Nutritional requirement of larvae and broodstock of commercially important fish and shell fish (1)

NUTRITIONAL REQUIREMENT OF
BROODSTOCK AND LARVAL NUTRITION OF
COMMERCIALLY IMPORTANT FISH AND
SHELL FISH
Submitted by,
Akhila S,MFSc 2018
Fish Nutrition and feed Technology

• One of the most poorly understood and
researched areas of finfish nutrition
• due to the necessity of suitable indoor or
outdoor culture facilities for maintaining large
groups of adult fish and the consequent higher
cost of running and conducting extended
broodstock feeding trials.
• many of the deficiencies and problems
encountered during the early rearing phases of
newly hatched finfish larvae are directly related
to the feeding regime (including nutrient level
and duration) of the broodstock.

Effects of Nutrition on
Fecundity of broodstock fish
• *Fecundity determines the egg quality, which can be
affected by some nutritional deficiency in the
broodstock diet.
• Reduced fecundity could be either caused by a nutrient
imbalance on the brain–pituitary–gonad endocrine
system or by the restriction in the availability of a
biochemical component for egg formation
• one of the major nutritional factors that has been
found to significantly affect reproductive performance
in fish is the dietary essential fatty acid content
• In most fishes, the fatty acid composition of the female
gonad is greatly affected by the dietary fatty acid
content, which in turn significantly influences egg
quality in a short period of time

• Polyunsaturated fatty acids can also regulate eicosanoid
production, particularly prostaglandins, which are
involved in several reproductive processes including the
production of steroid hormones and gonadal
development such as ovulation
• Fish ovaries have a high capacity to generate eicosanoids,
among them prostaglandin E (PGE) derived from
cycloxygenase action and *leukotrienes LTB4 and
LTB5 derived from **lipoxygenase action.
• The excess of EFA also have a negative effect on
reproductive performance of fish.˜
• High dietary n−3 HUFA levels could affect the brain–
pituitary–gonad endocrine axis since both EPA and DHA
have been found to reduce in vitro the steroidogenic
action of gonadotropin in the ovary of teleost fish

• Other nutrients which have been shown to
affect fecundity include vitamin E and
ascorbic acid
• Dietary antioxidant requirements increase
during reproduction
• Supplementation of 0.1% *tryptophan in
the diets of ayu Plecoglossus altivelis.
resulted in a significant increase in the
serum testosterone levels thus advancing
time of spermiation in males and induced
maturation of females

Effect of broodstock
nutrition on fertilization
• Dietary eicosapentaenoic (EPA)and arachidonic acid (AA)
levels show a correlation with fertilization
• Since sperm fatty acid composition depends upon the
essential fatty acid content of broodstock diet in species such
as rainbow trout and European Seabass it is possible that
sperm motility and in turn fertilization would be affected.
• The timing of spermiation may be delayed and subsequently
fertilization rates reduced by depressed steroidogenesis
caused by a broodstock EFA deficiency or imbalance
• prostaglandins are also recognized as important pheromones
in some teleost fish
• The antioxidant function of vitamins C and E can provide an
important protective role for the sperm cells during
spermatogenesis and until fertilization by reducing the risk of
lipid peroxidation, which is detrimental for sperm motility.

Effect of broodstock nutrition
on embryo development
• AA and EPA or DHA play an important structural role as
components of phospholipids in fish biomembranes and
are associated with the membrane fluidity and correct
physiological functions for bound membrane enzymes and
cell functions in marine fish
• the n-3 PUFA are regarded as major energy sources during
early embryonic development
• Free radicals are able to deteriorate egg membranes and
membrane integrity. Vitamin E, vitamin C and carotenoids
e.g. astaxanthin, are strong scavengers of active oxygen
species and have been shown to have a protective role
against the action of free radicals.
• vitamin E functions as an inter- and intra-cellular
antioxidant to maintain homeostasis of labile metabolites
in the cell and tissue plasma

• The carotenoid content of broodstock diets has also been
reported to be important for normal development of fish
embryo and larvae
• Carotenoids constitute one of the most important
pigment classes in fish, with a wide variety of functions
including protection from adverse lighting conditions, a
provitamin A source, chemotaxis of spermatozoa and
antioxidant functions including singlet oxygen quenching.
• vitamin C is necessary for the synthesis of collagen during
embryo development.
• Vitamin A is considered important for embryo and larval
development due to its important role in bone
development, retina formation and differentiation of
immune cells.
• Other dietary nutrients which have been found to affect
the reproductive performance of marine fish include
dietary protein intake.*

