Large-scale culture method for daphnia magna in Sulawesi province

Abstract

The invention provides a large-scale culture method for daphnia glabra. The culture method comprises the following steps: S1, disinfecting and cleaning a culture device; s2, water feeding: filtering and disinfecting seawater, then mixing the seawater with disinfected fresh water, and adding the mixed water into the culture device; s3, inoculating: inoculating the Sulawesi daphnia xiaoensis obtained by breeding in the water body of the culture device at the inoculation density of 460-550 daphnia xiaoensis / L, and controlling the concentration of dissolved oxygen in the water body to be 10-14 mg / L; s4, feeding; s5, changing water, namely changing the water body in the culture device by using the filtered and disinfected seawater every day. According to the method, through scientific culture condition and environment control, the production efficiency is remarkably improved, the culture cost is reduced, and stable and high-density culture of the Sulawesi daphnia magna is achieved. According to the technology, the defect of seawater live bait supply is overcome, a stable, reliable and high-quality live bait source can be provided for aquatic breeding, and wide application prospects are achieved.

Description

Technical Field

[0001] The invention belongs to the technical field of cultivation of Daphnia pulex, and in particular relates to a large-scale cultivation method of Daphnia pulex. Background Art

[0002] Existing aquatic baits are mainly rotifers, large daphnia (water spiders), and copepods, which are large and unpalatable to groupers. Freshwater daphnia are difficult to adapt to and survive in seawater due to their own osmotic pressure regulation mechanism. In the general grouper breeding process, the process of bait delivery is from ss rotifers (80-110 microns) to large rotifers (180-300 microns), and then directly feed newly hatched Artemia nauplii (600 microns). During this period, there is a large gap in bait. Although copepods can fill it, copepods face great difficulties in actual mass production due to sexual reproduction. When some species of groupers adapt to bait, they will not eat dead ones or unpalatable ones. Moreover, if only Artemia larvae are fed, gill stress is easily caused. The reason is that unsaturated fatty acids have some response mechanisms to fry stress, so fish death and fry reduction are easily caused during the breeding process.

[0003] The Sulawesi scleroderma is a member of the genus Scleroderma of the family Daphnia. Its larvae are 320 microns and its adults are 800 microns, which can just fill the gap between large rotifers and Artemia larvae. In addition, the nutrition of the cladoceran scleroderma is not worse than that of Artemia larvae, and its unsaturated fatty acid content is higher than that of Artemia larvae. The effect of breeding and feeding is better, and the survival rate of fry is higher. As a nutritious and highly palatable aquatic bait, the Sulawesi scleroderma is suitable for a variety of aquaculture species, especially in the nursery stage of marine fish and crustaceans. It can be used as a high-quality substitute for some traditional live baits (such as rotifers and Artemia), which helps to improve the survival rate of seedlings and the efficiency of nursery.

[0004] However, the Sulawesi sphenodontia is a pure marine species with a relatively low density in natural seawater, making it difficult to collect from nature, and the Sulawesi sphenodontia aquaculture industry is currently blank. Therefore, a large-scale aquaculture method for Sulawesi sphenodontia is needed in the art to carry out large-scale and batch aquaculture of Sulawesi sphenodontia, filling the gap in the Sulawesi sphenodontia aquaculture industry. Summary of the invention

[0005] Based on this, the object of the present invention is to provide a large-scale breeding method for Sulawesi Xiusotho Daphnia sulawesi, which can achieve a unit volume density of Sulawesi Xiusotho Daphnia sulawesi of more than 6000 pieces / L after 14 days of breeding, significantly improving production efficiency and reducing breeding costs.

[0006] To achieve the above objectives, the present invention adopts the following technical solutions.

[0007] A first aspect of the present invention provides a method for cultivating Daphnia sulawesiensis, the method comprising the following steps:

[0008] S1. Disinfect and clean the breeding equipment;

[0009] S2. Water entry: filtering and disinfecting seawater, then mixing it with disinfected fresh water, and adding the mixed water into the aquaculture device;

[0010] S3. Inoculation: Inoculate Daphnia sulawesiensis in the water of the aquaculture device at an inoculation density of 460 to 550 daphnia gracilis / L, and control the dissolved oxygen concentration in the water to be 10 mg / L to 14 mg / L;

[0011] S4. Feeding: feeding feed into the breeding device;

[0012] S5. Water change: Use filtered and disinfected seawater to replace the water in the aquaculture device every day.

[0013] In some embodiments, the feed comprises at least 60% (by weight) concentrated Nannochloropsis.

[0014] In some preferred embodiments, the feed comprises the following components in mass fractions: 60% to 80% concentrated Nannochloropsis edulis, 15% to 35% Isochrysis zhanjiangensis or marine photosynthetic bacteria or Chaetoceros muehii.

