Optimization of Zebra Danio Performance

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Optimization of Zebra DanioPerformance
BY Dallas E. Weaver, Ph.D. Scientific
Hatcheries Huntington Beach, Ca
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The Devil is in the Details

• The biology of zebra fish interacts with the
  culture system, water chemistry, husbandry
  details and nutrition.

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Aquatic Animals vs. Land Animals

• Designing and operating zebra systems is a lot
  like trying to design and operate mice systems in
  zero G. You dont have gravity to separate the
  waste products from the animals.

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Performance

• Definition of performance
• Egg production and fertility
• Embryo development and hatching
• Larval survival and initial growth
• Growth rate
• Maturation -- egg to egg time

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Egg production and fertility

• Number of eggs per gm of female
• Fertilization percentage
• Cannibalism
• Egg health -- yoke size

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Factors impacting egg production

• Water quality
• Metabolic waste products -- non-detectable
• Ammonia, nitrite lt 0.05 ppm in tank input water
• Other chemicals
• Nitrate 0 to 75 ppm
• Salinity 100 to 2,000 ppm
• pH 6.0 to 8.0
• Temperature 20.5 ºC to 28.5 ºC
• Pushing the limit on one variable reduces the
  range of other variables.

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Factors impacting egg production

• Nutrition
• No real research into zebra maturation diets
• Quality aquaculture diets all have short shelf
  life problems. Age dating, inventory cycling.
• Omega-3 fatty acid requirements
• Vitamin requirements
• Water soluble (C, B, etc) -- leaching problem
• Oil soluble (E)

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Factors impacting egg production

• Nutrition practice by Scientific Hatcheries
• Dry diet of custom formulated modified salmon
  diet. High fishmeal protein, unsaturated fats
  (high omega-3 fats).
• Frozen adult artemia
• Live moina, rotifers, artemia for maximum
  production.

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Factors impacting egg production

• Cycling
• Observation that off cycle fish have low
  fertility.
• May be related to over-ripe eggs
• Density
• At densities gt 10,000 / M2 of spawning surface
  fertility decreases.

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Performance

• Definition of performance
• Egg production and fertility
• Embryo development and hatching
• Larval survival and initial growth
• Growth rate
• Maturation -- egg to egg time

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Embryo development

• Impacts on development
• Chemical stress from water quality
• Non-viable embryo problem -- static systems
• Toxins from materials
• Interaction between embryos may impact sex ratio
• Unstable microbiology -- bacterial/fungal attack
  on the embryos

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Embryo development

• Chemical stress from water quality
• Static systems
• Non-viable embryos source of pollutants
• Food supply for bacteria
• Ratio of volume per embryo critical
• Depth irrelevant
• Surface tension will be relevant at hatch
• Flowing system
• Water quality determined by source water.

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Toxins from materials

• Bioassay all materials
• Test materials with embryos and larva
• Problems materials
• Rubber -- black rubber
• Vinyl tubing, brass valves at low alkalinity
• Cleaning solutions and sterilization solutions
• Solutions
• Rinsing, leaching and materials selection
• Flowing water

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Embryo development

• Interaction between embryos may impact sex ratio
• Static systems can have interactions
• Many researchers have sex ratio problem
• Hypothesis developing embryos can release
  chemicals that can impact the development of
  other embryos exposed to those chemicals.

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Embryo development

• Unstable microbiology -- bacterial/fungal attack
  on the embryos
• Static systems -- dishes, beakers, etc.
• Dead embryos and non-viable embryos provide
  nutrients for bacteria and fungal growth
• Once fungal or bacterial attach on non-viable,
  attack on viable embryos is possible
• Flowing systems
• Upflow system -- not a problem
• Non-upflow -- fungal problems

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Embryo development

• System options
• Static systems
• All the problems
• Easier to work with embryos
• Fluidized Bed Hatchers
• Eliminates the problems of static system
• Automatic non-viable removal
• Automatic fungus egg removal

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Embryo Development

• What is a Fluidized bed Hatcher
• A fluidized column of glass bead media with a
  fluidized column of embryos behaving like a
  second phase fluid
• With a constant up-flow velocity, non-viable
  embryos swell up and float out of the hatcher.
• Every embryo has a constant supply of clean water
  which decreases developmental differences caused
  by oxygen differences in static systems.

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Performance

• Definition of performance
• Egg production and fertility
• Embryo development and hatching
• Larval survival and initial growth
• Growth rate
• Maturation -- egg to egg time

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Larval survival

• Ideal is 100 survival for all normal embryos
• Factors causing unnecessary mortality
• Water quality
• Nutrition
• Weak larva

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Larval survival

• Factors causing unnecessary mortality
• Water quality
• Normal pollutants, ammonia, nitrite and metabolic
  waste products, low oxygen
• Surface tension -- oil film -- swim bladder
  inflation
• Toxic chemicals
• Bacterial/protozoan explosion

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Larval survival

• Factors causing unnecessary mortality
• Water quality (continued)
• Surface tension -- oil film -- swim bladder
  inflation
• Zebra fish need surface access for filling the
  swim bladder.
• Common problem with other aquaculture species
  (striped bass, white bass, etc.)
• Solution is keeping the surface free of oil
• Tank depth is irrelevant.