Effects of broodstock
nutrition on larval quality
• Few studies have been able to show the
improvement of seed quality through
implementation of broodstock nutrition
• *Increased n-3 HUFA ,particularly
docosahexaenoic acid levels in
broodstock diets were reported to
significantly enhance the weight of fish
larvae and their resistance to osmotic
shock

Timing of broodstock
nutrition
• In some fishes, egg composition is readily affected by the
diet within a few weeks of feeding
• In those species which are continuous spawners with
short vitellogenetic periods, it is possible to improve
spawning quality by modification of the nutritional quality
of broodstock diets even during the spawning season
• it is possible to improve egg quality and hatching rates of
seabass by feeding broodstock with appropriate amounts
of HUFA during the vitellogenetic period
• In batch spawners with up to 6 months of vitellogenesis,
such as in salmonids, broodstock must be fed a good
quality diet for several months before the spawning
season to improve their reproductive performance
• In some fishes like salmons, the muscle and egg fatty acid
profiles reflect the dietary FA profiles only after 2 months
of feeding while some take only 15 days

Valuable ingredients for
broodstock diets
• Several feedstuffs have been recognized as highly valuable
for broodstock nutrition
• Cuttle fish meal and squid meal
• High dietary value of cuttle fish meal was due to the fat-
insoluble fraction of the meal*
• Certain components were identified in squid meal that
improves egg quality
• Broodstock were fed with diets fish meal, squid meal,
defatted fish meal with squid oil or defatted squid meal
with fish oil
• **Raw krill are also included in broodstock diets, with
doubled percentage of buoyant eggs, total hatch and
normal larvae.The polar and non-polar lipid fractions of
krill contain important nutritional components like
phosphatidyl choline and astaxanthin

Broodstock feeding practices
• For most cultured fish species, the commercially available so-
called broodstock diets are just larger sized on-growing diets
• In most fish hatcheries, brood fish are fed with fresh marine
by-products or in combination with commercial feeds
• Most common fresh marine organisms fed to brood fish are
squid, cuttle fish, mussels, krill and small crustaceans
• Main problem include inadequate nutrient levels and high risk
of disease transmission to parents and off springs including
endo and ecto parasites, and bacterial and viral pathogens,
etc.
• The nutritional quality of formulated feeds can be effectively
improved.* but would lead to high production costs, and be
higher if developed for individual species
• However, the benefits of improving the survival and thus
increasing the production of marine fish larvae will have far
reaching economic return than the initial cost of feeding the
broodstock feed.

SUMMARY
• In summary, information on the nutrient requirements of
broodstock fish is limited to a few species.
• Certain nutrients such as essential fatty acids and antioxidant
nutrients have been shown to be particularly important in
broodstock nutrition.
• Their requirements during reproduction is higher than those
of juveniles, but excess amounts of nutrients or an imbalance
can be detrimental for reproduction. Some minerals, such as
phosphorous, and other nutritional aspects, such as protein
quality, are also known to be important for fish reproduction.
• The importance of many other nutrients such as vitamin A,
vitamin B6 and folic acid has not yet been established within
broodstock feeds and deserve future research.
• Future in vitro studies may provide clues to function and
certain unexplained biochemical mechanisms of certain
micronutrients in reproduction of fish; however, these studies
should complement in vivo research rather than substitute it.

PENAEID SHRIMP BROODSTOCK
NUTRITION
• Healthy females (25-30 cm body length and 200-320 g weight) and males (20-25
cm; 100-170 g) captured from the wild are preferably used as broodstock in the
induced ovarian maturation process.
• Broodstock from greater depths (60-80 m), or more than 20 miles offshore, are
preferable due to the lower prevalence of shrimp diseases, which are higher in
coastal shrimp farming areas.
• Once the shrimp have recovered from transport stress for a few days, they are
stocked in a circular maturation tank that is normally covered and kept in a dark
room. The same stocking density (2-3/m²) is used for both females and males.
• Shrimp are subsequently induced to moult by manipulating the salinity of the
water. After mating has occurred, which is easily determined by the presence of a
spermatophore in the thelycum and hardening of the shell, the eyestalk of females
is unilaterally ablated for endocrine stimulation
• Broodstock are fed with squid, mussel or cockle meat, supplemented by
polychaete or Artemia biomass to enhance reproductive performance.

