Fish and Aqua

FISH BREEDING (SEED PRODUCTION) & HATCHERY MANAGEMENT FOR COMMERCIAL PURPOSE IN INDIA

January 29, 2020

1778

FISH BREEDING (SEED PRODUCTION) & HATCHERY MANAGEMENT FOR COMMERCIAL PURPOSE IN INDIA

Post no-1469 Dt-29/01/2020
Compiled & shared by-DR RAJESH KUMAR SINGH ,JAMSHEDPUR, JHARKHAND,INDIA 9431309542,rajeshsinghvet@gmail.com

Bundh Breeding

Bundhs are specialized ponds where the condition of reverine flow are created by constructing embankments against the large catchment area, subjected to rapid flooding during monsoon. Bundh are special type of perennial and seasonal pond or impoundments where reverine conditions are simulated during monsoon months. The bundh after heavy rain, receive large quantities of rain water, washing from their extensive catchment and provide large shallow marginal areas which become the spawning ground for carps. Bundh-breeding, this is prevalent only in the States of Madhya Pradesh and West Bengal, accounted for 5.38% of the total fish seed production in the country. It is however, reported that wet bundhs are existent in certain parts of Andhra Pradesh, Uttar Pradesh and Bihar, wherein breeding of major carps reportedly takes place; but no definite information as to their occurrence and magnitude is available with the concerned State Governments.

Bundhs are Two Types:

1) Perennial Bundh – Wet Bundh
2) Seasonal Bundh – Dry Bundh

Bundhs

Bundhs are special type of tanks or impoundments where riverine conditions are simulated during monsoon months for breeding carps.
They may be perennial (wet bundhs) or seasonal (dry bundhs).
Bundh breeding, which accounted for 5.4% in 1964-65, contributed to about 63% in 1980.
Bundh breeding is popular in Madhya Pradesh and West Bengal.
Bundh breeding seems to have its origin in West Bengal.
Majority of bundh-type of tanks, where major carps are known to breed, are located in the districts of Midnapore and Bankura in West Bengal and around Nowgong in the Chhattarpur district of Madhya Pradesh.
The first dry bundh was located in Sonar Talliya in Nowgong district of Madhya Pradesh.
The initial success achieved by the Department of Fisheries, Government of Madhya Pradesh, during 1958 in dry bundh breeding of carps in Sonar Talliya led to the construction of many more dry bundhs with improved designs.
The most modern constructions are generally masonry structures with arrangements for a sluice gate in the deepest portion of bundh for complete draining and one or two waste weirs for overflow of excess water.
In most cases, apart from the bundh itself, a dry bundh unit consists of storage ponds for stocking breeders, an observation tower with arrangements for storing necessary equipment and a set of cemented hatcheries (2.4m x 1.2m x 0.3m) with a regular supply of water for a large number of eggs at a short time.
In some cases, the embankment is a pucca stone masonry with a small sluice gate and a portion of the embankment itself serves as the waste weir (Dubey, 1969).

Wet bundh

A typical wet bundh of Midnapore (West Bengal) is a perennial pond or tank, situated in the slope of a vast catchment area of undulating terrain, with proper embankments having an inlet towards the upland and an outlet towards the opposite lower end.

During summer, only the deeper portion of the bundh retains water where carp breeders are released for spawning.
The remaining portion is dry and is many times used for agricultural purposes.
After a heavy shower, a major portion of the bundh gets submerged with water from catchment area (catchment area: bundh, 20-100: 1), coming into it in the form of streamlets.
The depth of water where breeding takes place has been stated to vary from 8 cm to 1.2 m.
Though the fish breeds at any spot in the bundh, it may be advantageous to prepare `spawning grounds’ at different levels which could get flooded at different water levels in bundhs.
The bundhs can also be filled with water as and when desired and breeding operations undertaken without waiting for the fresh rainwater.

Method of breeding carps in a wet bundh

With the onset of monsoon the rain water from the catchment area gushes into the bundh, creating an artificial current.
The breeders, already present in the deeper area of the bundh, migrate to the shallower areas and start breeding.
After the breeding is over, egg/spawn collection is done.

Dry bundh

A typical dry bundh is a shallow, seasonal depression, having a bundh on one side and a catchment area on the other three sides.
The bundh may be of varying shape and size and made of earthen wall or masonry wall.
A dry bundh is smaller and shallower than a wet bundh which is bigger and deeper.
The bundh gets flooded in monsoon, but remains completely dry for a considerable period during a year.
It consists of a sluice gate for quickly draining the water and an outlet for the excess water to flow away. In dry bundhs, spawning sometimes takes place in deeper areas.

Technique of breeding major carps in a dry bundh

The mature carp breeders which are raised in perennial ponds elsewhere are introduced into the bundh at 1:2 (Female: Male).
The fish are left undisturbed for 2-3 days so that they get acclimatized to new environment.
After this, 10-20% of the fish is given intramuscular injection of pituitary extract or ready–to-use spawning agents which are synthetic hormones.
Water current is created in the bundh by drawing water from a store tank.
The following morning, the spent breeders are removed, eggs collected, water drained and the bundh dried for 2-3 days.
The bundh is then utilized for the next breeding by releasing a fresh batch of breeders.
Five to six spawning are generally conducted in each bundh during one season as opposed to only one spawning in a wet bundh.
Silver carp and grass carp have been successfully induced bred in bundhs without stripping.
Sinha et al. (1979) have reported natural spawning of both grass carp and silver carp in a dry bundh of Bankura District where they were able to spawn the two species without stripping.
They consider dry bundhs to be one of the reliable means of mass breeding of Chinese carps to meet the increasing demand of their seed.

