Seedling cultivation method for Jinhu hybrid spots

Abstract

The invention provides a method for cultivating hybrid fries of Jinhu and tiger. The method comprises the following steps: selecting a breeding environment; breeding and ripening female parents; management and control before oviposition; insemination; hatching fertilized eggs; fry rearing. According to the method, in the female parent breeding and ripening stage, specific biological bait is controlled to be fed in stages, feeding of the parent fish can be promoted, growth can be accelerated, in cooperation with specific compound feed in the management and control stage before oviposition, favorable conditions are created for subsequent oviposition, and the quality and number of ova can be improved. Low-frequency ultrasonic assisted fertilization is introduced in the artificial insemination stage, so that the activity of ovum sperms can be obviously improved, and the fertilization rate is increased; in the fertilized egg hatching stage, intermittent ultrasound is assisted, the hatching rate can be increased, and the seedling yield of the golden-tiger hybrid spots can be increased; after hatching, transferring into a specific culture pond, slowly releasing beneficial bacteria, improving the adaptability of the fry, adsorbing harmful substances, reducing pathogenic bacteria, stabilizing water quality, promoting the growth of the fry, improving the survival rate and reducing the malformation rate.

Description

Technical Field

[0001] The present invention relates to the technical field of aquaculture, and particularly relates to a method for culturing fry of hybrid grouper Epinephelus fuscoguttatus♀×Epinephelus lanceolatus♂. Background Art

[0002] Grouper is an important germplasm resource of marine fish in China and plays a pillar role in the industry. The annual demand for fry reaches 1 billion tails. However, problems such as many diseases and low survival rate of grouper fry have become bottlenecks in the industry's development. By constructing a technology for improving grouper varieties and cultivating excellent germplasms with fast growth and strong stress resistance, the problems of many diseases and low survival rate of fry in the industry can be solved. "Hybrid grouper Epinephelus fuscoguttatus♀×Epinephelus lanceolatus♂" is a hybrid variety bred with Epinephelus fuscoguttatus as the female parent and Epinephelus lanceolatus (Epinephelus tukula) as the male parent, and has the characteristics of fast growth rate, smaller coefficient of variation of body length and body weight, and stable inheritance of traits. Hybrid grouper Epinephelus fuscoguttatus♀×Epinephelus lanceolatus♂ will provide high-quality suitable cultured varieties for modern industrialized and deep-sea aquaculture in China, and will strongly promote the innovation of marine fishery seed industry and the sustainable development of the industry.

[0003] At present, the artificial breeding technology of this hybrid variety is not yet mature. The broodstock has difficulty in spawning, poor quality of sperm and eggs, low hatching rate of fertilized eggs, low survival rate of fry, some larvae are deformed, and the disease resistance is poor, which has a certain impact on the fry yield and quality of hybrid grouper Epinephelus fuscoguttatus♀×Epinephelus lanceolatus♂, and greatly restricts the large-scale breeding process of large grouper.

[0004] Therefore, it is necessary to further optimize the process of culturing grouper fry, which can improve the fry yield and quality of hybrid grouper Epinephelus fuscoguttatus♀×Epinephelus lanceolatus♂, promote the healthy development of the grouper industry, and promote the large-scale breeding process of grouper, which has practical significance. Summary of the Invention

[0005] In view of this, the present invention proposes a method for culturing fry of hybrid grouper Epinephelus fuscoguttatus♀×Epinephelus lanceolatus♂.

[0006] The technical solution of the present invention is realized as follows:

[0007] A method for culturing fry of hybrid grouper Epinephelus fuscoguttatus♀×Epinephelus lanceolatus♂, comprising the following steps:

[0008] S1. Optimize the breeding environment: Select a pond with a water temperature of 22-25°C and a water depth of 2-3 meters, and put the female parent Epinephelus fuscoguttatus;

[0009] S2. Cultivate and ripen the female parent: In the first stage, feed biological bait A, and the feeding amount is 4%-6% of the body weight of the female parent, 3-4 times a day, for 4-6 days. In the second stage, mix and feed biological bait B and nutritional preparation, and the feeding amount is 5%-7% of the body weight of the female parent, 2-3 times a day, for 3-5 days;

[0010] S3. Pre-spawning control: 5 - 7 days before the female parent spawns, raise the water temperature to 27 - 28 °C, feed with compound feed, and the feeding amount is 1% - 3% of the weight of the female parent, twice a day;

[0011] S4. Fertilization: After the female parent spawns, under the condition of a low-frequency ultrasonic field, fertilize with the sperm of Epinephelus tukula to obtain fertilized eggs;

[0012] S5. Incubation of fertilized eggs: Place the fertilized eggs in an incubator, perform intermittent ultrasonic treatment, and hatch to obtain fry;

[0013] S6. Fry cultivation: Select a culture pond, put in probiotic magnetic chitosan microspheres, and transfer the fry to the culture pond for preliminary cultivation.

