A method for co-culturing giant freshwater prawns with shrimp, grass, and fish based on probiotic regulation and its application

Abstract

This invention discloses a method for co-culturing giant freshwater prawns (Macrobrachium rosenbergii) with shrimp, grass, and fish based on probiotic regulation, and its application. This invention constructs a three-dimensional ecological system with shrimp as the main body, grass as the protector, and fish as a supplement. Elodea and water spinach purify the water, silver carp and crucian carp decompose uneaten feed and feces, and a compound microbial preparation of Lactococcus gasseri C6a2 regulates water quality, while a biological bottom conditioner improves the bottom sediment, achieving precise ecological regulation of the aquaculture environment. This invention does not rely on chemical agents, effectively inhibits ammonia nitrogen accumulation, reduces disease incidence, and significantly improves the survival rate and size of giant freshwater prawns, achieving a win-win situation for both ecological and economic benefits.

Description

Technical Field

[0001] This invention relates to the field of aquaculture, specifically to a method for the co-culture of giant freshwater prawns (Macrobrachium rosenbergii) based on probiotic regulation of shrimp-grass-fish and its application. Background Technology

[0002] As an important freshwater aquaculture species in my country, the giant freshwater prawn (Macrobrachium rosenbergii) has become one of the most economically valuable pillar categories in the aquaculture industry due to its delicious meat, rapid growth rate, and strong market demand. With the continuous increase in market demand for high-quality giant freshwater prawn products and the expansion of aquaculture scale, traditional giant freshwater prawn farming methods, lacking scientific ecosystem construction, especially the synergistic regulation of aquatic plants, fish, and probiotics, have gradually revealed many problems that urgently need to be addressed in long-term practice.

[0003] Currently, most mainstream giant freshwater prawn (Macrobrachium rosenbergii) farming methods employ a single, high-density intensive farming approach. This method is highly dependent on chemical agents and lacks an ecological regulation system that includes aquatic plants, filter-feeding fish, and probiotics. During the farming process, disinfectants, pesticides, and water quality improvers are frequently used to control diseases and improve water quality. This approach not only easily leads to drug residues in the giant freshwater prawns, affecting product quality and food safety, but also disrupts the aquatic microecological balance, exacerbating aquatic environmental degradation. Specifically, the traditional model has extremely low aquatic vegetation coverage, making it impossible for aquatic plants to absorb harmful substances such as ammonia nitrogen and nitrite in the water through purification, and also unable to provide shelter for giant freshwater prawns to reduce cannibalism. Without the addition of filter-feeding fish, it is difficult to decompose organic waste such as uneaten feed and prawn feces generated during the farming process, resulting in a large accumulation of such waste at the bottom of the pond, causing bottom acidification, blackening, and foul odor. At the same time, the lack of scientific regulation by probiotics makes it impossible to effectively degrade organic pollutants and inhibit the growth of pathogenic microorganisms, ultimately leading to frequent diseases in giant freshwater prawns, such as black gill disease and ciliate disease, which significantly reduces the survival rate of farmed prawns.

[0004] Furthermore, traditional aquaculture methods lack precision in water quality and bottom sediment control, relying heavily on experience-based judgment. The absence of synergistic regulation by aquatic plants, fish, and probiotics makes it difficult to adapt to the environmental needs of giant freshwater prawns at different growth stages, leading to significant fluctuations in key indicators such as dissolved oxygen, pH, ammonia nitrogen, and nitrite. This not only further impacts the growth and development of giant freshwater prawns, resulting in inconsistent adult sizes and unstable product quality, but also diminishes the water's self-purification capacity, necessitating continuous chemical interventions to maintain the aquaculture environment and further increasing costs. Simultaneously, single-species aquaculture models fail to fully leverage the synergistic effects of aquatic plants, fish, and giant freshwater prawns in a three-dimensional environment, resulting in low resource utilization and significant economic fluctuations due to factors such as disease and environmental changes, indicating insufficient stability and reliability for large-scale aquaculture.

[0005] As the aquaculture industry transforms towards green, ecological, and high-quality practices, the traditional giant freshwater prawn (Macrobrachium rosenbergii) farming model, lacking the synergistic ecological effects of aquatic plants, fish, and probiotics, has become a core bottleneck restricting industry upgrading. The industry urgently needs to construct an ecological system that relies on aquatic plants to purify water, fish to decompose waste, and probiotics to regulate the microecology, replacing the traditional chemical-dependent farming model. This system aims to achieve the goals of eliminating reliance on chemical agents, maintaining ecological balance, effectively controlling diseases, improving the aquatic environment, and reducing residual feed pollution, while simultaneously increasing survival rates and economic benefits. Therefore, how to overcome the limitations of existing models and develop a giant freshwater prawn farming technology that integrates the synergistic effects of aquatic plants, fish, and probiotics has become a pressing technical challenge in the aquaculture field, and is of great significance for promoting the large-scale and high-quality development of the giant freshwater prawn industry. Based on this, the development of an ecologically regulated, high-efficiency farming method, through a three-dimensional ecological system of "prawn as the main body, aquatic plants as protection, and fish as supplementary support" combined with probiotic regulation, aims to achieve a win-win situation for both ecological and economic benefits, thereby promoting the transformation and upgrading of the giant freshwater prawn industry towards a green and healthy direction. Summary of the Invention