Collection and hatching of eggs

After spawning is over, the eggs are collected from bundhs, after lowering the water level, by dragging a piece of mosquito netting cloth (gamcha) and released for hatching either in improvised pits or double-walled hatching hapas or cement hatcheries.
The hatching pits (448 cm x244cm x 46 cm) are excavated on the bank with arrangement for the supply of water.
Each pit may contain about 0.9 to 2.2 million eggs, of which 2.5-25% hatch successfully.
A double-walled hapa, which is fixed in the bundh itself, consists of an outer hapa (182 cm x91 cm x 91 cm) and an inner hapa (152 cm x 76 cm x 46 cm ), accounts for a spawn survival rate of 32 to 50%.
The provision of cement hatcheries (2.4 m x 1.2 m x 0.3 m) near the dry bundhs in Madhya Pradesh has aided in improving the survival of hatchlings to 97%.
A cement hatchery of Madhya Pradesh has three times more capacity than a double-walled hapa and is far more economical than the latter. Since the collection of all the eggs is impossible, especially in the case of wet bundhs in view of their larger size, fry and fingerling collection is equally important.

Factors responsible for breeding of fish in bundhs

Spawning may occur at night and during bright sun in the forenoon.
After a period of breeding behaviour, mating occurs with vigorous splashing of water and a number of scales may get dislodged while some fish may even sustain minor injuries.
After spawning is over, a thick blanket of eggs is left behind at the spawning site.
The spent fish in bundhs move to the deeper areas.
No single factors can probably be attributed to spawning of major carps in bundhs and rivers.
The act of spawning involves the completion of a chain of interrelated pre-conditions.
1. Heavy monsoon flood capable of inundating vast shallow areas is believed to be a primary factor responsible for spawning. Some workers believe the availability of shallow spawning ground to be a deciding factor for spawning. The rise in the level of water, naturally or artificially, is known to bring about spawning.
2. The temperature of water for spawning is found to be between 22 and 33°C.
3. Other factors like pH, high Dissolved Oxygen, alkalinity, chloride and minerals do not seem to play any significant role in spawning.
  Soil type is not very important.
4. Spawning is inhibited due to the presence of hormone-like secretion in captive waters.
5. Water that has flown through a dry bed of land rich in humus has stimulatory effect on spawning.

Induced breeding of warmwater finfishes and environmental factors affecting spawning

Houssay (1930) of Argentina was the first to attempt induced breedingof fish by using pituitary extract on a viviparous fish. He was successful in obtaining premature birth of young fish.
Subsequently, based on the lines of Houssay, Von Ihering and his team of Brazil, in 1934, successfully induced bred a catfish with pituitary hormones and hence credit for the present day concept of induced breedingof fish goes to Brazilians.
In India, Chaudhuri and Alikunhi (1957) successfully induced major carps to spawn through hypophysation technique.
Since then, the technique has been standardized and refined for the large-scale production of fish seed.
The Indian Major Carp, which normally spawn once a year either naturally or through hypophysation during monsoon, were successfully induced bred twice within an internal of about two months.
Chondar (1984;1990) described a method for the mass scale breeding of IMC and silver carp in `Bangla bundh’ through Human Chorionic Gonadotropin (HCG) and its combination with pituitary extract.

Environmental factors concerned with breeding of fishes

Environmental factors concerned with fish breeding are

Light
Temperature
Ecological factors
Meteorological conditions

These factors are known to play important roles in stimulating the release of pituitary gonadotropins, thereby controlling reproduction in fish.

Light

It is an important factor that controls reproduction in fish.
Early maturation and spawning of fish as a result of enhanced photoperiodic regimes.
In India, Cirrhinus reba was found to attain early maturity when subjected to artificial day lengths longer than natural day even at a low temperature of the winter months, viz. 19-20^o
The resorption of gonads in C. reba was delayed and spawning conditions could be maintained up to November.

Temperature

The role of environmental temperature on sexual maturation and spawning of fish in India has been studied.
All observations show that there are optimum temperature ranges for induced breedingof cultivable fishes and critical temperature limits, above and below which fish will not reproduce.
The Indian Major Carps are found to breed within a range of 24-31^o
Beyond this range fish do not spawn.
The Chinese silver and grass carp have been successfully induced bred at temperatures 28.2^oC to 34^o
It was observed natural spawning of pituitary injected grass carp at a water temperature varying between 28.9 and 31.1^oC, the optimum being 27^oC, as in the case of Indian Major Carps.

Other Factors

It was opined that fresh rainwater and flooded condition in a tank are the primary factors in triggering the spawning of carps.
The presence of repressive factors may be responsible for inhibiting spawning of carps in confined waters, but when this repressive factor is sufficiently diluted by the onrush of floods in bundhs or ponds, spawning occurs.
Some workers suggested that it is the sudden drop in the electrolytes level in the environment caused by heavy monsoon rain or water current which induces gonadal hydration, resulting in natural spawning of carps.
Rain water and weather condition are important factors for induced breedingof fish.
Successful spawning in the majority of fishes has been induced on cloudy and rainy days, especially after heavy showers.
The carps are known to breed at a fairly wide range of pH and dissolved oxygen content.

Sympathetic breeding

Sympathetic breeding refers to the breeding of uninjected fish at the sympathy of injected fish.
This is common in bundh breeding, wherein, only 10-20 brooders are injected with either pituitary extract or synthetic spawning agent and the rest are not injected.
After an interval of about 8-10 hours, the injected brooders first start spawning and subsequently the uninjected brooders are also stimulated to spawn, thereby leading to the complete spawning of all the brooders.
Sympathetic spawning leads to lesser use of hormone and reduced handling of brooders.
By this method, natural spawning of both grass carp and silver carp is possible in a dry bundh of Bankura District where they spawned naturally, without stripping.
Some consider sympathetic breeding as one of the reliable means of mass breeding of Chinese carps to meet the increasing demand of their seed.