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Larval survival

• Factors causing unnecessary mortality
• Water quality
• Nutrition
• Weak larva

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Larval survival

• Factors causing unnecessary mortality
• Nutrition
• First food is critical
• Notes from Aquaculture
• Larval fish/shrimp production is a multi-billion
  business
• Dry diets cant replace 100 of live feeds for
  all small larva species. Dry diets are only
  supplements to cut the cost of live feeds.
• When dry diets appear to work, it is often the
  bacterial/protozoan contamination of that diet
  that allows it to work.
• Leaching of critical nutrients from dry diets is
  very fast

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Larval survival

• Factors causing unnecessary mortality
• Nutrition (continued)
• Notes from Aquaculture (continued)
• Rotifers (brackish water species) is the most
  common small live feed. For species where new
  hatched artemia is too large.
• Rotifers are often enriched before feeding with
  algae or prepared omega-3 fat blends.
• Rotifers can be packed with antibiotics or other
  chemicals.
• Can be purchased alive or produced locally.

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Larval survival

• Factors causing unnecessary mortality
• Nutrition (continued)
• Notes from Aquaculture (continued)
• Artemia napulii
• Easy to hatch
• Nutrition value decreases with time when un-fed
• Nutrition value variable
• Enrichment is a common practice -- improved fatty
  acid profiles and amino acid profiles are
  possible
• Digestive enzymes are limited in many larval fish
• Depend upon live food to provide

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Larval survival

• Factors causing unnecessary mortality
• Water quality
• Nutrition
• Weak larva

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Larval survival

• Factors causing unnecessary mortality
• Weak Larva
• Can be caused by poor adult nutrition
• Stressing the larva during development
• Stressing the larva with high density
• Social interaction problem
• Too much turbulence wasting energy reserves
• A problem with McDonald jars for hatching

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Performance

• Definition of performance
• Egg production and fertility
• Embryo development and hatching
• Larval survival and initial growth
• Growth rate
• Maturation -- egg to egg time

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Growth rate

• Factors impacting growth rates
• Temperature
• Water quality
• Nutrition
• Density

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Growth rate

• Factors impacting growth rates
• Temperature
• There is a maximum above which metabolism is
  greater than digestible feed consumption.
• With a low quality diet, the rearing temperature
  could be above the maximum.
• Apparently parameters are not available for a
  zebra fish bio-energetic model.

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Growth rate

• Factors impacting growth rates
• Temperature
• Water quality
• Nutrition
• Density

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Growth rate

• Factors impacting growth rates
• Water quality
• Low Oxygen levels decrease growth
• Very high unionized ammonia levels
• gt 20 ppb unionized
• Pheromones give an apparent density dependence
  that is not real.

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Growth rate

• Factors impacting growth rates
• Temperature
• Water quality
• Nutrition
• Density

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Growth rate

• Factors impacting growth rates
• Nutrition
• This is a critical element
• Live feeds are the best
• Enrichment is desired
• All they can eat all the time
• 7 days a week
• Excess feed and old feed may decrease growth rate
  and support enteric diseases.

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Growth rate

• Factors impacting growth rates
• Temperature
• Water quality
• Nutrition
• Density

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Growth rate

• Factors impacting growth rates
• Density
• Often misinterpretation of observations
• Apparent impacts from water quality
• Apparent impacts from nutrition
• Real impacts result from social interaction
• Real impact density gt 35/liter for zebra fish or
  1.5 zebra fish in the water
• Below those levels, apparent density impacts of
  slow growth, growth variation, etc. are artifacts
  of other problems.

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Performance

• Definition of performance
• Egg production and fertility
• Embryo development and hatching
• Larval survival and initial growth
• Growth rate
• Maturation -- egg to egg time

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Maturation -- egg to egg time

• Putting it all together and adapting commercial
  aquaculture technology to zebra fish
• Use a standard Aquaneering FBB rack system
• Use standard zebra research artemia hatching tank
  -- tank for automated feeding
• Use de-chlorinated tap water at 28.5 ºC

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Maturation -- egg to egg time

• Special additions
• Fluidized Bed Hatchers for development
• Black tanks for experiment on tank color
  (significant for many species of fish)
• Peristaltic feed pump -- automated feeding
• Timer, rotifer supply, artemia supply (enriched)
• Inlet diffuser for mixing feed with main flow
• Turbulent flow control valves on each tank
• Fine discharge screen baffles -- 300 µ

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Maturation -- egg to egg time

• Additions (continued)
• Air injection behind screen (improved oxygen
  transfer without excessive turbulence)
• Monitoring of salinity -- live food additions
  were with brackish water

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Maturation -- egg to egg time

• The experiment
• Four clear tank, Four Black, Four density classes

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ResultsEGG to EGG 49 days
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