[0006] The purpose of this invention is to overcome the shortcomings of existing technologies and provide a method for the co-culture of giant freshwater prawns (Macrobrachium rosenbergii) based on probiotic regulation, and its application. This invention constructs a three-dimensional ecological system with "prawns as the main body, aquatic plants as the protectors, and fish as supplementary support," combined with precise control of water and bottom quality and refined feeding management. This achieves reduced disease incidence, efficient resource utilization, increased yield and quality of giant freshwater prawns, reduced farming costs, and ultimately a win-win situation for both ecology and economy, providing reliable technical support for large-scale, high-quality giant freshwater prawn farming.

[0007] To achieve the above objectives, the technical solution designed by the present invention is as follows: This invention provides a method for the co-culture of giant freshwater prawns (Macrobrachium rosenbergii) based on probiotic regulation, comprising the following steps: (1) Drain the remaining water in the pond, dry the pond and expose it to the sun and turn over the bottom mud; after the sun exposure, fill the pond with water and disinfect the whole pond with bleaching powder. After disinfection, let it stand and then carry out the first stage of aquatic plant planting. (2) Release giant freshwater prawn larvae, and simultaneously raise fish larvae in the same manner after release. Then, carry out the second stage of aquatic plant planting 7-10 days after release to construct a three-dimensional ecological system of shrimp-grass-fish. (3) Maintain dissolved oxygen in the water body ≥5 mg / L through oxygenation equipment management; spray compound microbial agents into the pond every 14-16 days after stocking to regulate water quality; apply biological bottom conditioner and sodium humate every 6-8 days after stocking to improve bottom sediment and detoxify water; and test the pH value, ammonia nitrogen and nitrite index of the water body. (4) After the seedlings are released, they are fed daily. After the feeding is completed, they are caught using a ground cage net. The mixed-culture fish are caught simultaneously with the later harvest of the giant freshwater prawn.

[0008] Furthermore, in step (1), the drying and sun exposure time of the pond is 15 to 25 days; the amount of bleaching powder used is 45 to 55 catties / mu; after disinfection, the pond is left to stand until the residual chlorine content drops below 0.05 mg / L before proceeding to the next step. The first stage of aquatic plant planting involves planting Elodea in the pond 3-5 days before releasing Giant freshwater prawn larvae using a hole planting method at intervals of 2-3 meters, with one plant per hole and a planting depth of 3-5 cm.

[0009] Furthermore, in step (2), the stocking density of giant freshwater prawn larvae is 40,000 to 50,000 larvae per mu. Before stocking, the transport bags of giant freshwater prawn larvae are soaked in the corresponding pond water for 20 to 30 minutes to adapt to the temperature. When the temperature difference between the water inside the bag and the pond water drops to below 2°C, the bag is opened to allow the giant freshwater prawn larvae to swim into the pond on their own. The mixed-culture fish fry are silver carp fry and crucian carp fry. The stocking density of silver carp fry is 5-7 fish / mu, and the stocking density of crucian carp fry is 6-8 fish / mu. The mixed-culture fish fry are evenly distributed in each pond area after adopting the same temperature adaptation method as the giant freshwater prawn fry.

[0010] Furthermore, in step (2), the second stage of aquatic plant planting specifically involves planting water spinach by cuttings at a depth of 10-20 cm along the banks of each pond, with a plant spacing of 15-20 cm.

[0011] Furthermore, in step (3), the oxygenation equipment includes a waterwheel aerator and an impeller aerator, with power of 10~15 kW and 30~40 kW per mu respectively.

[0012] Furthermore, in step (3), the amount of compound microbial preparation applied is 2-4 jin / mu. After fermentation, the compound microbial preparation is evenly applied to the entire pond. After application, the oxygenation equipment is turned on for 2-3 hours.

[0013] Furthermore, the compound microbial preparation contains EM bacteria and Lactococcus gasseri C6a2, and the content of EM bacteria in the compound microbial preparation is ≥5×10⁻⁶. 11 CFU / catties, viable count of Lactococcus gasseri C6a2 ≥1×10⁻⁶ 11 CFU / catties; The fermentation treatment conditions were: sealed and static culture at 30°C for 3-5 days. The preservation number of the Lactococcus gasseri C6a2 is CGMCC NO: M 31024.