[0015] Further preferably, the feed comprises the following components in mass fractions: 60% to 80% concentrated Nannochloropsis, 15% to 35% Isochrysis zhanjiangensis or marine photosynthetic bacteria or Chaetoceros muelleri, and 3% to 7% hydrolyzed fish oil or Schizochytrium powder.

[0016] More preferably, the feed comprises the following components in mass fractions: 65% to 75% concentrated Nannochloropsis, 20% to 30% Isochrysis zhanjiangensis or marine photosynthetic bacteria or Chaetoceros muelleri, and 4% to 6% hydrolyzed fish oil or Schizochytrium powder.

[0017] In some embodiments, the total amount of feed fed into the aquaculture device per day is 1 to 3 mg of dry weight feed per liter of water; preferably, the feed is fed in 2 to 3 times.

[0018] In some embodiments, the Daphnia sulawesiensis inoculated in step S3 is obtained by breeding using the following breeding method:

[0019] (1) Sampling: ① Collect water samples from the sea and place them in a sterilized container containing ultrafiltered seawater; ② Add mixed algae to the container so that the concentration of algae in the container is 1×10 5 / L~5×10 5 / L; the mixed algae liquid contains Chlorella, Nannochloropsis, and Thalassiosira, and the mass ratio of Chlorella, Nannochloropsis, and Thalassiosira is (1-2): (1-2): (1-2); ③ an air stone is placed in the container for micro-aeration culture; ④ the water is changed every day and the mixed algae is added to allow Daphnia sulawesiensis to develop into a dominant species;

[0020] (2) Screening: After Daphnia sulawesiensis develops into a dominant species, adult Daphnia gracilis are screened from it;

[0021] (3) Breeding and domestication: ① Domestication with feed: the adult Daphnia sulawesi obtained by screening in step (2) is domesticated to accept concentrated Nannochloropsis sulawesi; ② Selective breeding: the Daphnia sulawesi domesticated with feed containing a large amount of Daphnia sulawesi is selected and cultivated to obtain Daphnia sulawesi for inoculation.

[0022] In some embodiments, the gas volume in step (1) is controlled to be 1 to 2 bubbles per second.

[0023] In some embodiments, the step (3) is to obtain the concentrated Nannochloropsis sulweziensis by acclimatization for at least 5 generations.

[0024] The breeding method of the present invention can successfully propagate Daphnia sulawesiensis obtained from seawater samples in large quantities and accept concentrated Nannochloropsis sulawesiensis, which is convenient for subsequent large-scale cultivation.

[0025] In the breeding method of the present invention, the composition of the mixed algae is critical to the successful cultivation of the Sulawesi Daphnia sulawesi isolated from seawater. After extensive research, the inventors found that a mixed algae composed of Chlorella vulgaris, Nannochloropsis sulawesi and Thalassiosira marinum in a mass ratio of (1-2): (1-2): (1-2) is more conducive to the breeding and cultivation of the Sulawesi Daphnia sulawesi isolated from seawater.

[0026] In some embodiments, the dissolved oxygen concentration of water in the aquaculture device is controlled to be 12 mg / L to 14 mg / L.

[0027] In some embodiments, the salinity of the mixed water in step S2 is 20 ppt to 25 ppt.

[0028] In some embodiments, the volume of water replaced every day in step S5 is 15% to 25% of the total volume of the water in the aquaculture device. The inventors found during the aquaculture process that replacing new water every day stimulates the sulawesi Daphnia sulawesi to carry Daphnia sulawesi, especially when the volume of the replaced water is 15% to 25% of the total volume of the water in the aquaculture device, it can stimulate the sulawesi Daphnia sulawesi to produce a large number of carrying Daphnia sulawesi.

[0029] In some embodiments, the ammonia nitrogen concentration of the water in the aquaculture device is less than 0.5 mg / L, the pH value is 7.5-8.5, and the transparency is >50 cm; and / or,

[0030] The water level of the aquaculture device is 60 cm to 80 cm; and / or,

[0031] The temperature of the water in the aquaculture device is 30°C ± 3°C; and / or,

[0032] In step S3, Daphnia sulawesiensis is inoculated after the circulating water of the aquaculture device has been running for 3 to 5 days.

[0033] In some embodiments, the method for disinfecting seawater and / or freshwater is: treating with 0.8-1 ppm ozone for 20-30 minutes; preferably combined with ultraviolet disinfection.

[0034] A second aspect of the present invention provides the use of the above-mentioned aquaculture method in preparing fish or crustacean bait.

[0035] In some embodiments, the fish is grouper.

[0036] The third aspect of the present invention provides a feed for Daphnia sulawesiensis, which comprises the following components in mass fractions: 60% to 80% concentrated Nannochloropsis sulawesiensis, 15% to 35% Isochrysis zhanjiangensis or marine photosynthetic bacteria or Chaetoceros muelleri.

[0037] In some embodiments, the feed comprises the following components in mass fractions: 60% to 80% concentrated Nannochloropsis spp., 15% to 35% Isochrysis zhanjiangensis or marine photosynthetic bacteria or Chaetoceros muehii, and 3% to 7% hydrolyzed fish oil or Schizochytrium schizochytrium powder.