Amino acid requirement for
some species
Penaeus
japonicus
Penaeus
monodon
Penaeus
setiferus
Macrobrachium
rosenbergii
Lipid % ? 2.5-10.0 3-8 2.5-6.0
Protein % 48-60 35-39 20-32 20-25
Lysine 1/% 9.2 ? ? ?
Methionine 1/% 2.7 2/ ? ? ?
Essential
Components:
1-2% n-3 series fatty acids, particularly HUFA members of the series; high n-3/n-6
fatty acid ratio; marine protein for marine shrimp particularly.
Negative Factors: excessive cholesterol level; excessive vitamin C; Ca/P ratio >2.0
1/Based on the amino acid profile of mussel flesh (as % of
protein).
2/Cystine present.

Common Carp
Lipid up to 18% (higher levels
spare protein)
Protein 25-38%
Amino Acids lysine 5.7%)% of dietary
methionine (cystine absent) 3.1%) protein
Available Phosphorus 0.6-0.7%
DE 2 700-3 100 kcal/kg
Essential Components: at least 1% of each of n-3 and n-6 series fatty acids; high lipid diets
for oogenisis in broodfish.
Negative Factors: non-protein nitrogen: there is some evidence that carp may be able
to utilize this (this is disputed, however); rancid fat; high lipid or
carbohydrate after ovulation of broodfish.

Indian and Chinese Carps
fry & fingerlings juveniles & growers broodfish
Lipid (min %) 8 5 5
Protein (min %) 30 25 30
Calcium (min/max %) 0.8-1.5 0.5-1.8 0.8-1.5
Available Phosphorus
(min/max %)
0.6-1.0 0.5-1.0 0.6-1.0
Lysine (min., as % of
protein)
6.7 6.4 6.0
Methionine/Cystine
(min as % of protein)
4.0 3.6 3.3
DE (kcal/kg) 3 100 2 800 2 800

Tilapia
Lipid 10% (fry to 0.5g)
8% (0.5-35 g animals)
6% (35 g - market size)
Protein 50% (fry to 0.5 g size)
35% (0.5-35 g animals)
30% (35 g - market size)
Digestible Carbohydrate 25%
Fibre 8% (fry to 10g)
8-10% (10 g - market size)
Lysine
Methionine + 50% cystine
4.1
1.7
as % of dietary protein
DE 2 500-3 400 kcal/kg
Essential Components: not less than 1% each of n-3 and n-6 series fatty acids
Negative Factors: rancid fats

General specifications for warmwater carnivorous species were
given by ADCP (1983) as follows:
fry + fingerlings juveniles + growers broodfish
Protein (min %) 36 30 36
Calcium (min/max %) 1.0-1.5 1.0-1.5 1.0-1.5
Available Phosphorus
(min/max %)
0.5-0.8 0.5-0.8 0.5-0.8
Lysine (min % of
protein)
5.6 5.3 5.0
Methionine + Cystine
(min 7, of protein)
3.3 3.0 2.8

LIVE FEEDS
• Living capsule of larvae
• Artemia, rotifer, cladoceran, zooplankton etc..
• Small size, autolytic digestion
• High nutrient content
• A pre requisite is consistent supply throughout the year
• Usually procurement expenses and inconsistent supply
• Also deficient in crucial nutrients like DHA, often leading
to some nutritional defeciencies thereby needing
enrichment.
• Enrichment done with n-3 HUFAs(artemia,rotifers) and
lipid soluble vitamins like A,D, E, K and taurine also
required.

• The use of live feeds how ever restricts
the understanding of quantitative
nutrient requirements and interaction
of nutrient in larvae
• Also understanding larval nutrition is
complicated as the requirement varies
for the separate larval stages due the
the changing digestive tract anatomy
and associated digestive enzymes

DIGESTIVE ENZYMES
• Satisfaction of nutrient requirement is
definitely influenced by the type and
quantity of digestive enzyme produced by
different stages of larval development
• Although the array of digestive enzymes
available are different and quantitatively
restricted, particularly during early stages
of development, sufficient number and
concentration are eventually produced for
good growth and survival during different
stages of metamorphosis