[0014] Furthermore, in step (3), the application rate of the biological bottom conditioner is 2-4 catties / mu, and the application rate of sodium humate is 0.8-1.2 catties / mu; After bottom sediment improvement and water detoxification, the water quality indicators are maintained at: pH 7.0~8.5, ammonia nitrogen content ≤0.5 mg / L, and nitrite content ≤0.1 mg / L.

[0015] Furthermore, in step (4), the daily feeding is specifically as follows: When the weight of giant freshwater prawns is <1 g, the daily feeding rate is 15-20% of their body weight; when the weight of giant freshwater prawns is 1-5 g, the daily feeding rate is 8-10% of their body weight; and when the weight of giant freshwater prawns is >5 g, the daily feeding rate is 3-6% of their body weight.

[0016] This invention also provides an application of the aforementioned giant freshwater prawn farming method in large-scale ecological farming of giant freshwater prawns.

[0017] The beneficial effects of this invention are: 1. Constructing a virtuous ecological cycle and improving the aquaculture environment: This invention uses a three-dimensional ecological system of "shrimp + grass + fish". Elodea and water spinach purify the water quality, and filter-feeding fish decompose uneaten feed and feces, forming an ecological closed loop of shrimp removing grass and fish purifying water. This effectively solves the problems of easy deterioration of the water environment and serious pollution from uneaten feed in the existing aquaculture model, and reduces the risk of disease occurrence.

[0018] 2. Improve aquaculture efficiency: The method of this invention can significantly improve the survival rate and size of giant freshwater prawns, increase additional output by co-culturing with fish species, reduce waste of chemical agents and feed, lower aquaculture costs, achieve a double improvement in yield and quality, and enhance the economic benefits of large-scale aquaculture.

[0019] 3. Achieving a win-win situation for ecology and economy: This invention reduces the negative impact of aquaculture activities on the environment through biological regulation and ecological balance maintenance, taking into account both ecological protection and aquaculture benefits, providing reliable technical support for large-scale high-quality aquaculture of giant freshwater prawns, and promoting the green and sustainable development of the industry. Attached Figure Description

[0020] Figure 1 This is a spatial layout diagram of a shrimp-grass-fish co-culture pond based on probiotic regulation for giant freshwater prawns.

[0021] Figure 2 This is a schematic diagram illustrating the ecological balance of the giant freshwater prawn (Macrobrachium rosenbergii) farming model based on probiotic regulation of shrimp-grass-fish co-culture.

[0022] Figure 3 A graph showing the ammonia nitrogen index results in the seedling water; In the figure, R, G, K, and D represent the control group, Lactococcus gasseri group, Bacillus subtilis group, and Clostridium butyricum group, respectively. Detailed Implementation

[0023] The present invention will now be described in further detail with reference to specific embodiments, so that those skilled in the art can understand it.

[0024] Experimental materials: 1. The Lactococcus gasseri of this invention is Lactococcus gasseri C6a2, which is described in Chinese Invention Patent Application No. 202411206834.8. Its accession number is CGMCC NO: M 31024, and it was deposited on June 20, 2024, at the China General Microbiological Culture Collection Center, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, China.

[0025] Example 1: Screening of probiotics Lactococcus genomica C6a2 is a functional strain independently developed by our team. In order to clarify its ammonia nitrogen degradation efficiency in aquaculture water and its promoting effect on the development of giant freshwater prawn larvae, and to eliminate the randomness of a single experiment, this embodiment carried out a probiotic screening experiment, thereby providing a reliable experimental basis for the practical application of probiotics.

[0026] 1. Experimental Materials and Test Site 1.1 Experimental Location This experiment was conducted at the Yangdong Giant Freshwater Prawn Breeding Base in Nanxun District, Huzhou City, Zhejiang Province. The base has standardized seedling facilities and basic conditions for water quality control, which can meet the needs of the entire seedling breeding experiment of Giant Freshwater Prawn larvae.

[0027] 1.2 Laboratory Animals The giant freshwater prawn larvae used in the experiment were all sourced from Shufeng Aquatic Products Co., Ltd. in Gaoyou City, Jiangsu Province. The larvae were uniform in size, vigorous, and free from injury, disease, or deformities, meeting the standards for experimental seedlings.

[0028] 1.3 Experimental Probiotics Three probiotics were selected as experimental agents, namely Lactococcus gasseri C6a2 ( Lactococcus garvieae Bacillus subtilis ( Bacillus subtilis ) and Clostridium butyricum ( Clostridium butyricum Among them, Lactococcus gasseri C6a2 was isolated, screened and purified from the intestines of healthy giant freshwater prawns by our research group, and has been verified to have potential probiotic functions in the early stage; Bacillus subtilis and Clostridium butyricum were purchased from Hubei Fengtian Biotechnology Co., Ltd., and were further isolated and purified in the laboratory to ensure that the purity of the strains was ≥99% and the activity reached the standard before being put into experimental use.