[0014] Further, in step S1, the female parent Epinephelus fuscoguttatus is a healthy Epinephelus fuscoguttatus over 6 years old without diseases, and the stocking density is 100 - 200 tails / m 2 .

[0015] Further, in step S2, in the first stage, feed biological bait A, which includes the following raw materials by weight: 30 - 40 parts of fish meal, 5 - 10 parts of rotifers, 1 - 3 parts of vitamins, 1 - 3 parts of amino acids, 4 - 6 parts of olive fruit extract, 1 - 3 parts of withania somnifera polysaccharide, 4 - 6 parts of dendrobium nobile stem extract, 2 - 4 parts of coconut oil.

[0016] Further, the olive fruit extract is olive fruit water extract and / or olive fruit ethanol extract; the dendrobium nobile stem extract is dendrobium nobile stem ethanol extract and / or dendrobium nobile stem ethyl acetate extract.

[0017] Further, in step S2, in the second stage, feed biological bait B, which includes the following raw materials by weight: 20 - 30 parts of fish meal, 4 - 6 parts of shrimp powder, 1 - 3 parts of vitamins, 1 - 3 parts of camellia oleifera leaf polysaccharide, 1 - 3 parts of lentinan, 1 - 2 parts of date plum fruit extract, 3 - 5 parts of arundina graminifolia extract, 2 - 4 parts of vegetable blend oil.

[0018] Further, the date plum fruit extract is date plum fruit water extract and / or date plum fruit ethanol extract; the arundina graminifolia extract is arundina graminifolia water extract and / or arundina graminifolia ethanol extract.

[0019] Further, in step S2, the nutritional preparation includes the following raw materials by weight: 3 - 5 parts of earthworm fermentation broth, 3 - 5 parts of chitin fermentation broth;

[0020] The preparation method of the nutritional preparation is: Treat the earthworm fermentation broth and the chitin fermentation broth separately through a dynamic high-pressure microfluidic homogenizer at 80 - 120 MPa for 2 times, and mix to obtain.

[0021] Furthermore, the earthworm fermentation liquid is obtained by beating earthworms and then stirring them with lipase and ultrasound; the chitin fermentation liquid is obtained by crushing shrimp shells and then fermenting them with Trichoderma harzianum and Bacillus, the mass ratio of Trichoderma harzianum to Bacillus is preferably 1:1, and the fermentation temperature is preferably 35-45°C.

[0022] Furthermore, in step S3, the compound feed comprises the following raw materials in parts by weight: 10-20 parts of shrimp meal, 5-8 parts of clam pulp, 2-4 parts of carnosic acid nutrient solution, 3-4 parts of wild Ganoderma lucidum extract, 1-3 parts of Perilla leaf extract, 1-3 parts of Morinda citrifolia fruit extract, and 1-3 parts of longan polysaccharide.

[0023] Furthermore, the wild Ganoderma lucidum extract is a wild Ganoderma lucidum water extract and / or a wild Ganoderma lucidum ethanol extract; the Perilla leaf extract is a Perilla leaf water extract and / or a Perilla leaf ethanol extract; the Morinda citrifolia fruit extract is a Morinda citrifolia fruit ethanol extract and / or a Morinda citrifolia fruit ethyl acetate extract.

[0024] Furthermore, in step S4, the low-frequency ultrasonic field treatment is specifically as follows: ultrasonication at 20-30 kHz, 10-15 W for 20-30 seconds, after a 30-second pause, ultrasonication is continued for 20-30 seconds, and the above process is repeated until fertilization is completed.

[0025] Furthermore, in step S5, the intermittent ultrasonic treatment is first ultrasonicated at 20-30kHz, 20-30W for 30-60s, with a rest period of 3min, and then ultrasonicated at 50-60kHz, 50-60W for 20-40s, with a rest period of 5min, and the above process is repeated 3-5 times.

