Fish Nutrition Research
Differences and similarities with
livestock nutrition and what the
future holds. Part II.
Ronald W. Hardy, Director
Aquaculture Research Institute
University of Idaho University
of Idaho
Current areas of emphasis in fish
nutrition research
• Replacing marine protein and oil
• Effects of nutrition on food quality and
fish health
• Developing micro-particulate feeds for
small larvae at first feeding
• How can these areas be investigated
using new genomic tools?
Replacing marine protein and oil
• Rapid growth of aquaculture over the past 15 years is
due to:
– Industry growth in developed species
–expanded production of new species
–Switch from extensive to intensive production of pond fish
through higher feed inputs in SE Asia and China
• Proportion of annual global production of fish meal and
fish oil used by aquaculture has tripled
• Production of fish meal and oil cannot increase except
in Alaska by increased recovery of processing waste
• Only way to meet expected demand for protein and oil in
aquafeeds is to find alternatives to fish meal and oil
World fish meal use in livestock and
fish feeds in 1994, 2000 & 2006
60 Poultry
50 Swine
40 Aquaculture
30 Other
20
10
0
1994 2000 2006
World fish feed production
1995, 2000, and 2010 (predicted)
30,000,000 1995
25,000,000 2000
20,000,000 2010
15,000,000
10,000,000 Catfish Marine Cyprinids
5,000,000
0
Salmonids Shrimp
Summary of demand
• Average annual production of fish meal will be
equal to demand in fish feed by:
– 2015 at current incorporation levels in fish feeds
– But, if global production of fish meal decreases due to El
Nino or wild stock collapses, demand may equal supply
as soon as 2010
• More likely scenario
– Global production is adequate until at least 2020
if percentage use in fish feeds decreases
– Fish meal will no longer be used as the main
protein source, but rather as a feed supplement
• Palatability of plant-based feeds
• Source of taurine, carnitine, and other compounds
• Essential amino acid balance in plant-based feeds
Can carnivorous fish be
converted to a vegetarian diet?
• Trout fed a fish meal-free diet grow about
7% slower and are 10% less efficient
• We cannot formulate a fish meal-free diet
without adding several limiting amino acids
or adding rendered animal products
• Basing veggie trout feeds on plant protein
concentrate increases fiber (phytate) in diet,
thereby contributing to pollution
Similarities to past situation in
poultry feeds
• In USA, poultry feeds are based on soybean meal
and corn
• In 1970s, however, poultry performance could not be
sustained without 3-5% fish meal in the formulation
• Research over decades showed that fish meal
contained various micronutrients, mainly ultra-trace
elements, that were essential to chickens
• When these were supplemented, fish meal could be
eliminated from poultry feeds
Poultry feed analogy is not perfect
• Ultra-trace mineral aspect may not
be relevant to aquatic animals
• More likely the growth factors in
fish meal are related to:
–Amino acid imbalances
–Palatability
–Bioactive compounds in fish meal and plant
proteins
Challenges in feed formulation-1
• Replacing fish meal with plant proteins
–Corn or wheat gluten, soybean meal, soy
protein concentrate, canola protein concentrate
• Emphasis is on maintaining digestible
protein and limiting amino acid levels
• Fish meal contains bioactive compounds
–Gonads, nucleotides, others
• Oilseed meals contain bioactive compounds
– phytoestrogens and other compounds
Challenges in feed formulation-2
• Replacing fish oil with plant oils
–Canola, soy, flaxseed and others
• Fatty acid profiles of fish reflect dietary fatty
acid intake
• Fish oil contains bioactive fatty acids
–Long-chain omega-3s
• Plant oils contain relatively high amounts of
linoleic acid (C18:2, n-6)
–Interferes with omega-3s
–Can lead to production of inflammatory factors
Nutritional pathologies caused by
changing feed formulations
