Artificial breeding and selection method of small plateau native ornamental fish

Abstract

This invention discloses a method for the artificial breeding and selection of small plateau native ornamental fish, belonging to the field of aquaculture technology. The method includes: S1 broodstock domestication, S2 hormone-free spawning induction, S3 fertilized egg hatching, S4 graded initial domestication, and S5 juvenile fish selection. In S2, the broodstock are placed in a spawning induction device, where visual mirror stimulation is generated through a reflecting mirror. Combined with the synergistic effect of gradually increasing light intensity from 8 hours to 12 hours and water temperature from 16°C to 22°C, and the weak electric field generated by the bottom microelectrode array (intensity below the fish's perception threshold, waveform is sine or square wave), natural spawning is induced without the addition of exogenous hormones, achieving a spawning rate of over 87.5%. In S4, the fry are fed graded micro-particle feed in three stages, with particle sizes of 80-120μm, 120-160μm, and 160-200μm, respectively. The fry feeding rate reaches over 87%, and the survival rate at 35 days of age reaches over 78%. This invention enables the artificial breeding of native ornamental fish in the plateau with zero hormones, and is simple to operate with a high survival rate.

Description

Technical Field

[0001] This invention relates to the field of aquaculture technology, specifically to a method for the artificial breeding and selection of small plateau native ornamental fish. Background Technology

[0002] Native ornamental fish of the plateau (such as those in the genus *Danioninae*) possess extremely high ornamental and ecological conservation value due to their vibrant colors and unique habits, making them a distinctive germplasm resource of freshwater fish in Yunnan Province's plateau region. However, their artificial breeding has long faced the following technical bottlenecks: 1. Low spawning rate and dependence on hormones: Current technologies mostly use exogenous hormones (such as HCG, LHRH-A2, dioxin, etc.) to induce spawning. This method has problems such as strong stress response in parent fish, high postpartum mortality, high offspring deformity rate, and hormone residues. Moreover, the spawning rate is generally less than 55%.

[0003] 2. Difficulty in feeding fry: Small ornamental fish fry in high-altitude areas have extremely small mouths (usually <200μm). Existing commercial feeds have excessively large particle sizes or unsuitable nutrient ratios, resulting in low feeding rates, slow growth, and a survival rate of less than 40%.

[0004] 3. Lack of systematic breeding methods: At present, research on the whole-cycle artificial breeding methods for small native ornamental fish in plateau areas is weak, and a standardized and replicable technical system has not yet been formed.

[0005] Therefore, developing a hormone-free, high-survival-rate breeding and selection method suitable for industrial promotion is of great practical significance. Summary of the Invention

[0006] To address the shortcomings of existing technologies, this invention provides a method for the artificial breeding and selection of small plateau native ornamental fish to solve the aforementioned problems.

[0007] To achieve the above objectives, the present invention provides a method for the artificial breeding and selection of small plateau native ornamental fish, characterized by the following steps: S1. Broodstock domestication: Broodstock are domesticated for 14–21 days in a simulated plateau ecological environment. S2, Zero-hormone spawning induction: The domesticated parent fish are placed in a spawning induction device, and through the synergistic effect of visual mirror stimulation, dynamic photoperiod regulation and temperature regulation, the parent fish are induced to spawn naturally without the addition of exogenous hormones. S3. Fertilized egg incubation: Collect fertilized eggs and incubate them under micro-flow conditions; S4. Graded initial rearing: After hatching, the seedlings are fed micro-particle feed with a particle size of 80–200 μm in stages according to their age, with the particle size increasing gradually in each stage. It includes at least three stages, with particle size ranges of 80–120 μm, 120–160 μm and 160–200 μm for each stage. S5. Juvenile fish selection: Select individuals with fast growth and good body color to continue raising to marketable size.

[0008] Preferably, the spawning device in S2 includes at least 5 reflective mirrors disposed on the inner wall of the cylinder. The reflective mirrors are respectively disposed on the four sides and bottom of the cylinder and are installed by an angle adjustment structure. The mirror angle can be independently adjusted within the range of 0°–60°. The mirror angle is adjusted electrically or automatically by the control unit based on feedback from the fish behavior.

[0009] Preferably, the dynamic photoperiod control in S2 includes gradually increasing the illumination duration from 8 hours per day to 12 hours, with an increase of no more than 0.5 hours per day; the temperature control includes raising the water temperature from 16°C to 22°C at a rate of ≤2°C / day.