[0026] Furthermore, in step S6, the preparation method of the probiotic magnetic chitosan microspheres includes: adding chitosan to water, transferring to a high-pressure reactor, performing a hydrothermal reaction at 160-200° C. for 12-24 hours, cooling, washing, and drying, dispersing the product in deionized water, adding ferric chloride and PVP and stirring, adding concentrated ammonia water at 70-80° C., continuing to stir for 1-2 hours, cooling, centrifuging the microspheres, washing, and drying to obtain magnetic chitosan microspheres, and encapsulating the probiotics in the magnetic chitosan microspheres by a swelling-adsorption method;

[0027] The solid-liquid ratio of chitosan to water is 1:10-20 g / mL, and the solid-liquid ratio of the product to deionized water is 1:10-20 g / mL; the mass ratio of ferric chloride, PVP and product is 1-2:0.5-0.7:5; the amount of concentrated ammonia added is 0.2-0.4 times the mass of the product, and the concentration is 20-30 wt%; the loading amount of the probiotics on the magnetic chitosan microspheres is 20%-30%.

[0028] Further, in step S6, during the preliminary cultivation, the salinity of the breeding pond is controlled at 15-25‰, the temperature is 25-28°C, the pH value is 7.5-8.0, and the dissolved oxygen is greater than 6.0 mg / L.

[0029] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0030] 1. In the female parent cultivation and ripening stage of the present invention, by controlling the feeding of specific biological baits in stages, it can not only promote the feeding of broodstock, accelerate growth, and cooperate with the specific compound feed in the pre-spawning control stage to create favorable conditions for subsequent spawning, but also improve the quality and quantity of eggs.

[0031] 2. In the artificial insemination stage of the present invention, low-frequency ultrasonic-assisted fertilization is introduced, which can significantly improve the vitality of eggs and sperm and increase the fertilization rate; in the fertilized egg hatching stage, intermittent ultrasound is supplemented, which can improve the hatching rate and increase the fry yield of golden tiger hybrid grouper.

[0032] 3. After the fry of the present invention hatches, it is transferred to a breeding pond with beneficial probiotic magnetic chitosan microspheres, which slowly release beneficial bacteria, reduce loss, improve the adaptability of fry, adsorb harmful substances, reduce pathogenic bacteria, stabilize water quality, promote the growth of fry, increase the survival rate, reduce the malformation rate, improve the fry quality of golden tiger hybrid grouper. In addition, the release position of probiotics can be controlled by the action of magnetic field, so that the probiotics can be evenly distributed. Specific embodiments

[0033] In order to better understand the technical content of the present invention, specific embodiments are provided below to further illustrate the present invention.

[0034] The experimental methods used in the embodiments of the present invention are all conventional methods unless otherwise specified.

[0035] The materials, reagents, etc. used in the embodiments of the present invention can be obtained from commercial channels unless otherwise specified.

[0036] The preparation of biological bait B of the present invention is to mix the various raw materials evenly, extrude and granulate, and then grind and screen to obtain; the preparation of biological bait A is to mix the various raw materials evenly, extrude and granulate, and then grind and screen to obtain; the preparation of compound feed is to mix the various raw materials evenly, extrude and granulate, and then grind and screen to obtain.

[0037] The olive fruit extract of the present invention is an aqueous extract of olive fruit and / or an ethanol extract of olive fruit; the Dendrobium nobile Lindl. stem extract is an ethanol extract of Dendrobium nobile Lindl. stem and / or an ethyl acetate extract of Dendrobium nobile Lindl. stem; the date plum fruit extract is an aqueous extract of date plum fruit and / or an ethanol extract of date plum fruit; the Arundina graminifolia extract is an aqueous extract of Arundina graminifolia and / or an ethanol extract of Arundina graminifolia; the Ganoderma lucidum extract is an aqueous extract of Ganoderma lucidum and / or an ethanol extract of Ganoderma lucidum; the Perilla leaf extract is an aqueous extract of Perilla leaf and / or an ethanol extract of Perilla leaf; the Morinda citrifolia fruit extract is an ethanol extract of Morinda citrifolia fruit and / or an ethyl acetate extract of Morinda citrifolia fruit.

[0038] The earthworm fermentation broth of the present invention is prepared by pulping earthworms and then subjecting them to ultrasonic stirring with lipase; the chitin fermentation broth is obtained by pulverizing shrimp shells and then fermenting them with Trichoderma harzianum and Bacillus sp., and the mass ratio of Trichoderma harzianum to Bacillus sp. is preferably 1:1, and the fermentation temperature is preferably 35 - 45°C.