[0029] Experimental reagents and equipment The reagents used for preparing artificial seawater include sodium chloride (NaCl), magnesium sulfate (MgSO4), calcium chloride (CaCl), potassium chloride (KCl), sodium bicarbonate (NaHCO3), potassium bromide (KBr), boric acid (BH3O3), EDTA, and strong chlorine, all of analytical grade. The disinfectant is formaldehyde solution (purity ≥37%). The experimental equipment includes a sand filter, a UV water sterilizer, an aeration device, an Okdan water quality analyzer, a lux meter, a thermometer, a precision electronic balance, and seedling tanks (100L capacity). All equipment has been calibrated and disinfected to ensure experimental accuracy and safety.

[0030] 2. Experimental Design and Seedling Management Methods 2.1 Experimental Group Design The experiment consisted of one control group (R group, no probiotics added) and three probiotic treatment groups: Lactococcus gasseri (G group), Bacillus subtilis (K group), and Clostridium butyricum (D group). Each group had three replicates, for a total of 12 experimental units (seedling containers). The final concentration of all probiotic treatment groups was uniformly adjusted to 1×10⁻⁶. 5 The concentration of cfu / ml was evenly sprinkled into the water of the corresponding seedling tank every 5 days. The control group was only sprinkled with the same amount of sterile water, and the other treatments were the same.

[0031] 2.2 Preparation of Seedling Water and Environmental Control 2.2.1 Preparation of artificial seawater: Prepare seedling water according to the following formula, with the following reagent dosages per ton of water: 11,000 g sodium chloride, 3,700 g magnesium sulfate, 420 g calcium chloride, 190 g potassium chloride, 20 g sodium bicarbonate, 20 g potassium bromide, 120 g boric acid, 6 g EDTA, and 1 g strong chlorine.

[0032] 2.2.2 Water pretreatment: The prepared artificial seawater is circulated through a sand filter to remove impurities and particulate pollutants from the water. Then it is disinfected by an ultraviolet light water disinfection device. Finally, it is transferred to a seedling tank for a 3-day aeration and dechlorination to ensure that there is no harmful chlorine residue in the water and to increase the dissolved oxygen content of the water.

[0033] 2.2.3 Environmental Parameter Control: The initial water level for seedling cultivation was set at 60L. On the fourth day of the experiment, the water volume was replenished to 80L, and the salinity was maintained at approximately 14‰ throughout the experiment. During the experiment, environmental parameters were kept stable through a combination of automated control equipment and manual monitoring. The specific parameter ranges were: water temperature 29±1℃, pH value 8.12±0.02, light intensity 1800±300 lux, and dissolved oxygen content 5.65±0.2 mg / L. Various water quality and environmental indicators were regularly tested using a professional equipment such as an Aokdan water quality analyzer, illuminance meter, and thermometer. Data were recorded daily to ensure the consistency and reliability of the experimental conditions.

[0034] 2.3 Release and Feeding Management of Juveniles 2.3.1 Disinfection and release of larvae: Before the experiment, the larvae of giant freshwater prawns were immersed in a 150 ppm formaldehyde solution for 15 minutes to disinfect and kill harmful microorganisms on the surface of the larvae. They were then rinsed with pretreated artificial seawater and released evenly into each experimental seedling tank at a density of 5,000 larvae per tank.

[0035] 2.3.2 Feeding Management: A phased feeding strategy was adopted. For the first 10 days of the experiment, only nauplii of Artemia (Shandong Huitai Biotechnology Co., Ltd.) that had been hatched for 24 hours were fed twice a day, at 7:30 and 16:30 respectively. The amount of feed should be such that there is no uneaten food after the nauplii have finished eating. From the 11th day until the nauplii become seedlings, the feeding plan was adjusted. Egg custard was fed three times a day (at 5:00, 10:00 and 13:00) and Artemia nauplii twice a day (at 7:30 and 16:30) to avoid uneaten food polluting the water.

[0036] 2.3.3 Daily Management: Starting on day 4 of the experiment, sludge removal and water changes were performed. At 3:00 PM daily, uneaten feed and fecal contaminants were removed from the bottom of the seedling tanks using a siphon method. Water was changed every 3 days, with each change replacing 1 / 3 of the total water volume. Artificial seawater pretreated with the same formula was used for water changes to ensure no significant fluctuations in water environmental parameters before and after the water change. The feeding and management methods for all groups were completely identical to avoid interference from additional factors in the experimental results.