• Fin erosion
• Skeletal deformities
– Phosphorus deficiency, but
complicated etiology with ascorbic
acid or other factor possibly
involved
• Enteritis in distal intestine
– soybean meal and salmonids
Effect of diet on fin erosion of
rainbow trout
Standard Experimental
fish-meal based feed krill meal based feed
Atlantic salmon with jaw deformity
(Screamer)
“Screamer” at harvest
Skeletal deformities in Atlantic salmon
Normal Atlantic
rainbow salmon
trout screamer
Phosphorus
deficient
rainbow
trout
Skeletal deformities
• Deformities in farmed Atlantic salmon
– Behind head (chicken head)
– Mid-dorsal area (humpy)
– Caudal area (stumpy)
• Fish look normal at seawater transfer, but develop spinal
deformities during grow-out and are downgraded at harvest
• Examination of fry and fingerlings prior to seawater transfer
shows abnormal vertebra
– Abnormalities not grossly apparent, but seen in X-ray imaging
• Cause is thought to be inadequate mineralization (low dietary P
with other factors involved) at juvenile stage, followed by injury
during seawater transfer and size-grading
Skeletal deformities in Atlantic salmon
Example of problems with plant
proteins: soybean meal and enteritis
• Morphological abnormalities of intestinal villi
• Appearance related to dietary soybean meal
level and duration of feeding
• Can be induced in Atlantic salmon and
rainbow trout, but not in Atlantic cod
• Causative factors unknown but not present in
soy protein concentrate
SBM-induced enteritis- Distal intestine histology
STOMACH PROXIMAL
SECTION
PYLORIC DISTAL
SECTION SECTION
Soybean meal-induced enteritis after 24 weeks
Control vs. 40% SBM diet
Control diet: No SBM. 40% SBM diet: Villi are
Photo showing normal villi swollen and those in the
of distal intestine (X 75) center fused. Note
numerous large apical
vacuoles(X 75)
Summary of results in our laboratory
• Trout growth performance
• higher on diets containing 20% soybean meal than 40%
soybean meal
• Expression of immune factors
• Tumor necrosis factor (TNF) expression elevated in fish fed
40% soybean meal
• No differences in IL-8 or CD-8 expression
• Soybean meal-induced enteritis
– No evidence in any treatment after 12 weeks growth trial
– No evidence in Control fish (0% SBM) after 24 weeks
– Very low incidence in 20% SBM after 24 weeks
– High incidence in 40% SBM after 24 weeks
Chapter 10: Nutritional Pathology
Note to Dr. Roberts:
Chapter will need to
be expanded to
address new
pathological
conditions caused
by feed imbalances
Marine fish larval feeds
• Four problems with microparticulate feeds
– Larval fish cannot swim to catch feed
– Larval fish often do not recognize feeds as food
– Very small feed particles are susceptible to nutrient
leaching
– Larval fish do not have fully developed digestive
systems, so special forms of protein are required
Early fish larvae
Copepod: live prey for larval fish
Marine fish larval feeds
• Larval fish cannot swim to catch feed
• Larval fish often do not recognize
feeds as food
• One solution: add little glass balls to
increase pellet buoyancy and reflect
certain wavelengths of light that the
fish can detect
Making microparticulate feeds float and visible to larvae
by imbedding very small glass balls in the feed
Halibut larvae with microparticulate
feed (containing glass balls) in gut
Marine fish larval feeds
• Very small feed particles are
susceptible to nutrient leaching
Regular feed Coated feed
Marine fish larval feeds
Larval fish do not have fully developed
digestive systems, so special forms of
protein are required
• Line of research: Look at gene expression
of digestive enzymes and transporter
proteins in gut of developing larvae
– Pep1: peptide transporter expressed early in marine
larvae
• Develop diets containing amino acids and
peptides rather than intact proteins
Omega-3 fatty acid levels in farmed
fish
Problem: How to maintain omega-3 fatty
acid levels in fillets when plant oils are
used in the feed??