[0039] Example 1

[0040] The method for cultivating the seedlings of the tiger grouper hybrid includes the following steps:

[0041] S1. Optimize the breeding environment: Select a pond with a water temperature of 23°C and a water depth of 2 meters, and release healthy and disease-free Epinephelus fuscoguttatus over 6 years old, with a density of 150 tails / m 2 ;

[0042] S2. Cultivate and ripen female parents: In the first stage, feed biological bait A at a feeding amount of 5% of the weight of the female parents, 3 times a day for 5 days. In the second stage, mix and feed biological bait B and nutritional preparation at a feeding amount of 6% of the weight of the female parents, 3 times a day for 4 days;

[0043] Biological bait A, by weight, includes the following raw materials: 35 parts of fish meal, 8 parts of rotifers, 2 parts of vitamins, 2 parts of amino acids, 5 parts of olive fruit extract, 2 parts of withania somnifera polysaccharide, 5 parts of Dendrobium nobile Lindl. stem extract, 3 parts of coconut oil;

[0044] Biological bait B, by weight, includes the following raw materials: 25 parts of fish meal, 5 parts of shrimp powder, 2 parts of vitamins, 2 parts of camellia oleifera polysaccharide, 2 parts of lentinan, 1.5 parts of date plum fruit extract, 4 parts of Arundina graminifolia extract, 3 parts of vegetable blended oil;

[0045] The nutritional preparation, by weight, includes the following raw materials: 4 parts of earthworm fermentation broth, 4 parts of chitin fermentation broth; the nutritional preparation is prepared by separately treating the earthworm fermentation broth and the chitin fermentation broth 2 times with a dynamic high-pressure microfluidizer at 100 MPa and then mixing them;

[0046] S3. Pre-spawning control: 6 days before the female parent lays eggs, raise the water temperature to 27°C and feed the female parent with a compound feed at a rate of 2% of her body weight, twice a day.

[0047] The compound feed comprises the following ingredients in parts by weight: 15 parts of shrimp meal, 7 parts of clam pulp, 3 parts of carnosic acid nutrient solution, 3.5 parts of wild Ganoderma lucidum extract, 2 parts of Perilla leaf extract, 2 parts of Morinda citrifolia fruit extract, and 2 parts of longan polysaccharide;

[0048] S4. Fertilization: After the female parent lays eggs, they are fertilized with sperm from the blue-bodied giant grouper under the conditions of a low-frequency ultrasonic field to obtain fertilized eggs;

[0049] The specific conditions of the low-frequency ultrasound field are as follows: ultrasound at 25 kHz, 13 W for 25 s, a 30-s rest, and then ultrasound for 25 s, and the above process is repeated until fertilization is completed;

[0050] S5, fertilized egg hatching: the fertilized eggs were placed in an incubator, first ultrasonicated at 25kHz, 25W for 40s, with a rest period of 3min, then ultrasonicated at 55kHz, 55W for 30s, with a rest period of 5min, and the above process was repeated 4 times to hatch the fry;

[0051] S6. Fry cultivation: Select a culture pond with a salinity of 20‰, a temperature of 27°C, a pH of 7.7±0.2, and a dissolved oxygen level greater than 6.0 mg / L, add probiotic magnetic chitosan microspheres, transfer the fry into the culture pond, and carry out preliminary cultivation;

[0052] Among them, the preparation of probiotic magnetic chitosan microspheres: chitosan is added to water according to a solid-liquid ratio of 1:15g / mL, and then transferred to a high-pressure reactor, hydrothermally reacted at 180°C for 12h, cooled, washed and dried, and the product is dispersed in deionized water according to a solid-liquid ratio of 1:15g / mL, and ferric chloride and PVP are added. The mass ratio of ferric chloride, PVP and product is 1:0.6:5, and stirred. 25wt% concentrated ammonia water is added at 75°C, and the amount of concentrated ammonia water added is 0.3 times the mass of the product. Stirring is continued for 1.5h, cooled, centrifuged to separate the microspheres, washed and dried to obtain magnetic chitosan microspheres, and the probiotics are encapsulated in the magnetic chitosan microspheres by the swelling-adsorption method. The loading amount of probiotics in the magnetic chitosan microspheres is 25%.