Fish Pituitary gland

Pituitary gland is an endocrine (ductless) gland situated on the ventral side of the brain.
It is a small, soft, whitish body whose size and shape vary with species.
It is more or less round in carps; oval in catla and rohu and pear-shaped in mrigal.
The pituitary is located in a concave cavity known as Sella turcica and enclosed by a thin membrane known as duramater.
It may be attached to the brain by a short stalk called the Infundibular stalk.

Types of pituitary glands

Based on the presence or absence of the stalk, the pituitary is classified into

Leptobasic pituitary (with stalk)– eg. Carps and catfishes
Platybasic pituitary (without stalk)– eg. Murrels and glassfish (Ambasis species)

The teleost pituitary comprises of two parts-

The glandular part (the adenohypophysis)
The nervous part (the neurohypophysis)

Collection of pituitary gland

Fish pituitary gland can be collected by dissecting and removing a portion of the scalp or through the Foramen magnum.

Dissecting and removing a portion of the scalp

In this method, the brain case (cranium) is obliquely cut using a butcher’s knife/hand saw/bone cutter and the scalp removed.
The brain is then exposed by removing grey matter and fatty substance with forceps and cotton.
The anterior end (optic and olfactory nerves) of the brain is cut and the entire brain is lifted up and laid back, thus exposing the pituitary under a membrane.
After removing the membrane and the fluid, the pituitary is lifted up by inserting the blunt end of the forceps and carefully transferred to a vial containing a preservative.

The pituitary mounted on to a wrist

(2) Through the Foramen magnum

Foramen magnum is a large posterior aperture of the skull through which the spinal cord passes.
The grey matter and fatty substance are first removed with the help of forceps and cotton (they are pulled out posteriorly).
The brain is then exposed.
After this, the anterior end (optic and olfactory nerves) of the brain is cut and the entire brain is lifted up and laid back, thus exposing the pituitary.
After removing the fluid the membrane, the pituitary is lifted up by inserting the blunt end of the forceps and carefully transferred to a vial containing a preservative.

The first method is commonly practiced even though the second method is less time consuming and a large number of glands can be collected within a short time, with a good resale value of the fish.

Preservation of pituitary gland

Preservation in absolute alcohol

In this method, the gland, after collection, is immediately transferred to a vial/phial containing fresh absolute alcohol (ethanol).
After 24 hours, the alcohol is removed and fresh alcohol is added and stored at room temperature or in a refrigerator.

Preservation in acetone

Immediately after collection, the pituitary gland is kept in ice-chilled acetone and stored in a refrigerator for 2-3 days.
After this period, the acetone is changed and the gland stored in a refrigerator.
Both absolute alcohol and acetone have de-fattening and dehydrating effect.

Immediate freezing

In this method, the collected glands are frozen immediately and stored in a freezer.

Preparation of fish pituitary extract for injection

The extract preparation should be carried out just before injection.
The required quantity of glands is taken out of vial and they are dried on a filter paper by allowing the alcohol to evaporate.
The glands are then homogenized with distilled water or saline in a tissue homogenizer.
If acetone-dried glands are used, they can directly be taken for maceration.
One-third of the media is used for homogenization, while the remaining two-third is used for rinsing the homogenizer and the glass rod.
Recommended dilution rate is 20-30 mg in 1 ml of the media.
The extract is centrifuged at 5,000 rpm for 5 minutes.
The clear supernatant solution containing gonadotropins is taken in syringe for injection.

Types of injection

Homoplastic injection: Injecting pituitary from one fish to another fish closely related to the donor fish. E.g. carp pituitary gland extract to carps.

Heteroplastic injection: Injecting pituitary from one fish to another fish distantly related to the donor fish. E.g. carp pituitary gland extract to catfish and vice versa.

Methods of injecting fish brooders

There are three methods of injecting brooders.
They are :

Intra-muscular injection:

It is administered into the muscle on the caudal peduncle or behind the dorsal fin, but above the lateral line.
It is most effective, convenient, simple and less risky.
It is widely practiced.

Intra-peritoneal injection:

It is give through the soft regions of the body, generally at the base of the pelvic fin or the pectoral fin.
It is risky as it may damage the gonads or liver.

Intra-cranial injection:

In this method, the injection is given through the cranium and is also risky as it may damage the brain.
The pituitary extract is administered through a glass or disposable syringe, 2.0 ml capacity, having 0.1 ml graduation.
The size of the needle depends upon the weight of the brooder to be injected.
Needle number 22 is used for fish weighing 1-3 kg, No. 19 for larger fish and No. 24 for smaller fish.
When two injections are given, one is given on the side that did not receive the first injection.

Dosage of pituitary extract

Assessment of proper dosage is most important for successful spawning. In practice, the female receives two injections, while the male receives only one injection, i.e. at the time of second injection to the female.

I Dose or Provocative or preliminary dosage and II Dose or effective or resolving dosage.

The interval between the two doses is 6 hours.

Carp glands to major carps

	Female	Male
I Dose	2-3 mg/kg b.w.	nil
II Dose	5-8 mg/kg b.w.	2-3 mg/kg b.w.

Carp glands to exotic carps

	Female	Male
I Dose	4-6 mg/kg b.w.	nil
II Dose	10-16 mg/kg b.w.	4-6 mg/kg b.w.

Catfish glands to major carps

	Female	Male
I Dose	10 mg/kg b.w.	nil
II Dose	20 mg/kg b.w.	10 mg/kg b.w.

Catfish glands to exotic carps

	Female	Male
I Dose	20 mg/kg b.w.	nil
II Dose	40 mg/kg b.w.	20 mg/kg b.w.