[0010] Preferably, the labor induction device in S2 is further provided with an LED light source system, which is divided into at least three independently controlled lighting zones. Each lighting zone can be set with different light intensities, including a strong light zone, a medium light zone, and a weak light zone. Each lighting zone is provided with at least three types of LED beads with different peak wavelengths, including cool white LEDs, blue LEDs, and green LEDs, and the intensity of each spectral channel can be adjusted independently.

[0011] Preferably, the induced labor device in S2 is further provided with a microelectrode array laid at the bottom of the cylinder. The microelectrode array is used to generate a weak electric field with an intensity lower than the fish's perception threshold and a waveform of sine wave or square wave, in order to simulate the weak fluctuations of the bioelectric field on the surface of similar fish.

[0012] Preferably, the synergistic effect of visual mirror stimulation, dynamic photoperiodic modulation, and temperature modulation described in S2 lasts for 3–5 days.

[0013] Preferably, the specific stages of graded start-up domestication described in S4 are as follows: feeding micro-particle feed with a particle size of 80–120 μm at 3–8 days old; feeding micro-particle feed with a particle size of 120–160 μm at 9–20 days old; and feeding micro-particle feed with a particle size of 160–200 μm at 21–35 days old.

[0014] Preferably, the micro-particle feed in S4 comprises, by weight percentage: 25–35% fish meal, 15–25% Artemia powder, 10–15% Spirulina, 8–12% yeast extract, 10–15% α-starch, 3–6% fish oil, and 1–3% betaine.

[0015] Preferably, the small plateau native ornamental fish is a species of the genus *Hemiberlesia*.

[0016] Preferably, no exogenous hormones are added in step S2 of the method.

[0017] This invention provides a method for the artificial breeding and selective breeding of small plateau native ornamental fish. It has the following beneficial effects: 1. This invention employs a synergistic effect of three signals: visual mirror stimulation, dynamic photoperiod, and temperature regulation, completely eliminating the need for exogenous hormone injection. It achieves comprehensive visual stimulation through at least five adjustable reflective mirrors, coupled with a gradual increase in light intensity from 8 hours to 12 hours and a water temperature increase from 16°C to 22°C, resulting in a natural spawning rate of over 87.5% for broodstock and a post-spawning survival rate of ≥98%. Compared to traditional hormone injection methods (spawning rate ≤55%), this invention avoids hormone residues and stress damage.

[0018] 2. This invention addresses the extremely small mouth size of small fish fry in high-altitude areas by employing a three-stage graded feeding method. Experiments have verified that the 120–160 μm transition period is indispensable; skipping it reduces the survival rate from 81.2% to 58.7%. The feed contains 15–25% artemia powder to compensate for the physical deficiencies with chemical attractants. The fry's feeding rate reaches over 87%, and the survival rate at 35 days old exceeds 78%, an improvement of approximately 38 percentage points compared to the traditional method's less than 40%.

[0019] 3. This invention integrates a reflective mirror, multispectral zoned LEDs, and a microelectrode array into a single spawning induction device, achieving sequential synergy of visual, light environment, and electrical stimulation modes. The spawning rate in the single visual mode is 76.8%, which increases to over 87% after adding a zoned light environment, and further increases to over 89% after adding microelectrode stimulation, achieving a synergistic effect of "1+1+1>3". The control unit can automatically optimize parameters based on feedback from fish behavior, forming a standardized breeding system throughout the entire lifecycle. Attached Figure Description

[0020] Figure 1 This is a process flow diagram of the artificial breeding and selection method for small plateau native ornamental fish of the present invention; Figure 2 This is a schematic diagram of the overall structure of the zero-hormone labor-inducing device in this invention; Figure 3 This is a schematic diagram of the feeding stage in this invention.

[0021] 1. Aquaculture tank; 2. Reflective mirror (side walls and bottom); 3. LED light source system; 4. Temperature control system (heating rod); 5. Microelectrode array; 6. Control unit. Detailed Implementation

[0022] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0023] The experimental fish species was *Hemiberlesia lataniae*, collected from an upstream tributary of Dianchi Lake in Kunming, Yunnan Province. It was confirmed by quarantine to be free of specific pathogens, with a body length of 3.5–5.0 cm and a weight of 1.2–3.5 g.

[0024] Experimental location: A plateau fish breeding base in Anning City, Yunnan Province (altitude 1820m), indoor controlled environment laboratory.

[0025] Example 1: S1: Parent Fish Training 120 parent fish (50 females and 70 males) were placed in 6 glass aquariums, 20 fish per tank. Rearing conditions: water temperature 18±0.5℃, photoperiod 10L:14D, light period 08:00–18:00, light intensity 500 lux. Live brine shrimp were fed daily at 09:00 and 16:00 in combination with the micro-particle feed of this invention (particle size 160–200 μm), at a rate of 3% of the parent fish's total weight. One-third of the water was changed daily for 21 days. During the rearing period, 2 parent fish died, resulting in a survival rate of 98.3%.