[0053] Example 2

[0054] The method for cultivating seedlings of hybrid golden tiger spot comprises the following steps:

[0055] S1. Optimize the breeding environment: Select a pond with a water temperature of 22°C and a water depth of 2 meters, and stock it with healthy and disease-free brown-spotted groupers over 6 years old at a density of 100 per m 2 ;

[0056] S2. Female parent cultivation and ripening: In the first stage, feed biological bait A at a feeding amount of 4% of the weight of the female parent, 3 times a day for 4 days. In the second stage, mix and feed biological bait B and nutritional preparation at a feeding amount of 5% of the weight of the female parent, 2 times a day for 3 days;

[0057] Biological bait A, by weight parts, includes the following raw materials: 30 parts of fish meal, 5 parts of rotifers, 1 part of vitamins, 1 part of amino acids, 4 parts of olive fruit extract, 1 part of withania somnifera polysaccharide, 4 parts of dendrobium nobile stem extract, 2 parts of coconut oil;

[0058] Biological bait B, by weight parts, includes the following raw materials: 20 parts of fish meal, 4 parts of shrimp powder, 1 part of vitamins, 1 part of camellia oleifera leaf polysaccharide, 1 part of lentinan, 1 part of diospyros lotus fruit extract, 3 parts of arundina graminifolia extract, 2 parts of vegetable blend oil;

[0059] Nutritional preparation, by weight parts, includes the following raw materials: 3 parts of earthworm fermentation broth, 3 parts of chitin fermentation broth; The nutritional preparation is prepared by separately treating the earthworm fermentation broth and chitin fermentation broth with a dynamic high-pressure microfluidic homogenizer at 80 MPa for 2 times and then mixing them;

[0060] S3. Pre-spawning control: 5 days before the female parent lays eggs, raise the water temperature to 27 °C and feed with compound feed at a feeding amount of 1% of the weight of the female parent, 2 times a day;

[0061] Compound feed, by weight parts, includes the following raw materials: 10 parts of shrimp powder, 5 parts of clam meat slurry, 2 parts of carnosic acid nutrient solution, 3 parts of wild ganoderma extract, 1 part of perilla leaf extract, 1 part of morinda citrifolia fruit extract, 1 part of longan polysaccharide;

[0062] S4. Fertilization: After the female parent lays eggs, under the condition of a low-frequency ultrasonic field, fertilize with the sperm of Epinephelus tukula to obtain fertilized eggs;

[0063] The conditions of the low-frequency ultrasonic field are specifically as follows: Ultrasonic at 20 kHz and 10 W for 20 s, after an interval of 30 s, continue to ultrasonic for 20 s, and repeat the above process until fertilization is completed;

[0064] S5. Incubation of fertilized eggs: Place the fertilized eggs in an incubator, first ultrasonic at 20 kHz and 20 W for 30 s, with an interval of 3 min, then ultrasonic at 50 kHz and 50 W for 20 s, with an interval of 5 min, and repeat the above process 3 times to hatch fry;

[0065] S6. Fry cultivation: Select a culture pond with a salinity of 15‰, a temperature of 25 °C, a pH value of 7.7 ± 0.2, and a dissolved oxygen greater than 6.0 mg / L, put in probiotic magnetic chitosan microspheres, transfer the fry to the culture pond, and conduct preliminary cultivation;

[0066] Among them, the preparation of probiotic magnetic chitosan microspheres: Chitosan was added to water according to a solid-liquid ratio of 1:10 g / mL, and then transferred to a high-pressure reactor for hydrothermal reaction at $160^{\circ}C$ for 12 h. After cooling, washing and drying, the product was dispersed in deionized water according to a solid-liquid ratio of 1:10 g / mL, and ferric chloride and PVP were added. The mass ratio of ferric chloride, PVP to the product was 1:0.5:5, and stirring was carried out. 20 wt% concentrated ammonia water was added at $70^{\circ}C$, and the addition amount of concentrated ammonia water was 0.2 times the mass of the product. Stirring was continued for 1 h, followed by cooling and centrifuging to separate the microspheres, washing and drying to obtain magnetic chitosan microspheres. The probiotics were encapsulated in the magnetic chitosan microspheres by the swelling-adsorption method, and the loading amount of the probiotics in the magnetic chitosan microspheres was 20%.