Synthetic hormones for induced breeding of fishes

Studies conducted by numerous investigators on induced breedingof fishes have indicated the superiority of several ovulating agents over fish pituitary extract.
Although fish pituitary extract was initially used extensively for fish breeding all over the world, synthetic spawning hormones are now being increasingly used due to their efficacy and convenience.
Banerjee et al. (1989) succeeded in the purification of pituitary gonadotropic hormone from Channa punctatus andCatla catla.
Mammalian pituitary hormones in combination with fish pituitary gland extract precipitated spawning in fish.
Of all the mammalian hormones tested on fish, chorionic gonadotropin (CG) has given successful result in inducing fish to breed, probably because CG behaves primarily as a luteinising hormones (LH).
Synahorin (a mixture of CG and mammalian pituitary extract) in combination with pituitary gave positive results when injected to rohu.
Sinha (1969) reported the fractionisation of pituitary extract from carps and tilapia. He obtained success in spawning of carps.
Bhowmick et al. (1979) found mammalian hormones antuitrin-s, leutocyclin and RH-LH ineffective when injected singly or in combination with carp pituitary extract.
The CIFRI, Barrackpore undertook detailed studies on the use of LH-RH alone or in combination with progesterone and obtained breeding success which ranged between 25-49% in carps and 100% in catfish.

Synthetic spawning agents

The stimulation of pituitary gonadotropin secretion by synthetic LH-RH has been demonstrated in a number of teleosts.
Since LH-RH (natural or synthetic) alone is not very effective in inducing spawning in fish, a combination of LH-RH-a (GnRH-a) and a dopamine antagonist for induced ovulation and spawning in cultured fish is a highly effective procedure called the Linpe method.
Some workers reported successful spawning of catla, rohu and mrigal with LH-RH analogue at 10-20 mg/kg b.w. and also obtained 100% ovulation with pimozide at 10mg/kg b.w.
Parameswaran et al. (1988) achieved successful spawning in mrigal with LH-RH-a, buserelin acetate in combination with progesterone.
Investigations of Jose et al. (1989) with LH-RH-a indicated successful breeding of mrigal and Labeo fimbriatus.

The Linpe method and ovaprim

Both of these rapidly gained acceptance in fish farms in China and India and has now been commercialized by Syndel Laboratories, Inc., Vancouver, British Columbia, Canada, under the tradename ovaprim.
The ovaprim spawning kit is especially formulated for use with salmonids, cyprinids and other freshwater cultured fish.
It has been used successfully in a number of species in several countries and is gaining wide acceptance as the preferred method for induced ovulation and spawning of cultured freshwater fish.
For example, in India, based on field trials (during 1988-90) with ovaprim for induced spawning of Indian major carps, fringe lipped carp, silver carp, bighead carp and grass carp in various fish farms located in different agro-climatic regions, Nandeesha et al. (1990, 1991) concluded that in economic terms, the use of ovaprim is advantageous.
The spawning success, quantity of eggs obtained, the fertilization rate and hatching percentage remained consistently higher with ovaprim as compared to carp pituitary extract (CPE) or human chorionic gonadotropin (HCG) in almost all instances.
The results also indicate that nearly 40% more fry can be obtained by using ovaprim in place of commercial CPE.
Most of the carps tested generally spawned within 10-14 hours after injection. Ovulation and spawning has been successfully induced in India by the Linpe method in the Asian catfish, Clarias batrachus(Manickam and Joy, !989) and Indian catfish, Heteropneustes fossilis (Manickam, 1992).
Similarly, indigenous preparations, viz. Ovatide (M/s. Hemmopharma Ltd., Mumbai) and WOVA-FH (M/s. WOCKHARDT Ltd., Mumbai) are also being used commonly for the commercial spawning of carps and other fishes in India.
A combination of busereline (LHRH-a) and domperidone has been successfully used for the spawning of IMC (Basavaraja et al., 2007).

Dosage of ready-to-inject spawning agents (ovaprim, ovatide, WOVA-FH, etc.)

Females

Catla : 0.4-0.5 ml/kg b.w.
Silver carp : 0.4-0.7 ml/kg b.w.
Rohu : 0.3-0.4 ml/kg b.w.
Grass carp : 0.4-0.8 ml/kg b.w.
Mrigal : 0.25-0.3 ml/kg b.w.
Bighead carp : 0.4-0.5 ml/kg b.w.
Fringe-lipped carp : 0.3-0.4 ml/kg b.w.
Mahseers : 0.6-0.7 ml/kg b.w.
Catfishes : 0.6-0.8 ml/kg b.w.

Males (all species of carps) : 0.1-0.3 ml/kg b.w.

Males (catfishes) : 0.15-0.4 ml/kg b.w.

Steroids

An alternative approach is to use selected steroid hormones targeted at the oocytes. Most of the previous work on this subject is on induced ovulation in vitro.
The effects of steroid hormones on ovulation are seen primarily as germinal vesicle breakdown (GVBD).
GVBD is normally controlled by one or more steroids produced in the ovaries under gonadotropin stimulation, but the timing of ovulation related to that of GVBD varies.
In vitroovulation normally follows steroid induced GVBD in catfish ( fossilis). The action of pituitary gonadotropins on final oocyte maturation is known to be medicated through steroid hormones.
Deoxy corticosterone acetate (DOCA) and cortisone effectively stimulated in vitro ovulation in fossils.
The available reports indicate that steroid hormones are quite potent in inducing spawning in cultivated fishes, but are yet to find commercial applications.
The thyroid stimulating hormone is also reported to bring about ovulation in Indian catfish.
Although there are no reports on the effects of pheromones on the reproduction of IMC, there are circumstantial evidences which suggest that pheromones secreted by IMC help in effecting spawning.
Similarly, sympathetic spawning of carps in bundhs appears to be due to the release of pheromones.