[0038] In some preferred embodiments, the feed comprises the following components in mass fractions: 65% to 75% concentrated Nannochloropsis edulis, 20% to 30% Isochrysis zhanjiangensis or marine photosynthetic bacteria or Chaetoceros muelleri, and 4% to 6% hydrolyzed fish oil or Schizochytrium powder.

[0039] In some embodiments, the feed further comprises 1% to 3% yeast extract.

[0040] In some embodiments, the particle size of the feed is 10 μm to 50 μm.

[0041] The present invention provides a large-scale breeding method for Sulawesi sampaedia, which can effectively reduce environmental load and reduce feed waste by optimizing the breeding conditions of Sulawesi sampaedia, especially controlling a series of key parameters such as inoculation density, feed feeding amount and dissolved oxygen concentration of water body, while making full use of the high reproduction of Sulawesi sampaedia, shortening the production cycle, reducing dependence on natural resources, and providing technical support for the sustainable development of aquaculture industry. In addition, the breeding method of the present invention has low requirements on equipment and technical level, is easy to operate, and is suitable for promotion to small and medium-sized breeding farms.

[0042] The breeding method of the present invention is suitable for a multi-pond linkage breeding system, and the ponds are connected by a pipeline circulating water system, which can achieve unified water quality management and efficient water exchange, and further improve production efficiency.

[0043] Furthermore, the inventors of the present invention have obtained the above-mentioned feed for S. sulawesiensis based on their many years of breeding experience and a large number of experimental optimizations. The nutritional components of the feed are more conducive to the growth of S. sulawesiensis and can effectively increase the unit volume density of S. sulawesiensis.

[0044] The present invention, through scientific culture conditions and environmental control, and optimized feed feeding, can achieve a unit volume density of 6,000 pieces / L or more of Sulawesi sampanthellae in 14 days of cultivation, significantly improving production efficiency and reducing cultivation costs, achieving stable and high-density cultivation of Sulawesi sampanthellae, and providing feasibility for large-scale cultivation. In the context of the current prominent contradiction between supply and demand in the biological bait market, the technology of the present invention fills the shortage of live bait supply in seawater, can provide a stable, reliable and high-quality live bait source for aquatic seedling cultivation, and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0045] Figure 1 This is the Daphnia sulawesiensis placed in the culture dish during the breeding process of Example 1.

[0046] Figure 2 Sulawesi Daphnia obtained for breeding. DETAILED DESCRIPTION

[0047] The experimental methods in the following examples of the present invention, where no specific conditions are specified, are usually carried out under conventional conditions or under conditions recommended by the manufacturers. The various commonly used chemical reagents used in the examples are all commercially available products.

[0048] Unless otherwise defined, all technical and scientific terms used in the present invention have the same meaning as those commonly understood by those skilled in the art of the present invention. The terms used in the specification of the present invention are only for the purpose of describing specific embodiments and are not intended to limit the present invention.

[0049] The terms "including" and "having" and any variations thereof of the present invention are intended to cover non-exclusive inclusions. For example, a process, method, device, product or equipment comprising a series of steps is not limited to the listed steps or modules, but may optionally include steps not listed, or may optionally include other steps inherent to these processes, methods, products or equipment.

[0050] The "and / or" mentioned in the present invention describes the association relationship of the associated objects, indicating that there may be three relationships. For example, A and / or B can represent: A exists alone, A and B exist at the same time, and B exists alone. The character " / " generally indicates that the associated objects before and after are in an "or" relationship.

[0051] The following is a description with reference to specific implementation methods.

[0052] Example 1

[0053] This embodiment provides a large-scale cultivation method of Daphnia sulawesiensis, comprising the following steps:

[0054] Breeding

[0055] (1) Sampling

[0056] 1. First, process the water sample collected from the sea and put it into a disinfected 800L barrel, and put 1 / 4 fresh ultra-filtered seawater in the barrel in advance;

[0057] 2. Provide a mixed algae solution of fresh Chlorella, Nannochloropsis, and Thalassiosira (algae with high protein and unsaturated fatty acid content) with a mass ratio of 1:1:1. The algae concentration is controlled at 5×10 5 Second power, slightly green water concentration;

[0058] 3. Prepare an air stone and micro-aerate the culture. Do not use pure oxygen or nano oxygen. Particularly tiny bubbles will adhere to the surface of the insect body and cause its death. Adjust the air volume to 1-2 bubbles per second.

[0059] 4. During the culture period, observe its growth and development as well as the water conditions, change the water and add algae every day to maintain the algae concentration, let it develop and reproduce stably, form a certain biomass, and wait for the Sulawesi Daphnia to develop into a dominant species.