[0067] Example 3

[0068] The fry cultivation method of hybrid grouper includes the following steps:

[0069] S1. Select the breeding environment: Select a pond with a water temperature of $25^{\circ}C$ and a water depth of 3 m, and release healthy and disease-free Epinephelus fuscoguttatus over 6 years old, with a density of 200 tails / m 2 ;

[0070] S2. Cultivate and ripen female parents: In the first stage, feed biological bait A at a feeding amount of 6% of the weight of the female parents, 4 times a day for 6 days. In the second stage, mix and feed biological bait B and nutritional preparation at a feeding amount of 7% of the weight of the female parents, 3 times a day for 5 days;

[0071] Biological bait A, by weight, includes the following raw materials: 40 parts of fish meal, 10 parts of rotifers, 3 parts of vitamins, 3 parts of amino acids, 6 parts of olive fruit extract, 3 parts of withania somnifera polysaccharide, 6 parts of dendrobium officinale stem extract, 4 parts of coconut oil;

[0072] Biological bait B, by weight, includes the following raw materials: 30 parts of fish meal, 6 parts of shrimp meal, 3 parts of vitamins, 3 parts of camellia oleifera leaf polysaccharide, 3 parts of lentinan, 2 parts of diospyros lotus fruit extract, 5 parts of arundina graminifolia extract, 4 parts of vegetable blended oil;

[0073] Nutritional preparation, by weight, includes the following raw materials: 5 parts of earthworm fermentation broth, 5 parts of chitin fermentation broth; The nutritional preparation is prepared by separately treating the earthworm fermentation broth and the chitin fermentation broth 2 times with a dynamic high-pressure microfluidizer at 120 MPa and then mixing them;

[0074] S3. Control before spawning: 7 days before the female parents spawn, raise the water temperature to $28^{\circ}C$ and feed with compound feed at a feeding amount of 3% of the weight of the female parents, 2 times a day;

[0075] The compound feed comprises the following ingredients in parts by weight: 20 parts shrimp meal, 8 parts clam pulp, 4 parts carnosic acid nutrient solution, 4 parts wild Ganoderma lucidum extract, 3 parts Perilla leaf extract, 3 parts Morinda citrifolia fruit extract, and 3 parts longan polysaccharide;

[0076] S4. Fertilization: After the female parent lays eggs, they are fertilized with sperm from the blue-bodied giant grouper under the conditions of a low-frequency ultrasonic field to obtain fertilized eggs;

[0077] The specific conditions of the low-frequency ultrasound field are as follows: ultrasound at 30 kHz, 15 W for 30 s, rest for 30 s, and then continue ultrasound for 30 s, and repeat the above process until fertilization is completed;

[0078] S5, fertilized egg hatching: the fertilized eggs were placed in an incubator, first ultrasonicated at 30kHz, 30W for 60s, with a rest period of 3min, then ultrasonicated at 60kHz, 60W for 40s, with a rest period of 5min, and the above process was repeated 5 times to hatch the fry;

[0079] S6. Fry cultivation: Select a culture pond with a salinity of 25‰, a temperature of 28°C, a pH of 7.7±0.2, and a dissolved oxygen level greater than 6.0 mg / L, add probiotic magnetic chitosan microspheres, transfer the fry into the culture pond, and carry out preliminary cultivation;

[0080] Among them, the preparation of probiotic magnetic chitosan microspheres: chitosan is added to water according to a solid-liquid ratio of 1:20 g / mL, and then transferred to a high-pressure reactor, hydrothermally reacted at 200°C for 24 hours, cooled, washed and dried, and the product is dispersed in deionized water according to a solid-liquid ratio of 1:20 g / mL, and ferric chloride and PVP are added. The mass ratio of ferric chloride, PVP and product is 2:0.7:5, and stirred. 30wt% concentrated ammonia water is added at 80°C, and the amount of concentrated ammonia water added is 0.4 times the mass of the product. Stirring is continued for 2 hours, cooled, centrifuged to separate the microspheres, washed and dried to obtain magnetic chitosan microspheres, and the probiotics are encapsulated in the magnetic chitosan microspheres by the swelling-adsorption method. The loading amount of probiotics in the magnetic chitosan microspheres is 30%.

[0081] Comparative Example 1

[0082] The difference from Example 1 is that the biological bait A in step S2 lacks green fruit extract and Withania somnifera polysaccharide, and the rest is consistent with Example 1.

[0083] Comparative Example 2

[0084] The difference from Example 1 is that in the second stage of step S2, only biological bait B is fed, and the rest is the same as Example 1.