[0037] 3. Experimental Results and Analysis 3.1 Effect on the survival rate of giant freshwater prawn larvae After the experiment, the survival rate of giant freshwater prawn larvae in each group was statistically analyzed, and the results are shown in Table 1 below. Table 1 shows that compared with the control group (Group R, survival rate 78.2±2.0%), the survival rates of larvae in all three probiotic treatment groups were improved. Group G (Lactococcus gasseri) had the highest survival rate at 82.4±1.2%, while the survival rates in Group K (Bacillus subtilis) and Group D (Clostridium butyricum) were 80.5±1.5% and 79.7±1.7%, respectively. The results indicate that the addition of probiotics has a positive effect on improving the survival rate of giant freshwater prawn larvae during the seedling stage, and all probiotic treatments did not have adverse effects on larval growth and survival, demonstrating good safety.

[0038] Table 1. Statistics on the survival rate and hatching rate of giant freshwater prawn larvae. 3.2 Effect on emergence rate of giant freshwater prawn larvae Table 1 shows that the hatching rates of the three probiotic treatment groups were significantly higher than those of the control group (P<0.05). The hatching rate of the control group was 43.57±0.86%, while the hatching rates of groups G, K, and D reached 52.80±0.95%, 47.83±0.76%, and 45.20±0.46%, respectively. Among them, the Lactococcus gasseri group (group G) showed the most significant effect on improving the hatching rate, increasing it by 21.18% compared with the control group. The experimental results indicate that adding probiotics can effectively improve the hatching rate of giant freshwater prawn larvae, with Lactococcus gasseri C6a2 showing the best improvement effect.

[0039] 3.3 Effect of regulating ammonia nitrogen content in seedling water Throughout the experiment, the changes in ammonia nitrogen content in the water bodies of each group were monitored, and the results are as follows: Figure 3 As shown, the ammonia nitrogen content in the water of the four groups generally showed an increasing trend with the seedling time, but the increase rate differed significantly among the different treatment groups. The ammonia nitrogen content in the control group (R group) was low in the early stage of the experiment (3-13d), remaining at 0.1mg / L. It began to increase significantly in the middle stage (13-15d), and the increase accelerated after 15d. The ammonia nitrogen content reached 0.6mg / L at 21d, and continued to rise thereafter, reaching as high as 1.0mg / L at the end of the experiment (29d), which was the highest value among all groups.

[0040] Compared with the control group, the increase in ammonia nitrogen in the water of the three probiotic treatment groups (G, K, and D groups) was significantly moderated, demonstrating a good water quality regulation effect. Specifically, the ammonia nitrogen content in groups G and K remained at a low level from the beginning of the experiment, showing a slow linear increase throughout the seedling stage, with the ammonia nitrogen content at the end of the experiment (29 days) being only 0.32-0.37 mg / L. In the later stages of the experiment, the ammonia nitrogen content in group D was slightly higher than that in groups G and K, but still significantly lower than the control group, with the ammonia nitrogen content at the end being 0.52 mg / L. In summary, the addition of probiotics can effectively inhibit the excessive accumulation of ammonia nitrogen in the seedling water, with the Lactococcus gasseri group (G group) and the Bacillus subtilis group (K group) showing more prominent regulatory effects, maintaining a stable water environment during the seedling stage.

[0041] 4. Conclusion This embodiment demonstrates that by adding specific concentrations of *Lactococcus gasseri* C6a2, *Bacillus subtilis*, or *Clostridium butyricum* to the rearing water for giant freshwater prawn larvae, it can improve larval survival rate and significantly increase hatching rate while effectively inhibiting ammonia nitrogen accumulation in the water, thus improving the water quality environment for rearing and providing stable and suitable conditions for giant freshwater prawn larvae growth. Among these, *Lactococcus gasseri* C6a2 showed the best effect in improving hatching rate and also demonstrated outstanding performance in controlling ammonia nitrogen content in the water.

[0042] Lactococcus gasseri C6a2 significantly promotes the development of Macrobrachium rosenbergii and effectively inhibits the accumulation of ammonia nitrogen in the water, improving the water quality environment during seedling and aquaculture. It provides stable and suitable conditions for the growth of Macrobrachium rosenbergii larvae and the rearing of adults. Compared to two other bacterial agents, Lactococcus gasseri C6a2 has a more comprehensive and stable probiotic effect. Furthermore, it was isolated and screened by our research group, making it suitable for the growth needs and aquaculture environment of Macrobrachium rosenbergii. Its safety and applicability meet the requirements of large-scale aquaculture production. Therefore, applying Lactococcus gasseri C6a2 to large-scale Macrobrachium rosenbergii aquaculture production can improve the success rate and economic benefits, and has significant value for widespread application.