[0060] (2) Screening (separation and purification)

[0061] When the Sulawesi xiusoda develops into a dominant species (because when the environment is suitable, the Sulawesi xiusoda will reproduce parthenogenetically, without the need for mating, without the need for males, and can grow in large numbers, but the copepods screened at the same time need to mate before laying eggs, both the development time and the number of reproduction will be faster than the copepods, so after a period of breeding, the Sulawesi xiusoda will slowly become the dominant species. At this time, you can use a flashlight to shine on it from the side, and you can see a large number of Sulawesi xiusoda moving against the light. After filtering out the mixed state of various plankton (including copepods, rotifers, etc.), use a 200-mesh sieve to filter out the mixed state of various plankton (including copepods, rotifers, etc.). Prepare two 1L beakers and put them in filtered ultra-pure seawater. The temperature difference cannot be greater than 3°C. Put the worms in one beaker and put the water in another. Prepare three borosilicate culture dishes and a 5ml disposable pipette. Cut off some of the ends and trim them slightly to make them smooth (only a little trimming is needed, not too much trimming will affect the absorption) to prevent injury to the worms. Figure 1 As shown, choose a black acrylic plate as the background and put 3 culture dishes on it and put some seawater in it. Absorb some worms and place them in the first culture dish. Use a flashlight to shine a direct light source on the side and blow gently with a straw until it is even. After standing for a while, it will be obvious that copepods will be strongly attracted to light due to phototaxis, while Sulawesi S. sulawesi is a photophobic species (anti-phototaxis) and will escape. At this time, find a Sulawesi S. sulawesi that swims normally (use the swimming method to judge its health status) and absorb it into the second culture dish. Blow gently to avoid absorbing other or smaller organisms such as rotifers. After confirming that it is clean, absorb it into the third culture dish for use. The second culture dish needs to be replaced with new water after absorbing an average of about 10 worms. Do not put the young worms back into the culture bucket for continued cultivation. Only adult worms (normal swimming posture, fast speed, and no parasites and attached flocs on the body surface) are selected. Repeat the above operations to collect a sufficient amount of Sulawesi S. sulawesi.

[0062] (3) Breeding and domestication

[0063] Bait acclimation: slowly adjust the proportion of the mixed algae liquid, increase the proportion of Nannochloropsis, acclimate it to accept Nannochloropsis, and after it can accept only feeding Nannochloropsis, add concentrated Nannochloropsis little by little to replace fresh Nannochloropsis. The duration must be slow, at least 5 generations according to current experimental data.

[0064] During the bait conversion period, to determine whether the bait is palatable, you can use a side light source to observe its feces. If its feces are dragged in strips, it can be determined that this bait is palatable. If it is not palatable, it will be scattered in the water in the form of sparse fragments.

[0065] Selective breeding: When they grow to a certain number, use a 180-mesh scoop net to gently scoop them out and put them into a beaker. Use a disposable pipette to transfer them to a 6-well plate and observe the number of fleas under a stereomicroscope or inverted microscope (40 times). Be patient and select 3 of the best health and the most fleas from 1,000. Figure 2 ), gently suck it out with a disposable straw, and when it accumulates to a certain number, put it in a constant temperature incubator (the larger the water body in the front, the better, and you can try a small water body later) for separate cultivation. After it has a certain biomass, repeat the operation several times to increase its unit density for subsequent large-scale breeding.

[0066] Batch farming

[0067] S1. Construction of a culture pond: Build a 4-meter-long, 4-meter-wide, and 1.2-meter-high cement culture pond or set up a glass fiber reinforced polyester culture water tank of the same specification, using a 5% concentration of Ca(ClO) 2 Solution or one of the drugs with a concentration of 5% NaClO solution for disinfection, then clean thoroughly, drain the drugs and wash the breeding equipment with clean water;

[0068] S2. Water entry: natural seawater is filtered through a coarse filter to remove larger particles, and then finely filtered using a 100-mesh cotton filter bag, and then treated with 0.8-1ppm ozone for 20 minutes, combined with ultraviolet sterilization equipment disinfection (further improve the safety of water quality and reduce the risk of introducing pathogens), and naturally aerated for 24 hours. The treated natural seawater is then mixed with disinfected fresh water that has undergone the same treatment to reach a salinity of 20ppt-25ppt, and the water level of the aquaculture device is 60-80cm, and a heater is used to maintain a constant temperature of 30°C;

[0069] S3. Inoculation: After the water in the breeding pond circulates for 3 to 5 days to ensure that the water quality is stable, inoculate the Sulawesi Xiuso Daphnia obtained from the above breeding. Before inoculation, the health status of the population needs to be screened, and the Xiuso Daphnia with strong vitality and plump individuals are selected as the source of the population to ensure the excellent reproductive performance of the population. The inoculation density is controlled at about 500 / L to provide a sufficient initial population size, while avoiding excessive density that leads to increased individual competition. After inoculation, continuous oxygen supply is required to ensure that the dissolved oxygen concentration in the water is stably maintained at about 13 mg / L to meet the oxygen demand of the Sulawesi Xiuso Daphnia. Oxygen is evenly distributed through the microporous aeration device, and the bubbles are fine (the aeration device uses a larger air stone for micro-aeration culture, and can be matched with a nano air stone for pure oxygen. Aeration is the main method, and pure oxygen is the auxiliary method. The amount of air cannot be too large, because its own swimming ability is weak and it cannot resist the impact of larger water flow. The bubbles cannot be too small. If the bubbles are too small, they will adhere to the surface of the insect body and cause it to float on the surface of the water body), avoiding mechanical damage to the daphnia due to the flow of large bubbles;