Commonly used live feeds in
Aquahatcheries
YEAST
• Serve as primary sourse of feed for larvae as well as zooplankton
• Act as artificial larval diets
MICRO ALGAE
• Chlorophyll bearing multicellular/ unicellular plants
– Chlorophyta(Freshwater)
– Phaeophyta(Marine)
– Rhodophyta(Marine)
• Diatoms – chaetoceros, skeletonema
• Small plankters : isochrysis, tetraselmis, chlorella
• They mainly serve as food source for copepods and cladocerans,
rotifers and artemia
• Contain 30-40% protein, 10-20% lipids, 5-15% carbohydrates

INFUSORIA
• Microscopic, single-celled animalcules, mainly
ciliata
• Usually serve as started diets
• Freshwater : Paramecium, Stylonchia
• Marine : fabrea, Euplotes
ROTIFERS
• wheel animalcules
• Eg : Brachionus plicatilis, Brachionus
rotundiformes
• 52-59% protein, 13% fats, 3.1% n-3 HUFA
• They are fed with bakers yeast and artificial
diets to enhance nutritional QUALITY.

ARTEMIA
• obtained in hatcheries mainly in the form of cysts
• 90% of cysts obtained from salt lake of Utah
• 1 gram cyst produces about 2-3 lakh naupli
COPEPODS
• One of the main copepod species cultured in hatcheries
include Tigriopus japonicus
CLADOCERANS
• Mainly cultured species include Daphnia, Moina etc..
• Contain about 705 protein
• One of their main advantage is that they show high
reproductive capacity
Inspite of all the above considerations and species cultured,
one of the main problem include the difficulty to obtain
pure strains and also the possibility of disease occurance

MARINE FISH
• Marine fish can neither biosynthesize 22:6(n-3) de novo
nor from shorter chain precursors such as 18:3(n-3),
therefore 22:6(n-3) and 20:5(n-3) are essential dietary
constituents for marine fish
• In fish 22:6(n-3) is present in very high concentrations in
neural and visual cell membranes and synaptosomal
membranes, as in the case of mammals.
• An insufficiency of 22:6(n-3) in marine larval fish diet is
likely to impair neural and visual development with
significant if not serious consequences for a whole range
of physiological and behavioural processes including those
dependent on neuroendocrines.
• Abnormal pigmentation in cultured marine flatfishes is
related to HUFA deficiencies.

• From detailed studies, it was examined that
excess levels of AA(20:4(n-6)) was found to be
deleterious, because of a generalized
biochemically-induced stress in the fish through
excess eicosanoid production.
• In commercially available fish oils, 20:4(n-6) are
found to be consistently at low levels (< 1% of
the total fatty acids).
• Nearly all mariculture production systems rely
heavily on live feeds viz., rotifers, atremia
nauplii and copepods
• Artemia and Brachionus strains, being defecient
in EPA and also DHA makES (n-3) HUFA
enrichment necessary
• Enrichment done with emulsions of fish oils or
even some commercial products

• Current problems in enrichment of live feed include
1. content is very small in triacylglycerol micelles
generated in enrichment procedures and are
prone to autooxidation, especially under
vigorous aeration.
2. Natural antioxidants such as á-tocopheryl
acetate and ascorbyl palmitate are not
effective especially until hydrolysed in the
intestinal tract and absorbed
• Lecithin can be used to considerable advantage in
enriching the nauplii with 22:6(n-3) rich fish oils,
because lecithin acts as a natural emulsifying agent
and a natural protectant against autooxidation
• Lecithin derived from fish eggs is superior to soy
lecithin because fish egg lecithin contains readily
assimilable 22:6(n-3) and 20:5(n-3) in the ratio of
2:1.

• *Alternatives to fish oil fractions rich in 22:6(n-3) are
1. A heterotrophic dinoflagellate Crytothecodenium
cohnii which is mass produced commercially to
produce triacyl glycerol rich in 22:6(n-3) -
commercial product by MARTEK®;
2. Spray dried Schizochytrium spp. rich in PUFA is a
single celled heterotrophic marine protist of the
group labyrinthulomycota - commercial product
KELCO®.
3. Copepods cultures have to be developed because
they have a preponderance of phospholipids rather
than triacylglycerols in their body.
4. Copepods enriched with freeze thawed cells of C.
cohni or Schizotricodinium spp. is another
possibility ensuring the appropriate HUFA ratio
delivery to larval marine fishes