[0043] Example 2: A method for co-culturing giant freshwater prawns with grass carp and fish. This embodiment provides a method for co-culturing giant freshwater prawns (Macrobrachium rosenbergii) with grass carp and fish based on probiotic regulation, and conducts large-scale farming practices at a breeding base, combined with... Figure 1 and Figure 2 As shown, the specific steps are as follows: 1. Pond pretreatment stage (early March to late March 2025) 1.1 Pond cleaning operation (1) Before the start of the aquaculture cycle in this embodiment, all 18 ponds were cleaned in a standardized manner. The remaining water in each pond was drained and the ponds were dried and exposed to the sun for 20 days. During this period, the bottom mud was turned over regularly to ensure that the bottom mud was fully dry and cracked, effectively killing pathogenic microorganisms and harmful organism eggs in the bottom mud.

[0044] (2) On March 28, 2025 (one week before stocking), water was poured into each pond to a depth of 50 cm. Bleaching powder was used for full pond disinfection, with the dosage strictly following the standard of 50 catties / mu. After the bleaching powder was fully dissolved, it was evenly sprayed to all areas of each pond using mechanical spraying equipment to ensure disinfection without dead corners. After disinfection, the ponds were left to stand for 4 days, and the residual chlorine content of each pond was tested. The residual chlorine content dropped to a safe range of less than 0.05 mg / L, and the ponds were ready for subsequent aquatic plant planting.

[0045] 1.2 First stage of planting aquatic plants The aquatic plants of this invention adopt a two-stage planting mode. The first stage is carried out on March 30, 2025 (4 days before the seedling release) to plant Elodea. Elodea is planted in each pond at a spacing of 2.5 meters using the hole planting method, with one plant planted in each hole to ensure that the roots of Elodea are fully integrated with the bottom mud. The planting depth is controlled at 4cm to construct a three-dimensional vegetation purification system.

[0046] 1.3 Commissioning of oxygenation equipment Each pond is equipped with a 12kW paddlewheel aerator and a 36kW impeller aerator, evenly distributed according to the pond's shape to ensure aeration coverage of the entire pond. Installation and commissioning of all aeration equipment were completed by March 31, 2025. Trial runs were conducted on the aerators in each pond to check equipment stability and aeration effect. Testing showed that the dissolved oxygen level in each pond could be increased to above 8 mg / L. After commissioning, daily operating procedures were set to ensure stable power supply and continuous equipment operation during the aquaculture period.

[0047] 2. Seedling Release Phase (April 4, 2025) 2.1 Release of Giant Freshwater Prawn Larvae After pond pretreatment was completed and the water environment met standards, the stocking of giant freshwater prawn larvae was carried out on April 4, 2025. The stocking time was chosen on a sunny morning (9:00-11:00 AM) to avoid the period of high temperature and strong sunlight. A total of 2.45 million Guoliang brand high-quality giant freshwater prawn larvae were used for this stocking, averaging 40,800 larvae per acre, precisely matched to the actual area of ​​each pond.

[0048] Before releasing the shrimp larvae, soak the transport bags in the corresponding pond water for 25 minutes to allow them to adapt to the temperature. When the temperature difference between the water inside the bag and the pond water drops below 1.5℃, open the bag and slowly release the shrimp larvae into the shallow water area of ​​the pond, allowing them to crawl into the deep water area on their own. This avoids concentrated release that could lead to excessively high local density.

[0049] 2.2 Stocking of fish fry in mixed culture After the giant freshwater prawn larvae are released, polyculture of fish fry is carried out simultaneously. The fish species used for polyculture are silver carp fry and crucian carp fry. Based on the total culture area of ​​150 mu (approximately 10 hectares) and the actual size of each pond, a stocking density of 6 silver carp fry per mu and 7 crucian carp fry per mu is adopted. The same temperature acclimatization method as for the prawn larvae is used during stocking to ensure the fry have adapted to the aquatic environment before slow release. The stocking locations are evenly distributed across the ponds to avoid excessive concentration with the prawn stocking areas, thus creating a reasonable ecological distribution.

[0050] 2.3 Second stage of aquatic plant cultivation The second phase of aquatic plant planting will take place on April 11, 2025 (7 days after seedling release), with water spinach planted along the pond banks. Water spinach will be planted by cuttings in the shallow water area (15cm deep) along the banks of each pond, with a plant spacing of 18cm to ensure that the roots of the water spinach are in full contact with the water, providing sufficient shelter for the giant freshwater prawns and reducing cannibalism.

[0051] 3. Aquaculture process management phase (April-October 2025) 3.1 Oxygenation Management On a daily basis, the oxygenation equipment should be turned on according to the specifications to ensure that the oxygenation time is no less than 12 hours per day. During the high temperature period of summer (July-August), rainy days, and when the dissolved oxygen level in the water is low (≤5mg / L), the oxygenation time should be extended to more than 16 hours. During this period, waterwheel aerators and impeller aerators should be turned on simultaneously to improve the oxygenation efficiency. The dissolved oxygen level in the water should be tested regularly to ensure that it is always maintained above 5mg / L to ensure the normal growth of giant freshwater prawns and polyculture fish.