[0070] S4. Use artificial mixed feed for feeding. The feed formula is: 70% concentrated Nannochloropsis, 25% Zhanjiang Isochrysis and 5% eicosapentaenoic acid (EPA) oil. The feed particle size is controlled at 10-50μm to ensure that Sulawesi Daphnia can feed efficiently and meet its growth and reproduction needs. The recommended feeding frequency is 2-3 times a day to ensure that Daphnia individuals continue to feed and prevent water quality deterioration caused by overfeeding. The total daily feeding amount is controlled to 2 mg dry weight per liter of water. When feeding, the feed is evenly dispersed in the water to avoid excessive or insufficient local feed. Use an automatic quantitative feeding device to feed regularly at a frequency of once every 3 hours to ensure that the feed is evenly distributed in the water. During the feeding process, the residual feed concentration in the water should be regularly tested, and the residual feed should be observed by centrifugation of trace water samples to ensure that the residual feed is not excessive (less than 10% of the total feed amount). If residual bait is found to accumulate or water transparency is significantly reduced, the amount of feed should be appropriately reduced and part of the water should be replaced in time;

[0071] S5. Quantitatively replace 15-25% of the water in the culture pond every day, using natural seawater that has been filtered and disinfected multiple times to ensure stable water quality and avoid the introduction of pathogenic microorganisms. The water change process needs to be operated with caution, not only to ensure the renewal of water quality, but also to avoid disturbing the growth environment of Sulawesi Daphnia. Before changing the water, use a sand filter tank to preliminarily filter the natural seawater (flow rate 5-8L / min, filtration time 30 minutes) to remove suspended impurities and larger particulate matter. The seawater is then filtered through 60-mesh and 100-mesh sieves in turn, and then deep filtered using a cotton filter bag (200 mesh) to completely remove fine suspended matter and potential biological pollutants. The filtered seawater also needs to be treated with ozone, and ozone with a concentration of 0.5-1ppm is injected for 15-20 minutes to further kill microorganisms and pathogens. To eliminate ozone residues, the filtered seawater can be placed in the culture pond after standing for 1 hour. According to the water quality, inorganic salts can be added in appropriate amounts to maintain the stability of the water salinity (20-25ppt). When changing water, a fine screen of 80-100 mesh is needed to cover the drain outlet to prevent the loss of Sulawesi Daphnia pulex from being discharged with the water flow. Drainage should be carried out slowly, and the water flow rate should be controlled to ensure uniform exchange of water layers and avoid disturbing the sediments at the bottom of the pond. After the water change is completed, the water stirring intensity needs to be appropriately increased to ensure uniform distribution of Daphnia pulex and avoid hypoxia in areas with too high density. In order to monitor the effect of water change, the key parameters of the aquaculture pond water need to be tested after the water change, including: dissolved oxygen concentration (maintained at 13mg / L), ammonia nitrogen concentration (<0.5mg / L), pH value (7.5-8.5) and transparency (>50cm). If the water quality indicators are found to be abnormal, the water change ratio or frequency needs to be adjusted in time;

[0072] During the cultivation process, a mosquito-proof cover is usually added, otherwise mosquito larvae will be produced and compete for bait and living space.

[0073] S6. Stop feeding for 6 to 8 hours before harvesting to reduce suspended particles and undigested feed in the aquaculture water, improve collection efficiency and reduce the impact on subsequent water quality. To avoid sudden changes in the environment affecting the remaining population, do not inflate 2 hours before harvesting, maintain the dissolved oxygen concentration at 6 to 8 mg / L, and ensure that the water temperature is constant at 30°C. At this time, the daphnia will float on the surface of the water due to lack of oxygen in the water. When harvesting, place a wider 100-mesh silk mesh bag steadily into the aquaculture pond, and collect in batches by slowly stirring the mesh bag to avoid excessive squeezing and mechanical damage to individual daphnia. During the collection process, the water in the mesh bag needs to be dumped into the collection container at regular intervals, and the water temperature and salinity in the collection container must be kept consistent with the aquaculture pond to reduce the stress response of individual daphnia caused by sudden changes in the environment. The harvesting process is combined with a water flow guide device to guide the uncollected daphnia to the center of the pond to avoid too many individuals from scattering on the wall or bottom of the pond due to stirring. The harvest is carried out in batches to ensure the subsequent reproduction capacity of the remaining Daphnia population. The amount collected each time is controlled at 30% to 50% of the total amount of the breeding pond, and new water is added in time to maintain the stability of the environment in the pond. After collection, the harvested Sulawesi Daphnia can be temporarily reared for a short period of time. During the temporary rearing process, trace oxygen is provided, and the water temperature and salinity are kept stable to further restore the vitality of the Daphnia. The temporarily reared Daphnia can be directly used for aquatic seedling feeding or deep processing (such as frozen storage or drying preparation) to meet different breeding needs.