Fish broodstock management and transportation of broodfish
Management of broodfish ponds

Brood fish is a prerequisite for all induced breedingprogrammes, as it produces eggs and milt, which are required for the production of larvae.
Proper brood-stock will lead to better breeding responses, increased fecundity, fertilization, hatching and larval survival rates and more viable fish seed.
Hence, the subject of brood fish management has assumed great importance in hatchery management.
The number of brood fish ponds depends on hatchery requirements.
Large-scale of operation and sex-wise segregation of fish requires more ponds.

Carp broodfish pond

Carp brood-stock ponds are generally large (0.2-2.5 ha), 1.5-2.5m deep, 30-40m wide, rectangular, seasonal or drainable and earthen in nature.
Water inlet and outlet should be such that they simulate riverine/fluviatile conditions, which is the natural habitat of IMC and Chinese carps.

Source of broodfish
Since selective breeding and hybridization programmes of pedigreed fish are not carried out in fish seed farms, the source of future broodfish is stock ponds from the same farm or different farms or live adult of different species procured from capture fishery waters like rivers, lakes or reservoirs.

Care of broodfish

The carp brood-stock pond should be prepared following standard procedure to ensure sustained production of zooplankton.
The recommended stocking density of carp brood fish is 1,250-2,500 kg/ha, depending upon the species.
While rohu and mrigal are stocked at a higher rate, catla is stocked at a lower rate since it requires more space for proper gonadal development.
Stocking rates are manipulated to permit individual and collective care of broodfish, enabling them to get nutritional and environmental advantages for onset of right degree of maturity.
During immature stage, feed the fish with a traditional diet consisting of rice bran and oil cake (1:1) at a feeding rate of1- 2% of body weight daily.
During the maturing phase, feed the fish with a special feed containing rice bran, oil cake, fish meal, cereals, grams and mineral and vitamin mixture.
Alternatively, one can use commercial floating pelleted feed (protein content : 30%)
In addition to the artificial feed the grass carp is also given tender aquatic weeds/terrestrial grass.
However, the breeding habits of some species like common carp demand their separation from other carp species due to their natural breeding in ponds with aquatic vegetation.
As a result the common carp brood fish is segregated sex-wise and stocked in separate ponds to prevent accidental spawning in pond.
However, the rest of the species can be stocked in a communal pond or stocked in separate ponds after species-wise and/or sex-wise segregation.
Catla, in particular, needs to be separated from the rest of the species as it shows poor response to hormonal injection when stocked with other species.
A gravid fish when held by hand with tail up should practically ooze milt and also ova.
Paddle-wheel aerator, particularly in catla pond, can provide additional aeration, particularly during morning hours.
Segregation of sexes at least one month before increases the affinity between male and female during spawning.
Care should be taken to maintain water qualityand plankton level by periodic manuring, i.e. at one tenth of the initial dose.
Algal blooms and oxygen depletion are controlled by water exchange.
Parasites and pathogens should be controlled by periodic checking of brooders
Common parasites like Lernea and Argulus are common on major carps (catla is more susceptible) can be controlled by manually removing and disinfecting the affected fish with a solution of KMnO4 (about 5 ppm)

Broodstock management practice

Proper brood fish management forms the key to successful spawning. The number and quality of eggs produced are significantly affected by the conditions under which the brood-stock is maintained.

The quality of brood-stock diet, feeding regime, the quality of brood-stock and water management are the principal factors that influence the condition of the broodstock.
Most seed farms raise broodstock in their own farm (there are instances of inbreeding depression, as reported by Eknath and Doyle (1985) and maintain them in ponds at a density of 1,000-2,500 kg/ha.
The earthen brood-stock ponds vary in area from 0.2 to 1.0 ha, with depth ranging from 1 to 2 m.
The farms use water from perennial reservoirs.
The number of brood fish ponds varies with hatchery requirement.
The main basic steps in the preparation of broodstock ponds are : control of aquatic weeds, which in done manually; eradication of unwanted fish by applying mahua oilcake at 2,000-2,500 kg/ha and pond liming at 100-200 kg/ha depending on the pH of soil and water.
This is followed by fertilizing the pond with cattle dung, at 15,000-20,000 kg/ha/yr or poultry manure at 5,000-10,000 kg/ha/yr to enhance heterotrophic food production.
In addition, 200-400 kg/ha/yr NPK mixture is applied in split doses at fortnightly or monthly intervals.
The initial dose of organic manure is reduced by half if mahua oil cake is used as piscicide.
After stocking the pond with carps that are one-year-old or more, they are fed with a conventional feed containing a mixture of groundnut oil cake and rice bran (1:1 or 1:2 ratio) at 1-2% b. w., once daily.
To ensure better and timely development of gonads, fish breeders use a special broodstock diet (protein : 25-30%) prepared using locally available cheap ingredients.
This diet is nutritionally superior, advances maturation and spawning by one or two months and results in increased fecundity and better seed quality.

Ingredients	%
Rice bran	25
Groundnut oil cake	25
Fish meal	10
Maize	10
Broken rice	10
Horse gram	10
Blackgram	10

This diet is given at 2% b.w. daily, starting in December.
At some farms about a third of the broodstock will be injected with a low dose of HCG at 6-7 mg/kg body weight every 20 days, starting from mid-February for advancing maturation so as to induce spawning by the end of May.
At some seed farms, a few vitamin E tablets are mixed, in addition to a small quantity of commercially available vitamin and mineral mix, in the diet to facilitate gonad development.
Algal blooms and oxygen depletion are the most common problems encountered in broodstock ponds. These can be overcome by frequent water exchange.
Although no major diseases occur in these ponds, infestation by Lernaea and Argulus on catla are common and are controlled by manual removal of adult specimens, followed by a dip treatment in a mild solution of potassium permanganate.