[0052] 3.2 Water quality control Observe the water color of each pond regularly every 15 days. When slight turbidity is found in some ponds, promptly use a compound microbial preparation to adjust the water quality. Apply 3 catties / mu of fermented microbial agent (sealed and left to stand at 30℃ for 3-5 days) and then evenly sprinkle it throughout the pond. Sprinkle on sunny mornings only. After sprinkling, turn on the aeration equipment for 2.5 hours to promote full integration of the microbial preparation with the water and ensure stable water quality. The compound microbial preparation contains EM bacteria and Lactococcus gasseri C6a2, with the EM bacteria (Hubei Fengtian Biotechnology Co., Ltd.) having a live bacteria content ≥5×10⁻⁶. 11 CFU / catties, viable count of Lactococcus gasseri C6a2 ≥1×10⁻⁶ 11 CFU / catties.

[0053] 3.3 Bottom sediment improvement and water detoxification Bottom sediment improvement and water detoxification were carried out every 7 days. For bottom sediment improvement, a biological bottom conditioner (Jiangsu Lvke Biotechnology Co., Ltd.) was used, applied evenly to the entire pond at a dosage of 3 catties / mu (approximately 1.5 kg / acre). Special attention was paid to areas where shrimp larvae congregated and where uneaten feed and feces easily accumulated, ensuring the conditioner fully covered the pond bottom to improve the bottom environment and reduce the growth of pathogenic microorganisms. Simultaneously, sodium humate was used for water detoxification at a dosage of 1 catties / mu (approximately 0.5 kg / acre), fully dissolved and then applied to the entire pond. After detoxification, the pH, ammonia nitrogen, and nitrite levels of the water were tested and maintained within the suitable range for the growth of giant freshwater prawns (pH 7.0-8.5, ammonia nitrogen ≤0.5 mg / L, nitrite ≤0.1 mg / L).

[0054] 3.4 Daily monitoring and feeding management Observe the activity and feeding status of the giant freshwater prawns and polyculture fish daily, and adjust the feed amount according to the amount of uneaten food; regularly test water quality indicators such as temperature, pH, and dissolved oxygen, and keep detailed records; check the growth of aquatic plants weekly, and promptly remove withered and rotten plant debris to ensure the stability of the ecosystem. A phased, quantitative feeding model is adopted (Table 2), and the feed amount is rationally planned (feed purchased from Guangdong Haid Group Co., Ltd.) to reduce pollution from uneaten feed.

[0055] Table 2. Staged Feeding Standards for Giant Freshwater Prawns 4. Harvest Phase (August 21, 2025 - October 18, 2025) Based on the growth size of the giant freshwater prawns and market demand, harvesting began on August 21, 2025, and was completed by October 18. Feeding was stopped 1-2 days before harvest to reduce the fat content of the prawns and improve quality. A gillnet method was used for harvesting in batches, prioritizing the harvesting of larger adult prawns while allowing smaller prawns to continue growing, thus improving overall aquaculture efficiency. The polyculture fish were harvested simultaneously with the giant freshwater prawns during the later stages of the harvest.

[0056] 5. Aquaculture Results 5.1 Output and Output Value The total yield of the 150-mu (approximately 10 hectares) giant freshwater prawn farming project was 52,603 ​​jin (approximately 26,803 catties), with an average yield of 876.72 jin (approximately 488.3 kg) per mu (approximately 118.3 kg per hectare). The average price of the prawns during the sales phase was 23.28 yuan per jin (approximately 11.28 yuan per catties), and the total output value of the farming project reached 1,224,620 yuan.

[0057] 5.2 Costs and Profits The total cost of this aquaculture project was 783,900 yuan, including 287,800 yuan for feed (feed ratio 1.54), 108,000 yuan for shrimp fry, 156,000 yuan per year for pond rent, 31,000 yuan for electricity, 15,000 yuan for animal health products, 5,000 yuan for labor, and 181,100 yuan for other expenses. The net profit was 440,720 yuan, with an average profit of 7,345 yuan per mu, demonstrating significant economic benefits.

[0058] 5.3 Ecological Benefits No chemical agents were used in the aquaculture process. Through the shrimp-grass-fish three-dimensional ecological system and biological regulation measures, the ecological balance of the water body was effectively maintained, the water environment was stable, and the incidence of diseases was extremely low. This achieved the coordinated development of ecological protection and aquaculture production, and verified the core advantage of the invention in achieving a win-win situation for both ecology and economy.

