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Laboratory

Reproduction Laboratory : The Piscicola Station of the Palmeiras Fund, has been operating for seven years and is a unit of Silver Corporation SAC.

Within the company we work with native species of consumption. The species of consumption that are investigated and reproduced are: The paiche, the maiden, the paco, the gamitana, the boquichico and taro of red tail, counting on a laboratory of reproduction and having developed the technology of reproduction by hormonal induction in: Piaractus Brachypomus, Colosoma macropomun, Brycon cephalus, Prochilodus nigricans and Pseudoplastystoma fasciatum, while the Arapaima gigas are being tested and tested to begin with reproduction by hormonal induction.

One of the works of the Station has to do with the basic biology of fish, ie, the knowledge of the species in the natural habitats and their classification, in the nutritive part the work is focused on improving the feeding of the fish in Systems of cultivation, consisting of creating foods that are economic and that can be made with the products of the region. The work of the Aquaculture Station is already recognized at the national level and it is attended by students and researchers from different parts of the country to participate in the different projects, then we give a brief explanation on the process of Reproduction of our tropical fish.

INDUCTION TO OVULATION AND DEVELOPMENT

Of course, ovulation of a fish is performed through the action of the pituitary gland, which produces and stores its own hormones that stimulate ovulation. When the required conditions become favorable, the stored hormone is released into the blood. In the process of hypophysis, meanwhile, the gonadotropic hormone is obtained from one fish (donor) to be applied to another fish (receptor). Hypophysis will only become effective if the ovulus has reached the latency or rest phase that follows immediately after vitellogenesis, then with the eggs structurally ready follows the action of the gonadotropin that will develop later. One condition for the success of a hypophysis is the collection of fresh pituitary glands of donor fish; It is therefore necessary that they contain an adequate amount of stored gonadotropic hormones.

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Dosage

During a natural evolution the fish is able to precisely regulate the dosage of its own hormone. That way, there is no waste. In the process of hypophysis in which the hormone comes from another external source (donor) there is considerable waste. It becomes difficult to determine an exact dose resulting in more than necessary hormone application in the reproductive system. Ovulation begins with the disappearance of the nucleus membrane and the appearance of the chromosomes, ending with the first meiotic division, at the same time, the follicle that holds the egg fixed to the wall of the ovary is broken and partially dissolves, causing The release of the eggs into the ovary cavity; In that moment the spawning takes place with the flow of the eggs through the great opening. The variation in the necessary dosage of the hormone can vary significantly for each specimen of the same species, or as per the technique used. Actually, a dosage depends on the "degree of preparation" of the females, age, size, sensitivity and many other factors, according to these, "in tropical and subtropical areas where fish metabolism is higher (due to temperature Higher) and where the likelihood of hormone wastage is therefore greater than in temperate regions, 2 or more doses, the introductory and the final, are usually given.

Dose application in females

  • - Preparatory dose: 10% of the total dose (3-4mg pituitary / kg)
  • - Final dose: 90% of the total dose
  • Application interval between 10 and 14 hours. This will vary according to the degree of preparation of the fish.

    Application of doses in males

    A single dose (1-1.5 mg / kg) is recommended. Often 2 applications are made in the attempt to obtain a greater release of semen, leaving all males subject to release sperm before the females are ready to ovulate. Excessive doses should be avoided in the preparatory application, as it may induce the fish to partial ovulation to the detriment of normal programming. In the final decisive dose, a small excess can be applied as a safety measure in the order of 10 to 15%.

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    Diluent

    Physiological saline or distilled water may be used, for the decisive dose a solvent amount of 0.5 ml should be used for each gland (2.5-3 μg). The amount of hormones is only important when it is too large or too small. When small the loss of a drop of solution would mean the loss of a considerable amount of hormone in that the administration of a large volume of solution would become a difficult problem. It is therefore recommended that the amount vary between 1-5 ml.

    Preparation of pituitary solution

    To determine the dosage has to be considered the weight, number and sex of players. To macerate the gland to mortar and porcelain arm is used. The glands are placed and moistened with the diluent, more diluent is added when convenient. The graduated syringe is used, withdraw with the syringe all the mixture that is carried to the centrifuge. After centrifugation, the excess solution is collected with the syringe. Glycerin may also be used by adding a small amount of macerate. After that, the diluent is added in the required amount and the solution is collected with a graduated syringe, this solution does not need to be centrifuged, which can be applied immediately.

