department head, Dr. Ira Adelman. A list of authorized personnel is kept in Dr. Adelman's office.
Fish Containment Tanks Each tank is covered with a weighted lid. Lids are checked daily after tank cleaning, and also by the last person leaving at the end of the day. Each tank's standpipe is covered with a screen that will prevent the smallest fish from escaping. For embryos and fish less than 2 cm total length (equivalent to 2 mm minimum head diameter for the fathead minnow, Pimephales promelas), effluent water will be filtered through a sock filter (0.3- 0.8 mm mesh depending on species being incubated) external to the tank. This filter will remove any fish less than 2 cm which may have escaped the holding tank into the effluent water. The sock filter (Figure 1) will be changed and cleaned daily. (Filtrate collected in the sock filter will be disposed of in a manner described in the Fish/embryo/siphonate Disposal section, page 5.) Embryo incubators Each incubator tray is covered with a screened lid that locks into place. The effluent from each incubator will pass through the sock filter. Drains Floor drains have either a round or square opening. Round openings are 20 cm in diameter and are covered with a cast iron screen (1.2 cm between bars). The square opening is 45 x 45 cm and is also covered by a cast iron screen (1.5 cm between bars). These drains are located 1-2 m away from the nearest fish rearing tank. The floor drains are part of the sanitary sewer system.
Water Flow Pattern Fish shorter than 2 cm Water from either the flow-through or recirculating system will enter a tank containing the fish, pass through the standpipe/tank floor screen, enter the sock filter, empty into a floor drain and leave the laboratory through the sanitary sewer system (see Note 1 below).
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Figure 1. Larval fish/embryo trap sock filter. Any embryos or small fish leaving an incubator or rearing tank are removed from tank effluent with a 0.3 mm mesh net. An overflow net will filter the effluent if the lower net should become occluded.
Fish longer than 2 cm If water is from the flow-through system, it will enter a tank, pass through the standpipe/floor screen, drain into the sump, be filtered through the wedge wire screens (see Note 2), and leave the laboratory through the storm sewer system. If water is from the recirculating system, it will enter a tank, pass through the standpipe/floor screen, be filtered through the rotating drum microscreen (see Note 3), be pumped through the biofilter, pass through the ozonator and UV radiator and re-enter the tanks. Sludge leaving the rotating drum microscreen will enter the sanitary sewer system.
Notes 1 - Water leaving the laboratory enters the St. Paul sanitary sewer system. Near the junction of highways 280 & 94 an overflow weir allows water from the sanitary sewer system to enter the storm water drainage system during high water runoff events. The storm water drainage system directs water to the Mississippi River. In 1993, a particularly wet year, the sanitary sewer system overflowed into the storm water drainage system 4 times with a duration ranging from 15-90 minutes and an average volume ranging from 14,900-660,000 gallons. 2 - Wedge-wire screens will be vacuumed daily and more often if necessary. If the wedge-wire screens plug, the water will pass through a basket strainer (1 mm) and be discharged through the sanitary sewer system. A high water alarm will signal this event. 3 - The mesh size on the rotating drum microscreen is 0.05 mm.
Cleaning Protocols Tanks (fish longer than 2 cm) Daily maintenance and bimonthly tank cleaning 1) Lift up or remove tank cover (if needed). 2) Remove any dead/dying fish from the tank, record in mortality log, and dispose of them. (See Fish/embryo/siphonate Disposal section.) 3) Scrub standpipe/tank screens and floor with a brush. Scrub tank walls during bimonthly tank cleaning. 4) To partially drain the tank and then replace standpipe, use one of the following procedures: a) Flow-through system tank with an internal standpipe--Make sure the outer standpipe screen has no holes or gaps and is firmly secured to the standpipe. Making sure the outer standpipe is tightly sealed with the bottom of the tank, lift up the inner standpipe. Replace inner standpipe making sure it and the outer standpipe are securely in place. tank with an external standpipe--Clean the screen in the floor of the tank. Make sure floor screen has no holes and is securely sealed to the floor. Lift up and then replace external standpipe making sure it is securely in place. b)Recirculating system tank with an internal standpipe---Make sure the outer standpipe screen integrity and stability are good. Making sure the outer standpipe is tightly sealed with the bottom of the tank, siphon food and wastes from the bottom of the tank. Dispose of siphonate into the slurry- kill tank. (See Fish/embryo/siphonate Disposal section.) Lift up and then replace the inner standpipe making sure it and the outer standpipe are securely in place. tank with an external standpipe---Clean the screen in the floor of the tank. Make sure floor screen integrity and stability are good. Lift up and then replace external standpipe making sure it is securely in place. 5) Lift up and then return tank cover and check for a secure fit. Post-experiment cleaning 1) Remove all fish. 2) Before draining the tank, confirm that the outer standpipe or floor screen (for tanks with internal or external standpipe, respectively) has no holes in the screen, no gaps in the sealed edge of the screen, and is securely sealed with the bottom of the tank. 3) Drain tank by removing inner standpipe or external standpipe (for tanks with an internal or external standpipe respectively) only. 4) For tanks with an internal standpipe, once the tank is completely drained, replace inner standpipe and remove outer standpipe. Remove any fish still in the tank. 5) Clean tank. 6) Replace standpipe(s) when finished. 7) Cover empty tank if transgenic or exotic fish are in an adjacent tank.