Transportation of broodfish

It is not economically feasible to transport bigger fingerlings/yearlings and broodfish in small packing containers.
For this purpose, truck mounted open tanks with facilitates for mechanical aeration and/or circulation were initially used quite successfully.
Open canvas containers (1m x 1m x 1.25m) are used in Punjab and Madhya Pradesh for transporting major carp breeders. In those States galvanized iron drums of 180 l capacity are also used.
In India, two successful models of closed system of live-fish carrier were designed.
One is due to Mammen (1962), which he called `Splashless tank’.
The later model of the splashless tank is of a petrol tanks design of 1,150 l capacity with an autoclave-type lid.
It has a built-in aeration system for supplying compressed air, which works on a belt driven by the engine of the transporting vehicle.
An oxygen cylinder is carried only as a stand by for emergency.
The inner surface of the tank is lined with U-foam which prevents physical injury to live fish during transport.
A total weight of about 250 kg live fish can be transported at a time in the splashless tank, as also 90,000 carp fingerlings.
The load ratio of fish to water in this type of carrier in about 1 kg of fish per 4.5 l water.
Patro (1968) developed a tank which has an outer chamber of 120 cm diameter open from top and a slightly smaller one closed from top; the latter, during transport, fits inside the former.
The top of the inner chamber is provided with an air vent and an oxygen valve.
The outer chamber serves as a storage tank and is initially filled with water along with fish to be transported.
The inner chamber, which is shipped from the upper open end of the water serves as an oxygen holding chamber at its top and is lined throughout with U-foam to prevent fish from sustaining injury during transport.
This double-barrel type carrier as stated by Patro can transport a total weight of 100 kg of live fish at a time.

Breeding of major carps

Induced breeding of Indian major carps

Breeding of fish with pituitary gland (hypophysis) extract is termed as Hypophysation
The credit for developing the technique of hypophysation in the world goes to the Brazilians, while the pioneers of hypophysation of Indian major carps are H.L.Chaudhary and K.H.Alikunhi.
Induced breedingrefers to inducing fish to release gametes through the application of pituitary extract or hormones or chemicals.

Identification of sex of brooders

Identificationof sex is a prerequisite to induced spawning of the fish.
Fish is sexually dimorphic and sexual dimorphism is exhibited primarily by gonads and their ducts and this involves killing of fish.
Alternatively, the sex is identified based on certain morphological/external characteristics which include size, length, weight, colouration, fin characteristics, modification in the head in the form of nuptial dress, genital opening, width of mouth, etc.
Carps are sexually dimorphic i.e. mature male and female are morphologically different.
Some of the external morphological characters which are developed during breeding season could be used to identify sex in major carps which mature during their 2^ndor 3^rd

Characteristics	Male	Female
1. Scale, Operculum and pectoral fins	Rough to touch, particularly the dorsal surface of pectoral	Pectoral smooth to slippery
2. Abdomen	Round and firm	Swollen and soft
3. Genital opening swollen	Elongated slit, white in colour, not swollen	Round and pink
4. When pressure applied on abdomen opening	milky white fluid oozes through genital opening	a few ova may ooze through genital
5. Shape of body and size	Body linear, swollen	stouter, slightly larger

Breeding technique

Induced breedingof carps starts with the onset of south-west monsoon, June.
The male and female brooders are conditioned for a few hours prior to injection.
Sets of brooders are formed, each consisting of 1 : 2 (female : male) ratio.
The injected brooders are released in the breeding hapa.

Breeding hapa

A Breeding hapa is a box-shaped cloth enclosure made of long cloth, generally of size 2 x 1 x 1 m with provision to close its top after releasing brooders.
The upper flap is attached to one side and the other sides are either tied or buttoned.
The hapa is fixed in a canal or pond or cement cistern.
The four bottom and four top corners are tied to four poles such that the bottom of the hapa should not touch the ground and one-third of the hapa remain above the water level.

Injection of brooders

Intra-muscular injection

It is administered into the muscle on the caudal peduncle or behind the dorsal fin, but above the lateral line.
It is most effective, convenient, simple and less risky.
It is widely practised.

Intra-peritoneal injection

It is give through the soft regions of the body, generally at the base of the pelvic fin or the pectoral fin.
It is risky as it may damage the goads or liver.

Intra-cranial injection

In this method, the injection is given through the cranium and is also risky as it may damage the brain.
The pituitary extract is administered through a glass or disposable syringe, 2.0 ml capacity, having 0.1 ml graduation.
The size of the needle depends upon the weight of the brooder to be injected.
Needle number 22 is used for fish weighing 1-3 kg, No. 19 for larger fish and No. 24 for smaller fish.
Intra-muscular injection is commonly practiced.
The hormone injection (pituitary/ovaprim/ovatide) is given at the caudal peduncle region in between posterior end of dorsal fin and base of caudal fin, above the lateral line, avoiding the lateral line.

Spawning

After releasing the brooders in the hapa, they should not be disturbed.
After about 6 hours, splashing will commence for breeding and be involved in courtship which will continue for one hour.
At the climax of the courtship, both the partners will be seen in an embrace with their bodies twisted around each other. This exerts pressure on the abdomen, resulting the extrusion of gametes.
The following morning, the spent brooders are removed and then the eggs are collected and transferred for hatching in a suitable hatching device.

Examination of eggs

After the eggs are water-hardened, a sample of eggs is taken in a beaker for assessing quality and quantity.