[0059] All other parts not described in detail are existing technologies. Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, not all embodiments. People can obtain other embodiments based on these embodiments without creative effort, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A method for cultivating giant freshwater prawns in a shrimp-grass-fish co-culture system based on probiotic regulation, characterized in that: Includes the following steps: (1) Drain the remaining water in the pond, dry the pond and expose it to the sun and turn over the bottom mud; after the sun exposure, fill the pond with water and disinfect the whole pond with bleaching powder. After disinfection, let it stand and then carry out the first stage of aquatic plant planting. (2) Release giant freshwater prawn larvae, and simultaneously raise fish larvae in the same manner after release. Then, carry out the second stage of aquatic plant planting 7-10 days after release to construct a three-dimensional ecological system of shrimp-grass-fish. (3) Maintain dissolved oxygen in the water body ≥5 mg / L through oxygenation equipment management; spray compound microbial agents into the pond every 14-16 days after stocking to regulate water quality; apply biological bottom conditioner and sodium humate every 6-8 days after stocking to improve bottom sediment and detoxify water; and test the pH value, ammonia nitrogen and nitrite index of the water body. (4) After the seedlings are released, they are fed daily. After the feeding is completed, they are caught using a ground cage net. The mixed-culture fish are caught simultaneously with the later harvest of the giant freshwater prawn.

2. The method for farming giant freshwater prawns according to claim 1, characterized in that: In step (1), the drying and sun exposure of the pond lasts for 15 to 25 days; the amount of bleaching powder used is 45 to 55 catties / mu; after disinfection, the pond is left to stand until the residual chlorine content drops below 0.05 mg / L before proceeding to the next step. The first stage of aquatic plant planting involves planting Elodea in the pond 3-5 days before releasing Giant freshwater prawn larvae using a hole planting method at intervals of 2-3 meters, with one plant per hole and a planting depth of 3-5 cm.

3. The method for farming giant freshwater prawns according to claim 1, characterized in that: In step (2), the stocking density of giant freshwater prawn larvae is 40,000 to 50,000 prawns per mu. Before stocking, the transport bags of giant freshwater prawn larvae are placed in the corresponding pond water and soaked for 20 to 30 minutes to adapt to the temperature. When the temperature difference between the water inside the bag and the pond water drops to below 2°C, the bag is opened to allow the giant freshwater prawn larvae to swim into the pond on their own. The mixed-culture fish fry are silver carp fry and crucian carp fry. The stocking density of silver carp fry is 5-7 fish / mu, and the stocking density of crucian carp fry is 6-8 fish / mu. The mixed-culture fish fry are evenly distributed in each pond area after adopting the same temperature adaptation method as the giant freshwater prawn fry.

4. The method for farming giant freshwater prawns according to claim 1, characterized in that: In step (2), the second stage of aquatic plant planting specifically involves planting water spinach by cuttings at a depth of 10-20 cm along the banks of each pond, with a plant spacing of 15-20 cm.

5. The method for farming giant freshwater prawns according to claim 1, characterized in that: In step (3), the oxygenation equipment includes a waterwheel aerator and an impeller aerator, with power of 10~15 kW and 30~40 kW per mu respectively.

6. The method for farming giant freshwater prawns according to claim 1, characterized in that: In step (3), the amount of compound microbial preparation applied is 2-4 catties / mu. After fermentation, the compound microbial preparation is evenly applied to the entire pond. After application, the oxygenation equipment is turned on for 2-3 hours.

7. The method for farming giant freshwater prawns according to claim 6, characterized in that: The compound microbial preparation contains EM bacteria and Lactococcus gasseri C6a2, and the viable count of EM bacteria in the compound microbial preparation is ≥5×10⁻⁶. 11 CFU / catties, viable count of Lactococcus gasseri C6a2 ≥1×10⁻⁶ 11 CFU / catties; The fermentation treatment conditions were: sealed and static culture at 30°C for 3-5 days. The preservation number of the Lactococcus gasseri C6a2 is CGMCC NO: M 31024.

8. The method for farming giant freshwater prawns according to claim 1, characterized in that: In step (3), the application rate of the biological bottom conditioner is 2-4 catties / mu, and the application rate of sodium humate is 0.8-1.2 catties / mu. After bottom sediment improvement and water detoxification, the water quality indicators are maintained at: pH 7.0~8.5, ammonia nitrogen content ≤0.5 mg / L, and nitrite content ≤0.1 mg / L.

9. The method for farming giant freshwater prawns according to claim 1, characterized in that: In step (4), the daily feeding is specifically as follows: When the weight of giant freshwater prawns is <1 g, the daily feeding rate is 15-20% of their body weight; when the weight of giant freshwater prawns is 1-5 g, the daily feeding rate is 8-10% of their body weight; and when the weight of giant freshwater prawns is >5 g, the daily feeding rate is 3-6% of their body weight.

10. The application of the Macrobrachium rosenbergii farming method according to any one of claims 1 to 9 in the large-scale ecological farming of Macrobrachium rosenbergii.

Images