[0085] Comparative Example 3

[0086] The difference from Example 1 is that in the pre-spawning control in step S3, the compound feed is replaced with biological bait B and nutritional preparations, and the rest is the same as Example 1.

[0087] Comparative Example 4

[0088] The difference from Example 1 is that the low-frequency ultrasonic field is missing in step S4, and the rest is consistent with Example 1.

[0089] Comparative Example 5

[0090] The difference from Example 1 is that intermittent ultrasonic treatment is missing in step S5, and the rest is consistent with Example 1.

[0091] Comparative Example 6

[0092] The difference from Example 1 is that in step S6, the probiotic magnetic chitosan microspheres are replaced with probiotics, and the rest is the same as Example 1.

[0093] Test Case

[0094] According to the breeding methods of Examples 1-3 and Comparative Examples 1-6, the golden tiger hybrid grouper was bred. Experimental groups 1-9 were set, corresponding to Examples 1-3 and Comparative Examples 1-6, respectively. Each group included 300 female parent brown-spotted groupers aged 6 or above, and golden tiger hybrid grouper fry were obtained according to the corresponding breeding methods.

[0095] 1. Measure the egg production, fertilization rate, hatching rate and fry deformity rate respectively.

[0096] The results are shown in Table 1.

[0097] Table 1

[0098] Group Spawning amount Fertilization rate Hatching rate Malformation rate of fry Example 1 143.4 Kg 99.5％ 97.5％ 0％ Example 2 138.6 Kg 99.3％ 97.2％ 0％ Example 3 135.7 Kg 99.1％ 96.9％ 0％ Comparative Example 1 112.5 Kg 92.8％ 92.2％ 0.8％ Comparative Example 2 98.4 Kg 91.1％ 91.5％ 1.1％ Comparative Example 3 105.6 Kg 90.3％ 88.9％ 1.4％ Comparative Example 4 - 80.2％ 87.1％ 1.5％ Comparative Example 5 - - 80.6％ 2.4％ Comparative Example 6 - - - -

[0099] As can be seen from Table 1, the breeding methods of Examples 1-3 of the present invention significantly increased egg production and egg quality, fertilization rate, and hatching rate by feeding specific feeds, and the hatched fry showed no deformities. Comparative Examples 1-3, which varied the feed formula composition, resulted in decreased egg production and egg quality, and subsequently affected fertilization rate, hatching rate, and deformity rate.

[0100] 2. After 5 months of breeding, the survival rate and morbidity rate were calculated and the results are shown in Table 2.

[0101] Table 2

[0102]

[0103]

[0104] As can be seen from Table 2, the artificial cultivation method of the present invention can significantly improve the survival rate of fry, improve the quality of healthy fry, and the fry cultured for 5 months are almost disease-free.

[0105] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention shall be included within the protection scope of the present invention.

Claims

1. A method for culturing fry of hybrid tiger grouper, characterized in that, It includes the following steps: S1. Select a breeding environment: Select a pond with a water temperature of 22 - 25°C and a water depth of 2 - 3 meters, and release female parent Epinephelus fuscoguttatus. S2. Cultivate and ripen the female parent: In the first stage, feed biological bait A, and the feeding amount is 4% - 6% of the weight of the female parent, 3 - 4 times a day for 4 - 6 days. In the second stage, mix and feed biological bait B and a nutritional preparation, and the feeding amount is 5% - 7% of the weight of the female parent, 2 - 3 times a day for 3 - 5 days. S3. Control before spawning: 5 - 7 days before the female parent spawns, raise the water temperature to 27 - 28°C, and feed with compound feed. The feeding amount is 1% - 3% of the weight of the female parent, 2 times a day. S4. Fertilization: After the female parent spawns, under the condition of a low-frequency ultrasonic field, fertilize with the sperm of Epinephelus lanceolatus to obtain fertilized eggs. S5. Incubate the fertilized eggs: Place the fertilized eggs in an incubator and perform intermittent ultrasonic treatment to hatch fry. S6. Cultivate the fry: Select a culture pond, put in probiotic magnetic chitosan microspheres, and transfer the fry to the culture pond for preliminary cultivation.

2. The method for cultivating fry of the golden tiger hybrid grouper according to claim 1, characterized in that In step S1, the female parent Epinephelus fuscoguttatus is an Epinephelus fuscoguttatus over 6 years old, healthy and disease-free, and the stocking density is 100 - 200 tails / m 2 .