    Where to apply the pituitary solution?

    Generally, the hormone is applied to the dorsal muscles above the lateral line and below the front of the dorsal fin. It is also customary to apply the hormone in the dorsal part of the caudal peduncle in case of sensitive fish. In flax fish, the introduction of the needle should be applied between the overlapping of the scales. Intraperitoneal application is also common, below the abdominal flaps, in this place the reflux of the solution is avoided when it is commonly applied to the muscles. In order to be successful in hypophysis, it is necessary to take into consideration the environmental conditions that are important, such as: adequate temperature, high free O 2 content and tranquility. When a fish receives hormonal treatment it needs more than 50% of O2, which normally, because its metabolism is more pronounced, demanding more demand for this gas, this is also valid in relation to high temperatures. The excess of manipulation results in greater consumption of O2, being therefore necessary to keep the fish in calm environments avoiding traumas and with constant and abundant renewal of water.

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    Use of human chorionic gonadotropin (HCG)

    The pre-ovulation phase can generally be readily obtained by administering GCH, however, it is more difficult to achieve full ovulation through this method in most fish, although many fish react positively. Well-prepared fish are those that offer conditions for spawning with the use of GCH. For Colossoma, we recommend: 1a. Dose: 2 mg pituitary / 1 kg body weight 2a. Dose: 5 U.I / g body weight You can go to a 3rd. Dose with 5 U.I./g. Application interval: 12 hours. Males are injected when females receive their second application at a dose of 5 U.I./g.

    Grade time

    As normally the final process of maturing of the eggs depends on the temperature of the water, it takes some time to do so. From the time interval between the last application of hormone and total ovulation, that time interval is expressed in "hour degree". Each hour is measured after the last injection until complete ovulation. The hourly readings are added to get the "grade time", this helps the farmer know exactly when to ovulate from the last injection. The value of the "degree time" depends on the species of the fish, the type of treatment, the size of the female and whether the fish begins to spawn immediately after ovulation or not. In the case of Colossoma at a temperature of 28 ° C, the degree time is 250-300.

    Extrusion of sexual products

    Some fish do not spawn naturally, even though they are induced to spawn with applications of pituitary serum, as in the case of paco and colossoma reaches this state. However, for a better use of the sexual products, the collection of this material can be made by extrusion. The ovulation or final process of maturing of the eggs can not be interrupted or canceled. Once the process has begun the eggs have to be extruded or subjected to the extrusion process; If that does not happen, it becomes too mature, without the condition of being fertilized. In this way, every female in this condition has to undergo this process, because the oocytes already reach the maturation stage that occurs immediately. This maturation undergoes variations according to the species, being that in tropical fish this happens earlier than in the fish of temperate zones. Most ovules mature or develop and fall into the ovary cavity at the same time, this facilitates extrusion so that they flow easily. The hour-grade knowledge of the fish, helps to establish the exact time of ovulation, within a variation of 10-20 minutes on average. In saturated females, the sagging of the belly and the presence of some eggs between the saturas would indicate the maturation, being able to perform the extrusion, sometimes the males are used as indicators for the mature females, they begin the courtship to the mature females Flanking them constantly. In order to collect the eggs, clean and dry plastic tanks are used. Dry the female with a soft towel, removing the urine and taking care of the sutures. Mature females let the eggs flow in firm, thick jets. Only the last litter of eggs that need to undergo slight pressure is removed for complete withdrawal. A forced extraction indicates that the fish is not in condition and that should be avoided, as the eggs would not be submitted to fertilization. Simultaneously, the semen of the males is combined with the use of a special pipette. You can also collect the semen directly from the male, depositing on the collected eggs by slowly mixing the products with a duck feather or a plastic spoon; In that case they have to be cleaned and wiped the males. Shortly after adding water and with a certain period of 5 to 10 minutes, the already fertilized eggs are placed in incubators. Incubators with capacity for 100 to 200 liters are the most indicated. Never place more than 300,000 eggs in a 200 liter incubator. At birth, larvae may have problems with O2 dissolved in water and may die.