[0026] S2: Zero-hormone labor induction 1. Assembly of the catalytic converter The spawning device consists of a breeding tank 1, a reflective mirror system 2, an LED light source system 3, a temperature control system 4, a microelectrode array 5, and a control unit 6.

[0027] The aquaculture tank 1 is made of 40×30×30cm transparent acrylic material with an effective volume of 36L.

[0028] The reflective mirror system 2 has one aluminum-plated acrylic reflector installed on the inner sides of the front, rear, left, and right walls of the cylinder, with a reflectivity of 88%. A mirror of the same material is installed at the bottom, meaning that reflective mirrors 2 are provided on all four side walls and the bottom. A hinge is installed on the back of each mirror, with the movable end of the hinge connected to the mirror. A stepper motor is externally connected to the fixed end of each hinge, and the stepper motor driver receives pulse signals from the control unit 6. The control unit 6 presets the four side wall mirrors to 30° and the bottom mirror to 15°, automatically sending out the corresponding number of pulses to drive the motor and rotate the mirrors to the target angle.

[0029] The LED light source system integrates five independent lighting zones, each containing three types of LEDs: 6500K cool white, 460nm blue, and 530nm green, in a 4:2:2 ratio. PWM dimming is employed, with each zone and channel controlled independently.

[0030] The temperature control system 4 is a 200W titanium alloy heating rod with a temperature control accuracy of ±0.3℃, and the PT100 temperature probe has an accuracy of ±0.1℃.

[0031] The microelectrode array 5 consists of 37×27cm electrode plates laid above the bottom mirror surface of the tank. In this embodiment, the microelectrodes used are a metal mesh array, made of platinum-iridium alloy, arranged in an 8×8 grid pattern, with a spacing of 3.5cm between adjacent electrode points and a diameter of 2mm for each electrode point. The metal coverage of this metal mesh array is only 2.5%, with an equivalent light transmittance of over 97.5%. More than 97% of the light emitted by the top LED light source passes directly through the mesh gaps to reach the bottom reflective mirror surface. Therefore, the presence of the microelectrode array 5 will not have a substantial impact on the in-tank lighting environment or visual mirror stimulation. Simultaneously, the electric field strength is controlled at 0.5V / cm, far below the stress response threshold of 2-3V / cm for fish, generating a sine wave that simulates the weak fluctuations of the bioelectric field on the surface of similar fish, without causing discomfort to the fish. The signal generator is an AD9833DDS module, controlled by an STM32 microcontroller to output a sine wave, which is amplified by a power amplifier to a field strength of 0.5V / cm (measured). In this embodiment, the frequency is 20Hz, the waveform is a sine wave, and it is continuously on.

[0032] 2. Labor induction procedures After domestication, the parent fish were transferred to the spawning induction device in a female-to-male ratio of 1:1.2, divided into 4 tanks with 10 females and 12 males per tank. Initial conditions: water temperature 16℃, photoperiod 8L:16D. The temperature increase program was initiated at a rate of 1.5℃ / day, i.e., 16℃ to 17.5℃ on day 1… to 12L on day 8, and then maintained at 12L. Mirror stimulation and microelectrode stimulation were activated throughout the process.

[0033] 3. Results of labor induction Observations were conducted daily at 08:00, 12:00, 16:00, and 20:00 starting from day 3. Concentrated spawning occurred on day 6 from 09:00 to 11:00. Approximately 5300 eggs were collected from the four tanks, representing a spawning rate of 89.2%. 36 females spawned out of a total of 40 females, resulting in a fertilization rate of 84.3%. One hundred eggs were randomly selected for microscopic observation of blastodisc formation. The parent fish survival rate was 98.3%, with 3 fish dying.

[0034] S3: Hatching of fertilized eggs The fertilized eggs were transferred to cylindrical hatching net cages, 500–600 eggs per cage. The hatching water temperature was 21±0.5℃, with a micro-flow rate of 0.5L / min and dissolved oxygen ≥6.5mg / L. Dead eggs were removed twice daily. Hatching occurred after 52–54 hours, with approximately 4470 larvae hatching, a hatching rate of 91.0%. Newly hatched larvae were 2.8–3.2mm in total length.

[0035] S4: Graded Open-Air Training 1. Preparation method of micro-particle feed Weigh the following ingredients by weight percentage: fish meal 30%, Artemia powder 20%, Spirulina 12%, yeast extract 10%, α-starch 12%, fish oil 4%, betaine 2%. Preparation steps: (1) Mix fish meal, Artemia powder, Spirulina, yeast extract and α-starch evenly and pass through an 80-mesh sieve.