The fertilized (good) eggs are transparent with a clearly visible nucleus at the centre and look-like pearls.
The unfertilized (bad) eggs are opaque white and the nucleus disintegrate within one hour.
Fertilization rate
It indicates the quality of developing eggs and is estimated using the formula :
Fertilization rate (%) = No. of fertilized eggs/Total no. eggs x 100
Hatching rate
It can be estimated by knowing the total volume of spawn /number of spawn in a known volume
Hatching rate (%) : Total no. of spawn obtained/Total no. of fertilized eggs x 100

Induced breeding of exotic carps (silver carp and grass carp)

The silver carp and grass carp are native to China and hence are known as exotic carps.
They were first introduced into India in the year 1959.
Both were successfully induced bred in 1963, through hypophysation.
The method of hypophysation of these species is similar to that of IMC.
However, there are a few minor differences – the dosage of pituitary is double compared to IMC and, unlike IMC, they do not spawn naturally after injection, in captivity.
The sex of brooder is identified based on the morphological characteristics such as fins, abdomen, genital opening, etc. The same criteria that are used for IMC can also be used for silver carp and grass carp.
The grass carp can spawn naturally after injection if fed on artificial diet, by avoiding plants, at least two months before spawning.
To facilitate better fertilization, the eggs need to be stripped by applying pressure on the abdomen and eggs fertilized by mixing with similarly stripped milt.

Fertilization of eggs

After hormone injection, the male and female brooders are released separately in breeding hapas.
The female brooder is checked for ovulation at 3-4 hours interval.
After ovulation, the female is stripped and the eggs are collected on to a clean basin/tray, avoiding any contamination with excreta, mucus, blood, etc.
Eggs are then mixed with milt stripped from more than one male (preferably 2-3 males) to ensure better fertilization as these fishes produce comparatively less milt.
The water-hardened developing eggs are transferred to incubators for hatching.
The method of hatching of eggsand hatching period are same as that of IMC.

Major carp egg and embryonic developmental stages

The fertilization of major carp eggs is external.
The fertilized eggs absorb water and swell considerably in 10-15 minutes and they becomes a little hard and this process is known as water hardening.
They are bead-like, non-adhesive, semi-buoyant and have a large perivitelline space.
The size of the eggs varies between 2.5 and 6.5 mm.

Important major carp egg and embryonic stages and time required for each stage are as follows

Sl. no.

Time required at 24-31⁰C (Cumulative)

Stage

0 min

Fertilization

10-15 min

Water hardening and blastodisc formation

30-50 min

First cleavage (2 cells)

40-65 min

Second cleavage (4 cells)

50-80 min

Third cleavage (8 cells)

60-90 min

Fourth cleavage (16 cells)

70-120 min

Fifth cleavage (32 cells)

2-2½ hrs

Morula stage – less than half of yolk invaded by germ layer

2½-3 hrs

Blastula – about half of yolk invaded by germ layer

3-3½ hrs

Gastrula – about three-fourth of yolk is invaded by germ layer

3-4 hrs

Yolk-plug stage – complete invasion of yolk by germ layer excepting a small pore (blastopore)

4-6 hrs

Elongation of yolk mass-embryonic stage starts

5-6 hrs

Pea-shaped embryo – somites and myotomes appear

8-10 hrs

Comma-shaped embryo- optic vesicles appear, occasional twitching movement starts, gill rudiments and pectoral fin bud appear, heart appear

14-15 hrs

Advanced embryo – increase in the number of somites and myotomes, twitching movement quite rapid

18-20 hrs

Hatching – it takes place over a period of 2-3 hrs at 24-31⁰C

Causes of mortality of fish eggs and spawn and their treatment

Improper production and delivery of seed to farmers or poor management of broodfish and fish seed by farmers once stocked, may lead to decline in fish production.

Some of the causes of mortality of fish eggs and spawn are:

Poor water quality
Poor pond hygiene
Presence of Pests
Poor management of broodfish and seed
Transportation stress
Conditioning of fish seed
Mortality of eggs and hatchlings
Diseases and parasites

Poor water quality

It is known that a lot of major carp eggs perish during incubation as a result of heavy siltation (particularly so during the first few weeks after the onset of monsoon), oxygen depletion, high hydrogen sulfide level, etc.
This problem is all the more serious if reservoir water is taken directly to hatching tank.
Similarly, newly hatched spawn also suffers from the above mentioned problems.
High density of eggs and spawn also contributes to high mortality, particularly in Chinese type of carp hatchery where a large quantity of unfiltered water is used.
Use of filtered water will greatly help reduce mortality of fish eggs and spawn.

Poor pond hygiene

Mass mortality of seed in carp nursery ponds is frequently encountered in Karnataka (Mohan and Shankar, 1995).
Mortality of seed is attributed mainly to prevalence of protozoan parasites, particularly in ponds which are not dried properly prior to stocking.
Pond drying followed by liming is known to considerably reduce mortality and improve seed quality.

Presence of Pests

The presence of fairly shrimp (Streptocephalus spp) in carp nurseries is known to hamper the growth and survival by competing with fry for food, space and oxygen.
Presence of weed fishes in nursery pond leads to the production of stunted fingerlings.

Poor management of broodfish and seed

Competition among fish seed producers to meet demand some times leads to poor management of broodfish and fish seed may negatively affect seed quality.
Substandard quality seed is frequently observed as a result of high stocking density in nurseries.
Fish hatcheries in India are concerned more about the quantity rather than the quality of fish seed and produce them without following any selection norms.
Consequently, the seed suffer from high rates of mortality, poor growth and are prone to diseases and parasites.

Transportation stress

During transportation fish seed is subjected to confined environment, higher metabolic load, stress, strain and exhaustion. As a result, the seed becomes susceptible to diseases and parasites.
Chowdhury (1996) used scale loss and tail damage of carp after transportation as quality indicators.

Conditioning of fish seed

Conditioning is acclimatizing seed to a restricted environment prior to packing and transportation.
During this period the seed is stocked at a very high stocking density in a hapa or a pond with running water, but without provision for food so that the weak seed dies and only the healthy fry survives.
The seed that survives the `stress test’ only is selected for transport to a required destination. This type of conditioning of fish seed is commonly practiced in several States of India.