    Incubation of eggs

    This phase is of importance, because the survival of the eggs will depend on their good development. Therefore, the need for special care during this development, with temperature, oxygen, etc., which may intervene in the quality of the same is necessary. During this phase a high concentration of oxygen is required, being that in the initial states the consumption is insignificant, becoming more accentuated as its development advances. The temperature plays a very important role, having to be in levels identical to those of the environment of origin of the fish, avoiding minimum or maximum excesses, as these would jeopardize the success of its normal development. Maintaining a constant flow of water will eliminate the toxic substances produced by eggs, such as CO2 and NH3 that could compromise their life. However, the flow of water must be controlled to avoid mechanical cokes that could at risk the eggs of the fish, in their early stages of cell segmentation to morula are extremely sensitive. In addition, gaseous changes are more intense when eggs enter the blastula phase, requiring a higher concentration of oxygen. When it comes to luminosity, eggs and larvae of some species of fish are sensitive to ultraviolet rays, as a consequence, they must be protected from excess light.

    Incubators

    There are several types of incubators that can be used in the incubation of eggs and larvae of fish of the genus Colossoma, which have free eggs and have a density greater than that of water. However, the ideal shape is the conical, with part of the cylindrical body (10%) and the remaining conical (90%) that format helps considerably to keep the larvae in constant movement, thus preventing the eggs from depositing in the bottom of The incubator, which would be detrimental to them. Before absorption of the yolk sac the movement of larvae is vertical with periodic impulses. It is advisable to use incubators made of fiberglass, because they are resistant and light, easy to be transported, zinc incubators can be used as long as the inner part is painted neutral. Both can be handcrafted to significantly reduce costs. The water inlet should be placed in the bottom of the incubator with the use of a domestic shower, so that the flow of moving water is vertical, from the bottom up. The water outlet will be given above the incubator, being careful to use a mesh fabric of 350-400 microns, in order to prevent the escape of eggs or larvae allowing with this that the flow of the water is not interrupted. Meshes of this dimension will favor the passage of residues from the disintegration of the egg chamber. The attachment of this flow will cause the water to flow with the inevitable loss of the incubation material.

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    Filtration of water for incubation

    One of the serious problems during the incubation phase is the presence of undesirable organisms such as Cyclops that cause damage to fish eggs and larvae, causing them to die. The construction of a filter capable of retaining such organisms becomes necessary. It must be built in bricks and concrete, consisting of individual sections with coarse sand and solid rods with dimensions according to the needs of use the water intended for the incubators must pass through that filter.

    Permanence of larvae in incubators

    Fish larvae of the genus Colossoma maintain the yolk sac until the 5th day of life. Meanwhile, in the 3rd it is already greatly reduced in that phase, since the fish already has mouth opening, feeding simultaneously the internal and external environment, nothing horizontal and seeks food from the environment, and can be given to larvae, egg Cooked liquefied several times a day. The ration of the egg should not exceed the 3rd day, as it will cause the death of the larvae that would have difficulty digesting them and whose use is made to distend the intestine facilitating the passage of food. Filled zooplankton can be offered in 60 to 80 micron meshes, yet it is a slow and costly job. Ideally, in which the larvae will reside, they will find abundant food, indispensable for a rapid development, avoiding possible problems due to the deficiency of feeding.

    Cultivation of larvae in nursery

    Here basically lies the success of all upbringing, and involves special care. Placement of larvae in nursery, will occur between the 3rd and 4th days of life, often to avoid predation of copepods larvae are kept in incubators not more than 48 hours. At that stage, the nutritional reserve is still sufficient to support the larva. The best time to populate the nursery will be in the morning, when the temperature is low and without environmental shock problems. The collection of larvae will be done by means of siphoning system and plastic hoses, from the incubator to buckets of the same material, being careful to avoid mechanical shocks during that manipulation can also be used plastic bags. When bringing the containers with the larvae to the nursery, the water should be allowed to flow slowly on the shelves, in order to balance the water temperature in that way, the larvae are also placed in the nursery, returning from the bucket or sack Water.