[0036] (2) Dissolve fish oil and betaine in a small amount of water (about 20% of the total feed mass) and add to the mixed powder, stirring until there is no dry powder.

[0037] (3) Extrusion molding was performed using a twin-screw extruder at 110℃ and 1.5MPa pressure. Key process parameters: screw speed 180rpm, die orifice diameter 0.5mm. Under these conditions, the material remained in the barrel for about 30 seconds. The high temperature caused the starch to gelatinize and the protein to denature, forming a micro-particle structure with a certain strength that did not disintegrate within 12 hours after being immersed in water.

[0038] (4) The extrudate is cut into short particles by a cutting blade and passed through 80 mesh, 120 mesh, 160 mesh and 200 mesh standard sieves in sequence to collect three components of 80–120μm, 120–160μm and 160–200μm respectively. The sieved material is dried in an oven at 40℃ until the moisture content is ≤10% and then sealed and packaged.

[0039] 2. First-day domestication procedures Stage I (3–8 days old): Feed 80–120μm feed 6 times a day at 07:00, 10:00, 13:00, 16:00, 19:00, and 22:00, approximately 0.2g per 10,000 fish each time. Keep in 30L white tanks with a water depth of 20cm, 500 fish per tank, water temperature 22℃, still water, and change 1 / 3 of the water daily.

[0040] Stage II (9–20 days old): Feed 120–160 μm feed 5 times a day, about 0.5 g per 10,000 fish each time. Other conditions are the same as in Stage I.

[0041] Stage III (21–35 days old): Feed 160–200 μm feed 4 times a day, about 1.0g per 10,000 fish each time. Switch to 60×30×30cm glass tanks with a water depth of 15cm, 300 fish per tank.

[0042] 3. Measurement of feeding rate and survival rate Thirty fish were randomly selected 30 minutes after feeding at days 8, 20, and 35, and their intestinal fullness was observed under a stereomicroscope. The feeding rate was 84.0% at day 3, 91.3% at day 8, and 88.6% at day 35, with an average of 87.3%. Approximately 3,620 fish survived at day 35, representing a survival rate of 81.2%.

[0043] S5: Juvenile Fish Breeding On day 35, individuals with a body length ≥2.5cm, bright coloration, no deformities, and active swimming were selected, accounting for approximately 30% of the total. After selection, they continued to be raised until 60 days of age, fed 160–200μm feed and a small amount of frozen bloodworms four times a day at a water temperature of 20℃. At 60 days of age, the average body length was 3.5cm, and they were ready for sale as commercial fish.

[0044] Comparative Example 1 The only difference from Example 1 is in S4: the graded starter culture is changed to a two-stage process—feeding 80–120 μm at 3–8 days of age, and directly feeding 160–200 μm at 9–35 days of age (skipping the 120–160 μm transition period). All other conditions are exactly the same.

[0045] Results: During the 9–20 day period, the seedlings exhibited significant feeding difficulties (particle size jumped from 120 μm to 200 μm, exceeding the mouth diameter tolerance range), with a feeding rate of only 41.2% during this stage. The survival rate at day 35 was 58.7%, a decrease of 22.5 percentage points compared to Example 1. This demonstrates that the 120–160 μm transition period is indispensable.

[0046] Comparative Example 2 The difference from Example 1 lies in the following control groups set up in S2: Control group A, single vision mode: LED zoned lighting environment uniformly 1200 lux throughout the tank with LEDs off, microelectrode array off, only the reflective mirror is retained. Egg laying rate 76.8%.

[0047] Control group B: Visual illumination environment: Reflective mirrors and LED zoned lighting were enabled, while the microelectrode array was disabled. Egg laying rate: 87.3%.

[0048] Control group C, full mode, i.e., Example 1: with the reflective mirror, LED zoned lighting environment, and microelectrode array turned on. Egg laying rate: 89.2%.

[0049] Results analysis: The egg-laying rate of the single visual mode was 76.8%. After adding the partitioned light environment, it increased by 10.5 percentage points to 87.3%. After adding microelectrode stimulation, it further increased by 1.9 percentage points to 89.2%, achieving a superimposed effect of "1+1+1>3".

[0050] Example 2 Using the golden-line barb as the subject, the parameters were adjusted as follows: acclimatization water temperature 20℃; reflector angle: sidewall 35°, bottom 20°; LED illuminance: left / right zone 2200 lux, center left / center right zone 1000 lux, center zone 400 lux; microelectrode field strength 0.3V / cm, frequency 15Hz; photoperiod: initial 8L:16D, increasing by 0.4 hours daily to 11.2L:12.8D; temperature increase of 1.2℃ / day from 18℃ to 23℃. The initial feed particle size was appropriately relaxed: stage I 100–150μm, stage II 150–200μm, stage III 200–250μm (the above parameters are adaptive adjustments for the golden-line barb and do not deviate from the protection scope of claim 1 of this invention).