Mortality of eggs and hatchlings

Fish farmers in West Bengal at times encounter heavy mortality of eggs and hatchlings during incubation period.
This has been found to be due to immature bursting of egg shells and release of premature hatchlings before the anticipated period of hatching. Such hatchlings either do not survive or suffer mortality at subsequent stages.
To overcome this problem, fish farmers of the State have been using a solution which is a mixture of extract of catechu (Acacia catechu) and Myrobalan (Myrobolus indica).
The plant extract enhances hatching period and prevents hatchlings from premature release due to the presence of tannin which helps harden the egg membrane (chorion).

Diseases and parasites

High stocking density, artificial feeding, water fertilization, etc. have become common husbandry practices in carp nursery and rearing systems to optimize returns.
These high density systems offer the ideal environment for disease outbreak because such systems have stressed host and virulent pathogen.
Depending on the nature and severity, the disease may cause mass mortality of the affected population in a short time, produce protracted small scale mortality, reduce growth, make the larvae unsuitable for stocking.
The need for adopting suitable health management measures to reduce the loss due to diseases is being increasingly felt by hatchery operators.
Diseases and parasites in carp hatcheries
The nursery and rearing systems of carps are often very rich organically and provide an ideal environment for many of the pathogens. Important problems in hatcheries and early rearing systems are caused by some of the following pathogens:
Protozoan ciliates like Ichthyophthirius multifilis (white spot disease), Trichodina complex and ectodermal ciliates like Epistylis, Vorticella. All these ectoparasites can cause mass mortality of younger stages of carp very quickly and the situation becomes worst in waters with low oxygen and high organic matter.

2. Disease caused by Myxosporidians are a serious threat to the fish seed farms. These sporozoan spores present in the pond soil are normally ingested by the developing fry. Once inside the target tissue the sporozoans cause massive destruction of their target tissue and produce large scale mortality.

3. Worm parasites like Dactylogyrus (gill fluke) and Gyrodactylus (skin fluke) with their well developed attachment haptor and feeding apparatus can cause mortality in early developmental stages of carps.

4. Opportunistic secondary bacteria (Aeromonas) and fungi (Saprolegnia) can become serious problem in fish larvae which are heavily parasitized.

In carp rearing ponds major problems are because of larger ectoparasites, secondary invaders, systemic bacterial pathogens and viruses. Ectoparasites like Dactylogyrus and Argulus (fish lice) and endoparasites like sporozoans are very important. Bacterial problems like surface ulcerative conditions and acute systemic diseases are common in carp rearing systems. Many a times mortalities seen in carp culture systems are a result of ectoparasitic and systemic bacterial diseases. External fungal problems are normally associated with fish which are poorly handled. The possibility of viral diseases causing mortalities in carp nurseries cannot be ruled out.

Bacterial and fungal diseases in carp rearing systems

Bacteria can cause diseases either as secondary invaders or as primary pathogens.
Bacterial diseases in larvae can be broadly classsified as surface ulcerative, acute systemic and chronic granulomatous type.
Surface ulcerative type of diseases are characterized by haemorrhagic surface ulcers and are normally caused by Aeromonas, Pseudomanas, Vibrios, Flexibacteria, Myxobacteria, Surface ulcerative disease conditions at times develop to acute systemic disease.
These are characterized by the presence and proliferation of bacteria in internal organs like kidney, heart, spleen, blood and other visceral organs.
These diseases produce significant necrotic changes in all the affected organs and can cause mortality in a short time scale.
Bacterial haemorrhagic septicaemia caused by numerous serotypes of Aeromonas hydrophilais a major problem.

Treatment chart for common disease conditions of carp larval rearing systems

Disease agent	Chemical	Method	Concentration/time
1. Ectoparasitic protozoans
a. Ichthyophthirius sp.	Formalin	Short bath	60-100 ppm for 30 minutes
b. Trichodina sp.	Formalin	Long bath	20-30 ppm
c. Epistylis & Vorticella	Formalin	Dip	200-300 ppm for 1 minute
d. Ichthyobodos sp.	Formalin	Dip	20-30 pm for 30 minutes
2. Monogenetic worms
a. Dactylogyrus	Organophoshporus pesticides	Dip	10 ppm for < 1 minute
b. Gyrodactylus	Nuvan, Dipterex	Long bath	0.5 ppm for 24 hours
3. Crustaceans
a. Lernaea	Nuvan, Dipterex	Long bath	0.5 ppm for 24 hours
b. Argulus	Nuvan, Dipterex	Long bath	0.5 ppm for 24 hours
4. Endoparasites
a. Sporozoans	Nuvan, Dipterex	Long bath	0.5 ppm for 24 hours
b. Cercaria and metacercaria of digenetic trematodes	Nuvan, Dipterex	Long bath	0.5 ppm for 24 hours
5. External Mycosis – Saprolegnia	Malachite green Formalin	Dip Bath Bath Dip	60 ppm for < 1 minute 1-2 ppm for 1 hr 50-75 ppm for 30 minutes 100-200 ppm for 1-3 minutes
6. Surface bacterial diseases	Proflavine Oxytetracycline & Furnace	Bath Short bath	20 ppm 1-5 ppm
7. Systemic bacterial diseases	Furazolidone Oxonilic acid Chloramphenicol	In feed In feed In feed	50mg /kg fish/day 10mg/kg fish/day 50 mg/kg fish/day

Gas disease

Heavy mortality of fish seed due to `gas disease’ has been observed on several occasions. The symptoms of the disease are the presence of gas bubbles in gill filaments, heart and blood vessels and also in the gut. The air bladder gets highly distended. The `gas disease’ also referred to as ‘gas embolism’ occurs due to supersaturation of water with either oxygen or nitrogen and is generally encountered in fish nurseries.

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