Commercial fish oils
• Commercial fish oils are the richest source of fats and
fatty acids
• Commercial fish oils are enriched with EPA and DHA by
fractional distillation, solvent extraction and also urea
adduction or by a combination of these methods
• (n-3) PUFA’s are available as ethyl ester, free fatty acids
and rarely as triacylglycerols among which ethyl esters are
already used to enrich artemia
• Commercial fish oils can meet enrichment requirements
because saturated and monounsaturated fatty acids in
fish oils are as important as energy yielding molecules and
(n-3) PUFA are useful for structural purposes
• over enrichment with PUFA could conceivably result in
insufficient energy content in the diet.*

• The only DHA rich natural fish oil known so far
is tuna orbital oil (TOO), which contains 30%
DHA, 7% EPA and 2% AA
• Blending of 90% TOO with 10% lecithin from
fish roe produces the most ideal enrichment
emulsion known to date
• maintenance of the levels of DHA: EPA: AA in
artemia till the larval fish feeds on it has not
been successful because all these fatty acids
especially DHA is metabolized by artemia after
bioencapsulation leading to lowering of its
content in the enriched organism.
• a strain of artemia from China designated as
Artemia sinica is found to retain the levels of
DHA up to 24 h post-enrichment.*

MICROPARTICULATE DIET
• Quantitative and qualitative nutritional requirement
of larvae differs according to age, feeding habit and
capacity of digestive system.
• Size of larval feed depends on feeding behaviour of
larvae
– Filter feeders(5-150µm)
– Particle feeders(40-700µm)
• Benefits
• Uniform nutrition
• Year round availability
• Convenience of suitable particle size
• Cost effective compared to live food culture

Types of micro-particulate diets
Micro- encapsulated
• Encapsulating the feed particles in a coat, preferably a
lipid walled microcapsule or cross linked proteins
Micro bound
• Common type
• Produced by binding all the ingredients together using a
binder
– Crumbled feeders(cake or pellet produced first and
then crumbled)
– On size feeders
Micro-coated
• Coating is usually done using a lipid for watersoluble
nutrients like vitamins, minerals and amino acids

Taurine – a crucial nutrient in
larval nutrition
• Aminoethylsulphonic acid
• Enhances growth and survival of
larvae
• Enhances the pigmentation of larvae
• Facilitates proper eye functioning

Selenium
• Trace mineral in anti-oxidant system
• Prevents damage due to oxidative
stress and also accelerates growth
and development of larvae

Protein hydrolysates
• Enzymes like Protease, Lipase and amylase remain scarce
during early stages of development.
• Results in insufficient digestion and absorption of protein,
lipid and other nutrients into the body
• Consists of low-molecular weight protein resulting from
protein pre digestion which are more likely to be
absorbed by the enterocytes
• Protein hydrolysates of raw materials like fish meal,
chicken viscera and yeast can be made following satndard
protocol
• Proteins are hydrolysed in-vitro using proteolytic enzyme
papain(endopeptidase) leading to formation of di and tri-
peptides, which can be used as supplements in larval
feeds

Phospholipids
• Major structural lipid componentin the
cell membranes, helping to maintain its
fluidity and permeability
• Aids transportation of substances across
cell(nutrients from intestine to other
cells of larvae)
• Improves stress-resistance,skeleton
development in flatfish metamorphosis
and pigmentation

Wax spray beds(WSBs)
Improve delivery of water soluble
substances to marine larvae
Liposomes
Important role in delivering water
soluble nutrients to marine
suspension feeders

REFERENCE
• Effect of broodstock nutrition on reproductive performance of fish
Author links open overlay panelM.SIzquierdoa
HFernández-
Palaciosa
A.G.Jtaconb(
Aquaculture Volume 197, Issues 1–4, 1 June
2001, Pages 25-42)
• CMFRI - Winter School Course Manual on “Recent Advances in
Breeding and Larviculture of Marine Finfish and Shellfish”. 30.12.2008
- 19.1.2009
• ADCP/REP/87/26 - Feed and Feeding of Fish and Shrimp
• Nutrient requirements of fish and shrimp,NRC
• Live feed culture and larval rearing of marine finfishes G. Gopakumar,
A. K. Abdul Nazar and R. Jayakumar Mandapam Regional Centre of
CMFRI Mandapam Camp - 623520, Tamil Nadu, India
• Micro –particulate diet, a boon for larval culture- sikandra Kumar,
CIFE,Versova

Nutritional requirement of larvae and broodstock of commercially important fish and shell fish (1)

Nutritional requirement of larvae and broodstock of commercially important fish and shell fish (1)

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