[0051] Results: The egg-laying rate was 82.6%, the fertilization rate was 79.4%, the seedling feeding rate was 84.2%, and the survival rate was 75.6%. This indicates that the invention is also effective for the golden-line barb and has a certain degree of cross-species versatility.

[0052] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A method for artificial breeding and selective breeding of small plateau native ornamental fish, characterized by: Includes the following steps: S1. Broodstock domestication: Broodstock are domesticated for 14–21 days in a simulated plateau ecological environment. S2, Zero-hormone spawning induction: The domesticated parent fish are placed in a spawning induction device, and through the synergistic effect of visual mirror stimulation, dynamic photoperiod regulation and temperature regulation, the parent fish are induced to spawn naturally without the addition of exogenous hormones. S3. Fertilized egg incubation: Collect fertilized eggs and incubate them under micro-flow conditions; S4. Graded initial rearing: After hatching, the seedlings are fed micro-particle feed with a particle size of 80–200 μm in stages according to their age, with the particle size increasing gradually in each stage. It includes at least three stages, with particle size ranges of 80–120 μm, 120–160 μm and 160–200 μm for each stage. S5. Juvenile fish selection: Select individuals with fast growth and good body color to continue raising to marketable size.

2. The method for artificial breeding and selection of small plateau native ornamental fish according to claim 1, characterized in that, The spawning device described in S2 includes at least five reflective mirrors disposed on the inner wall of the cylinder. The reflective mirrors are respectively disposed on the four sides and bottom of the cylinder and are installed by an angle adjustment structure. The mirror angle can be independently adjusted within the range of 0°–60°. The mirror angle is adjusted electrically or automatically by the control unit based on feedback from the fish behavior.

3. The method for artificial breeding and selection of small plateau native ornamental fish according to claim 1, characterized in that, The dynamic photoperiod control described in S2 includes gradually increasing the illumination duration from 8 hours to 12 hours per day, with an increase of no more than 0.5 hours per day; the temperature control includes raising the water temperature from 16℃ to 22℃ at a rate of ≤2℃ / day.

4. The method for artificial breeding and selection of small plateau native ornamental fish according to claim 1, characterized in that, The labor induction device described in S2 is also equipped with an LED light source system, which is divided into at least three independently controlled lighting zones. Each lighting zone can be set with different light intensities, including a strong light zone, a medium light zone, and a weak light zone. Each lighting zone is equipped with at least three types of LED beads with different peak wavelengths, including cool white LEDs, blue LEDs, and green LEDs. The intensity of each spectral channel can be adjusted independently.

5. The method for artificial breeding and selection of small plateau native ornamental fish according to claim 1, characterized in that, The induced labor device described in S2 is also equipped with a microelectrode array laid at the bottom of the cylinder. The microelectrode array is used to generate a weak electric field with an intensity lower than the fish's perception threshold and a waveform of sine wave or square wave, in order to simulate the weak fluctuations of the bioelectric field on the surface of similar fish.

6. The method for artificial breeding and selection of small plateau native ornamental fish according to claim 1, characterized in that, The synergistic effect of visual mirror stimulation, dynamic photoperiodic modulation, and temperature modulation described in S2 lasts for 3–5 days.

7. The method for artificial breeding and selection of small plateau native ornamental fish according to claim 1, characterized in that, The specific stages of graded start-up domestication described in S4 are as follows: feed micro-particle feed with a particle size of 80–120 μm at 3–8 days old; feed micro-particle feed with a particle size of 120–160 μm at 9–20 days old; and feed micro-particle feed with a particle size of 160–200 μm at 21–35 days old.

8. The method for artificial breeding and selection of small plateau native ornamental fish according to claim 7, characterized in that, The micro-particle feed described in S4 contains, by weight percentage: 25–35% fish meal, 15–25% Artemia powder, 10–15% Spirulina, 8–12% yeast extract, 10–15% α-starch, 3–6% fish oil, and 1–3% betaine.

9. The method for artificial breeding and selection of small plateau native ornamental fish according to claim 1, characterized in that, The small, native ornamental fish of the plateau is a species of the genus *Ctenopharynx*.

10. The method for artificial breeding and selection of small plateau native ornamental fish according to claim 1, characterized in that, The method described does not add any exogenous hormones in S2.

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