[0074] During the breeding process, an intelligent water quality monitoring system is installed to record water temperature, dissolved oxygen, pH value and ammonia nitrogen concentration in real time, and automatically adjust the water quality parameters to an appropriate range when abnormalities occur.

[0075] During the breeding process, a small amount of sodium chloride (1g / L) is added every 5 days to regulate the water to inhibit the reproduction of pathogenic microorganisms. At the same time, the health status of the daphnia is regularly observed under a microscope, and any abnormalities are immediately isolated and handled.

[0076] In this embodiment, a total of 8 culture ponds were built for culture. During the culture process, the unit volume density of Daphnia sulwesiensis in the culture ponds was tested on days 0, 2, 4, 6, 8, 10, 12 and 14.

[0077] The detection method is: increase the inflation volume 5 minutes in advance to make it evenly distributed in the water, use a beaker to quickly pour 1 liter of water into a measuring cylinder, measure 1000ml of water, and then pour it into a new measuring cylinder to fix it with formalin in equal proportion, let it settle for 30 minutes to wait for it to die, and suck 1ml of plankton from the bottom and count it in the technical counting box.

[0078] Fill in the test results in Table 1.

[0079] Table 1

[0080]

[0081]

[0082] As can be seen from Table 1, the breeding method of the present invention can achieve a unit volume density of 6000-6400 Daphnia sulawesiensis / L after 14 days of cultivation, which significantly improves production efficiency, reduces breeding costs, and provides feasibility for large-scale breeding.

[0083] Example 2

[0084] This embodiment provides a large-scale cultivation method of Daphnia sulawesiensis. The cultivation method is basically the same as that of Example 1 except for the feed formula, and the feed formula is: 100% concentrated Nannochloropsis.

[0085] The bottom was drained and the water was changed regularly every day (same as in Example 1).

[0086] In this embodiment, a total of 8 culture ponds were built for culture. During the culture process, the unit volume density of Daphnia sulwesiensis in the culture ponds was tested on days 0, 2, 4, 6, 8, 10, 12 and 14.

[0087] The detection method is: increase the inflation volume 5 minutes in advance to make it evenly distributed in the water, use a beaker to quickly pour 1 liter of water into a measuring cylinder, measure 1000ml of water, and then pour it into a new measuring cylinder to fix it with formalin in equal proportion, let it settle for 30 minutes to wait for it to die, and suck 1ml of plankton from the bottom and count it in the technical counting box.

[0088] Fill in the test results in Table 2.

[0089] Table 2

[0090] date 0 2 4 6 8 10 12 14 Breeding pond 1 533 762 1234 1987 2567 2980 3545 4023 Breeding pond 2 455 957 1011 1326 1980 2444 4105 4328 Breeding pond 3 511 982 1123 1268 1756 1958 2741 2890 Breeding pond 4 477 785 1354 1577 1625 2541 3012 3742 Breeding pond 5 429 775 1659 1950 2011 2633 2644 3410 Breeding pond 6 463 958 999 1753 1987 2256 2461 2676 Breeding pond 7 529 852 857 1666 2068 2468 3037 3480 Breeding pond 8 533 980 985 1456 1576 2107 3485 4190

[0091] The above results show that the maximum unit volume density of Daphnia sulawesi can reach 4328ind / L after 14 days of cultivation using the method of this embodiment.

[0092] Example 3

[0093] This embodiment provides a large-scale cultivation method of Daphnia sulawesiensis, which is basically the same as that of Example 1 except for the feed formula, and the feed formula is: 75% concentrated Nannochloropsis sulawesi and 25% Chaetoceros muelleri (diatom).

[0094] The bottom was drained and the water was changed regularly every day (same as in Example 1).

[0095] In this embodiment, a total of 8 culture ponds were built for culture. During the culture process, the unit volume density of Daphnia sulwesiensis in the culture ponds was tested on days 0, 2, 4, 6, 8, 10, 12 and 14.

[0096] The detection method is: increase the inflation volume 5 minutes in advance to make it evenly distributed in the water, use a beaker to quickly pour 1 liter of water into a measuring cylinder, measure 1000ml of water, and then pour it into a new measuring cylinder to fix it with formalin in equal proportion, let it settle for 30 minutes to wait for it to die, and suck 1ml of plankton from the bottom and count it in the technical counting box.

[0097] Fill in the test results in Table 3.