3. The method for cultivating fry of the hybrid tiger grouper as described in claim 1, characterized in that, In step S2, when feeding biological bait A in the first stage, according to weight parts, it includes the following raw materials: 30 - 40 parts of fish meal, 5 - 10 parts of rotifers, 1 - 3 parts of vitamins, 1 - 3 parts of amino acids, 4 - 6 parts of olive fruit extract, 1 - 3 parts of withania somnifera polysaccharide, 4 - 6 parts of dendrobium officinale stem extract, 2 - 4 parts of coconut oil.

4. The method for cultivating the fry of the golden tiger hybrid grouper as described in claim 1, characterized in that, In step S2, when feeding biological bait B in the second stage, according to weight parts, it includes the following raw materials: 20 - 30 parts of fish meal, 4 - 6 parts of shrimp meal, 1 - 3 parts of vitamins, 1 - 3 parts of camellia oleifera leaf polysaccharide, 1 - 3 parts of lentinan, 1 - 2 parts of diospyros lotus fruit extract, 3 - 5 parts of arundina graminifolia extract, 2 - 4 parts of vegetable blend oil.

5. The method for culturing fry of the golden tiger hybrid grouper according to claim 1, characterized in that, In step S2, the nutritional preparation, according to weight parts, includes the following raw materials: 3 - 5 parts of earthworm fermentation broth, 3 - 5 parts of chitin fermentation broth. The preparation method of the nutritional preparation is: Treat the earthworm fermentation broth and the chitin fermentation broth respectively through a dynamic high-pressure microfluidic homogenizer at 80 - 120 MPa for 2 times, and mix them to obtain.

6. The method for cultivating fry of the golden tiger hybrid grouper as described in claim 1, characterized in that, In step S3, the compound feed, according to weight parts, includes the following raw materials: 10 - 20 parts of shrimp meal, 5 - 8 parts of clam meat slurry, 2 - 4 parts of carnosic acid nutrient solution, 3 - 4 parts of wild ganoderma extract, 1 - 3 parts of perilla leaf extract, 1 - 3 parts of morinda citrifolia fruit extract, 1 - 3 parts of longan polysaccharide.

7. The method for cultivating fry of the golden tiger hybrid grouper according to claim 1, characterized in that, In step S4, the specific low-frequency ultrasonic field treatment is: Ultrasonic at 20 - 30 kHz and 10 - 15 W for 20 - 30 s, after an interval of 30 s, continue to ultrasonic for 20 - 30 s, and repeat the above process until fertilization is completed.

8. The method for cultivating the fry of the golden tiger hybrid grouper according to claim 1, characterized in that, In step S5, the intermittent ultrasonic treatment is to first ultrasonic at 20 - 30 kHz and 20 - 30 W for 30 - 60 s, with an interval of 3 min, then ultrasonic at 50 - 60 kHz and 50 - 60 W for 20 - 40 s, with an interval of 5 min, and repeat the above process 3 - 5 times.

9. The method for cultivating the fry of the golden tiger hybrid grouper as described in claim 1, characterized in that, In step S6, the preparation method of the probiotic magnetic chitosan microspheres includes: adding chitosan into water, transferring it to a high-pressure reactor, carrying out hydrothermal reaction at 160 - 200 °C for 12 - 24 h, cooling, washing, and drying. Dispersing the product in deionized water, adding ferric chloride and PVP and stirring, adding concentrated ammonia water under the condition of 70 - 80 °C, continuing to stir for 1 - 2 h, cooling, centrifuging to separate the microspheres, washing, and drying to obtain magnetic chitosan microspheres, and encapsulating the probiotics in the magnetic chitosan microspheres by the swelling-adsorption method; The solid-liquid ratio of the chitosan to water is 1:10 - 20 g / mL, and the solid-liquid ratio of the product to deionized water is 1:10 - 20 g / mL; the mass ratio of the ferric chloride, PVP to the product is 1 - 2:0.5 - 0.7:5; the addition amount of the concentrated ammonia water is 0.2 - 0.4 times the mass of the product, and the concentration is 20 - 30 wt%; the loading amount of the probiotics in the magnetic chitosan microspheres is 20% - 30%.

10. The method for culturing the fry of the hybrid grouper as claimed in claim 1, characterized in that, In step S6, during the preliminary cultivation, control the salinity of the aquaculture pond to be 15 - 25‰, the temperature to be 25 - 28 °C, the pH value to be 7.5 - 8.0, and the dissolved oxygen to be greater than 6.0 mg / L.

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