• Line of research: Use diets that
contain fish oil at the end of the grow-
out period
• Genomic research: Look at expression
levels of fatty acid desaturase enzymes
Fillet eicosapentenoic acid (EPA) composition
mg EPA 100/g edible tissue 500
Canola to menhaden oil
400 Menhaden to canola oil
300
200 *
100
0
Initial 6 weeks
* Denotes significant differences within dietary groups over time (One factor
ANOVA, P<0.05).
Diet history: Previously fed oil source for 17 weeks then switched
Initial weight 807g. At the completion of the experiment fish had attained an
average weight of 1118 g fish-1 with an SGR of 0.905 and an FCR of 1.17.
Fillet n3/n6 fatty acid ratio
Ratio of n3/n6 fatty acids 4.0
* Canola to menhaden oil
Menhaden to canola oil
3.0
2.0
*
1.0
0.0 6 weeks
Initial
* Denotes significant differences within dietary groups over time (One factor
ANOVA, P<0.05).
D-6 fatty acid desaturase expression
d6FAD-MGB Expression 1.0E+09 Canola to menhaden oil *
8.0E+08 Menhaden to canola oil 6 weeks
6.0E+08
4.0E+08 *
2.0E+08
0.0E+00 Initial
Genomics in fish nutrition research
• Nutritional genomics (nutrigenomics)
involves measuring expression of genes that
respond to different dietary factors
– Digestion, nutrient transport, metabolism, nutrient
partitioning, protein synthesis, protein turnover, and
so on respond to nutritional inputs
– Studying expression of regulatory genes in various
pathways will provide insight into physiological
processes
• Nutrigenomics is relevant to growth, immune
function, reproduction, and just about
everything
Genomic tools
• Micro-arrays
– Zebrafish array (genome fully sequenced)
– GRASP chip for salmonids
• RT-PCR
• Proteomics
Problem with existing microarrays
Have to sort through up to 25,000 genes to connect the dots in
metabolic pathways of interest, plus not all genes of interest
exist on available arrays
Strategy to utilize genomics and
proteomics in fish nutrition
• Conduct feeding studies
• Measure physiological responses and correlate these
with changes in gene expression using existing micro-
arrays
• Identify genes of interest, e.g., key regulatory genes
• Create groupings (panels) of key regulatory genes in
specific pathways of interest
• Incorporate panels into mini-arrays that allow us to
design precise, targeted experiments to test specific
hypotheses
Mini-arrays solve the problem of
gene expression overload
• Mini-arrays measure gene expression in key
regulatory or rate-limiting enzymes that are up or
down regulated in specific metabolic pathways
• Additional panels being developed by others can be
added to expand to stress, effects of pollutants,
intestinal enzyme expression
• Mini-arrays will simplify nutritional studies in fish
compared to using global microarrays
Food restriction in zebrafish
(why we need mini-arrays)
Microarrays ALDOLASE FRUCTOSE-1,6-
BIPHOSPHATASE
(gluconeogenesis)
Genes Brain Liver PYRUVATE
5 KINASE
genes 1041
genes MALATE
DEHYDROGENASE
(glucogenesis)
In LiverSUCCINATE
DEHYDROGENASE
Persistent organic pollutants (POPs)
• PCBs, dioxin, etc. were present in fish oil and the residual oil in
fish meal
• Wild fish from Baltic and North Sea are known to have much
higher concentrations of POPs than products from the Pacific
• Industry now removes POPs by extracting residual oil from meal
with isohexane, then treating with activated carbon (same for oil)
• After treatment, meals and oils are below EU and EPA action
limits
• Farmed salmon have lower POP levels than wild salmon,
especially those from Washington State
• All salmon have lower POP levels than English muffins
Farmed fish and contaminants
Farming fish is really the only hope to produce
fish with reduced levels of POPs
– Dietary inputs can be managed
– High percentage of diet is plant protein in
contrast to diet of wild fish which cannot be
controlled