Tanks (fish shorter than 2 cm) Daily maintenance and bimonthly tank cleaning 1) Lift up or remove tank cover (if needed). 2) Remove any dead/dying fish from the tank, record in mortality log, and dispose of them. (See Fish/embryo/siphonate Disposal section.) 3) Scrub standpipe/tank screens and floor with a brush. Scrub tank walls during bimonthly tank cleaning. 4) Make sure outer standpipe screen integrity and stability are good. Making sure outer standpipe is tightly sealed with the bottom of the tank, siphon food and wastes from the bottom of the tank. Dispose of siphonate into the slurry-kill tank. (See Fish/embryo/siphonate Disposal section.) Lift up and then replace inner standpipe making sure it and the other standpipe are securely in place. 5) Clean and change filter socks in larval fish trap. Clean trap effluent holes. 6) Return tank cover checking for a secure fit. Post-experiment cleaning 1) Remove all fish. 2) Before draining the tank, confirm that the outer standpipe or floor screen (for tanks with internal or external standpipe, respectively) has no holes in the screen, no gaps in the sealed edge of the screen, and is securely sealed with the bottom of the tank. 3) Drain tank by removing inner standpipe or external standpipe (for tanks with an internal or external standpipe respectively) only. 4) For tanks with an internal standpipe, once the tank is completely drained, replace inner standpipe and remove outer standpipe. Remove any fish still in the tank. 5) Clean tank. 6) Thoroughly clean larval fish trap. 7) Replace standpipe(s) when finished. 8) Cover empty tank if transgenic or exotic fish are in an adjacent tank. Embryo incubators Daily maintenance 1) Remove dead/dying embryos and larval fish, record notes in mortality log, and dispose of dead fish. (See Fish/embryo/siphonate Disposal section.) 2) Clean and change filter socks in larval fish trap. 3) Make sure tray screen has no holes, no gaps in its sealed edge, and is securely fastened within the surrounding frame. Removal of dead embryos 1) Pull out tray to remove it from water current. 2) Drain half of the water in the tray before removing lid or tray from the stack. 3) Remove tray from the stack. 4) After mechanical shocking of the embryos (causes dead embryos to become opaque), remove dead embryos making sure no live or dead ones are dropped outside of the tray or dead-embryo receptacle. 5) Replace the lid, locking it into place, and return tray to incubator. Post-experiment cleaning 1) Remove dead/dying embryos and fry from the incubation tray and dispose of them. (See Fish/embryo/siphonate Disposal section.) 2) Clean tray(s) and cover thoroughly. 3) Thoroughly clean larval fish trap. 4) Return tray(s) to incubator.
Fish/embryo/siphonate Disposal To ensure 100% mortality, place fish, embryos, filtrate, and siphonate (a.k.a. "materials") into a slurry-kill tank (a 200 ppm chlorine bath) and later freeze materials for ò 24 hr. Disposal of transgenic fish materials (-> will send to incinerator) 1) Place materials into the portion of the slurry - kill tank labeled "transgenic fish". Estimate 30% of the volume added to the tank, and add this much bleach to the slurry-kill tank to maintain chlorine concentration. 2) Once enough material has been accumulated in the slurry-kill tank, separate the material from the slurry and move it to the freezer for ò 24 hr. Material should be held in an orange bag with "biohazard" printed on the side. These bags can be purchased from University Stores; they alert UMN Waste Management that the contents are to be incinerated. 3) Once enough material has been accumulated in the freezer, Facilities Support should be contacted at 625-7361 to organize collection of the waste. Facilities Support should be contacted at least 1 day in advance of pick up. They collect waste Tuesday through Saturday, usually early in the morning.
Disposal of non-transgenic fish materials (-> will send to rendering plant) 1) Place materials into the portion of the slurry - kill tank labeled "non-transgenic fish". Estimate 30% of the volume added to the tank, and add this much bleach to the slurry-kill tank to maintain chlorine concentration. 2) Once enough material has been accumulated in the slurry-kill tank, separate the material from the slurry and move it to the freezer for ò 24 hr. Material can be held in any convenient, sturdy bag. The bag should be strong because the frozen material will have to be separated from the bag when the material is deposited in the infectious waste disposal truck. 3) Once enough material has been accumulated in the freezer, the material should be deposited in the infectious waste disposal truck at one of the UMN Infectious Waste Docks (Figure 2). Call Ron Joki at 625-0255 to learn where the infectious waste disposal truck is located that day. Go to the dock, separate the waste material from the holding container, and place the waste inside the truck.
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Figure 2. Locations (docks) where infectious waste disposal truck may be located on the UMN St. Paul campus.
Accidental fish release protocol
Fish may accidentally escape from a tank, due to hole(s) in a screen, accidental standpipe removal, tank overflow, or fish jumping out of the tank. If fish are on the floor collect them and return them to the tank they escaped from (if known), otherwise, euthanize them, and place the bodies in the slurry-kill tank. If fish are in a single or common effluent line, they should be flushed from the line and collected from the effluent sump using the following procedure:
Flow-through system 1) Turn water flow off to effluent line. 2) Pour bleach down each access to effluent line starting with the access furthest "upstream" in order to kill and/or force the fish down the line to the effleunt sump. 3) Collect fish from the effluent sump area with a dip net or the wet vacuum.
Recirculating system 1) Call Jay Maher (home: 729-4611, pager: 538-5302).
Notify the Institutional Biosafety Committee in the event of any unfiltered release of embryos, fry, or fish from the facility. Fisheries and Aquaculture Facility - key holders
1. plumber 2. J. Maher 3. J. Maher 4. J. Maher 5. P. Sorenson 6. I. Adelman 7. A. Kapuscinski 8. C. Markee 9. Olson 10. P. Maniak 11. Phillips 12. C. Appelt 13. M. Hove 14. ? 15. D. Brister 16. Facilities Management (Nordia 5-3791) 17. "..." 18. "..." 19. Y. Cai 20. K. Kalina 21. ? 22. ? 23. ? 24. ? 25. R. Bjerselius 26. J. Grant
Date of last revision June 1995