[0098] Table 3

[0099]

[0100] According to the test, the maximum unit volume density of Daphnia sulawesiensis after 14 days of cultivation according to the cultivation method of this embodiment is about 3611ind / L.

[0101] Example 4

[0102] This embodiment provides a large-scale cultivation method of Daphnia sulawesiensis, which is basically the same as that of Example 1 except for the feed formula, and the feed formula is: 75% concentrated Nannochloropsis sulawesiensis and 25% Isochrysis zhanjiangensis.

[0103] The bottom was drained and the water was changed regularly every day (same as in Example 1).

[0104] In this embodiment, a total of 8 culture ponds were built for culture. During the culture process, the unit volume density of Daphnia sulwesiensis in the culture ponds was tested on days 0, 2, 4, 6, 8, 10, 12 and 14.

[0105] The detection method is: increase the inflation volume 5 minutes in advance to make it evenly distributed in the water, use a beaker to quickly pour 1 liter of water into a measuring cylinder, measure 1000ml of water, and then pour it into a new measuring cylinder to fix it with formalin in equal proportion, let it settle for 30 minutes to wait for it to die, and suck 1ml of plankton from the bottom and count it in the technical counting box.

[0106] Fill in the test results in Table 4.

[0107] Table 4

[0108] date 0 2 4 6 8 10 12 14 Breeding pond 1 482 885 1025 1415 2015 3612 4580 5101 Breeding pond 2 560 843 1305 1621 2564 3329 4010 4826 Breeding pond 3 427 868 1515 1915 2215 3543 4684 4514 Breeding pond 4 549 945 1336 1548 2651 3154 4265 5341 Breeding pond 5 496 954 1454 1648 2211 3298 4800 5548 Breeding pond 6 551 868 1348 1646 2341 3531 4334 5641 Breeding pond 7 452 1065 1565 1743 2278 3564 5617 4868 Breeding pond 8 586 1095 1864 2134 2616 3405 3848 4215

[0109] According to the test, the maximum unit volume density of Daphnia sulawesiensis after 14 days of cultivation according to the cultivation method of this embodiment is about 5641ind / L.

[0110] Example 5

[0111] This embodiment provides a large-scale cultivation method of Daphnia sulawesiensis, which is basically the same as that of Example 1 except for the feed formula, and the feed formula is: 75% concentrated Nannochloropsis sulawesiensis and 25% marine photosynthetic bacteria.

[0112] The bottom was drained and the water was changed regularly every day (same as in Example 1).

[0113] In this embodiment, a total of 8 culture ponds were built for culture. During the culture process, the unit volume density of Daphnia sulwesiensis in the culture ponds was tested on days 0, 2, 4, 6, 8, 10, 12 and 14.

[0114] The detection method is: increase the inflation volume 5 minutes in advance to make it evenly distributed in the water, use a beaker to quickly pour 1 liter of water into a measuring cylinder, measure 1000ml of water, and then pour it into a new measuring cylinder to fix it with formalin in equal proportion, let it settle for 30 minutes to wait for it to die, and suck 1ml of plankton from the bottom and count it in the technical counting box.

[0115] Fill in the test results in Table 5.

[0116] Table 5

[0117]

[0118]

[0119] According to the test, the maximum unit volume density of Daphnia sulawesiensis after 14 days of cultivation according to the cultivation method of this embodiment is about 4432 ind / L.

[0120] Example 6

[0121] This embodiment provides a large-scale cultivation method of Daphnia sulawesiensis, which is basically the same as that of Example 1 except for the feed formula, and the feed formula is: 70% concentrated Nannochloropsis sulawesiensis, 25% Isochrysis zhanjiangensis, and 5% Schizochytrium sphaeroides powder.

[0122] The bottom was drained and the water was changed regularly every day (same as in Example 1).

[0123] In this embodiment, a total of 8 culture ponds were built for culture. During the culture process, the unit volume density of Daphnia sulwesiensis in the culture ponds was tested on days 0, 2, 4, 6, 8, 10, 12 and 14.

[0124] The detection method is: increase the inflation volume 5 minutes in advance to make it evenly distributed in the water, use a beaker to quickly pour 1 liter of water into a measuring cylinder, measure 1000ml of water, and then pour it into a new measuring cylinder to fix it with formalin in equal proportion, let it settle for 30 minutes to wait for it to die, and suck 1ml of plankton from the bottom and count it in the technical counting box.

[0125] Fill in the test results in Table 6.

[0126] Table 6

[0127]

[0128] According to the test, the maximum unit volume density of Daphnia sulawesiensis after 14 days of cultivation according to the cultivation method of this embodiment is about 3333 ind / L.

[0129] Comparative Example 1

[0130] This comparative example provides a large-scale culture method for Daphnia sulawesiensis. Compared with Example 1, this comparative example does not install a pure oxygen device for oxygen supply, and only micro-aeration culture is performed, and other aspects are the same.

[0131] During the breeding process, we conducted a concentration gradient experiment on dissolved oxygen, and the results are shown in Table 7.

[0132] Table 7

[0133]

[0134] At the same time, the unit volume density of Daphnia sulwesi in the culture pond was tested on days 0, 2, 4, 6, 8, 10, 12 and 14 during the culture process.

[0135] The detection method is: increase the inflation volume 5 minutes in advance to make it evenly distributed in the water, use a beaker to quickly pour 1 liter of water into a measuring cylinder, measure 1000ml of water, and then pour it into a new measuring cylinder to fix it with formalin in equal proportion, let it settle for 30 minutes to wait for it to die, and suck 1ml of plankton from the bottom and count it in the technical counting box.

[0136] Fill in the test results in Table 8.

[0137] Table 8

[0138]

[0139] The results showed that if pure oxygen is not added, the dissolved oxygen content in the water will drop sharply as the feed concentration increases and the insect density grows. Although the Sulawesi Xiusoda is tolerant to low oxygen, it is not conducive to production. The unit volume density of the Sulawesi Xiusoda obtained by farming has dropped significantly.

[0140] Comparative Example 2

[0141] This comparative example provides a large-scale cultivation method of Daphnia sulawesiensis, which is basically the same as Example 1 except for the feed formula, and the feed formula is: 100% Chlorella.

[0142] The bottom was drained and the water was changed regularly every day (same as in Example 1).

[0143] In this embodiment, a total of 8 culture ponds were built for culture. During the culture process, the unit volume density of Daphnia sulwesiensis in the culture ponds was tested on days 0, 2, 4, 6, 8, 10, 12 and 14.

[0144] The detection method is: increase the inflation volume 5 minutes in advance to make it evenly distributed in the water, use a beaker to quickly pour 1 liter of water into a measuring cylinder, measure 1000ml of water, and then pour it into a new measuring cylinder to fix it with formalin in equal proportion, let it settle for 30 minutes to wait for it to die, and suck 1ml of plankton from the bottom and count it in the technical counting box.

[0145] Fill in the test results in Table 9.

[0146] Table 9

[0147]

[0148] When only Chlorella was fed during the culture process, the growth rate of S. sulawesiensis was slow. After 14 days of culture, the maximum unit volume density of S. sulawesiensis was about 1998ind / L.

[0149] Comparative Example 3

[0150] This comparative example provides a large-scale culture method of Daphnia sulawesiensis, which is basically the same as Example 1 except for the feed formula, and the feed formula is: 100% Chaetoceros muelleri (diatom).

[0151] The bottom was drained and the water was changed regularly every day (same as in Example 1).

[0152] In this embodiment, a total of 8 culture ponds were built for culture. During the culture process, the unit volume density of Daphnia sulwesiensis in the culture ponds was tested on days 0, 2, 4, 6, 8, 10, 12 and 14.

[0153] The detection method is: increase the inflation volume 5 minutes in advance to make it evenly distributed in the water, use a beaker to quickly pour 1 liter of water into a measuring cylinder, measure 1000ml of water, and then pour it into a new measuring cylinder to fix it with formalin in equal proportion, let it settle for 30 minutes to wait for it to die, and suck 1ml of plankton from the bottom and count it in the technical counting box.

[0154] Fill in the test results in Table 10.

[0155] Table 10

[0156]

[0157] During the breeding process, there were no abnormalities in water temperature, dissolved oxygen, pH value and other indicators. However, when fed with Chaetoceros muelleri (diatom) alone, the Sulawesi S. sulawesi Daphnia could not grow normally, and even showed negative growth. After 4 days of breeding, almost all the Sulawesi S. sulawesi Daphnia in the breeding pond died, and the unit volume density dropped to 0ind / L.

[0158] The technical features of the above-described embodiments may be arbitrarily combined. To make the description concise, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0159] The above-mentioned embodiments only express several implementation methods of the present invention, and the description thereof is relatively specific and detailed, but it cannot be understood as limiting the scope of the patent of the present invention. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the concept of the present invention, which all belong to the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention shall be subject to the attached claims.

Claims

1. A method for cultivating Daphnia sulawesiensis, characterized in that: The breeding method comprises the following steps: S1. Disinfect and clean the breeding equipment; S2. Water entry: filtering and disinfecting seawater, then mixing it with disinfected fresh water, and adding the mixed water into the aquaculture device; S3. Inoculation: Inoculate Daphnia sulawesiensis in the water of the aquaculture device at an inoculation density of 460 to 550 daphnia gracilis / L, and control the dissolved oxygen concentration in the water to be 10 mg / L to 14 mg / L; S4. Feeding: feeding feed into the breeding device; S5. Water change: Use filtered and disinfected seawater to replace the water in the aquaculture device every day.

2. The breeding method according to claim 1, characterized in that: The feed comprises at least 60% concentrated Nannochloropsis algae.

3. The breeding method according to claim 2, characterized in that: The feed comprises the following components in mass fractions: 60% to 80% of concentrated Nannochloropsis edulis, 15% to 35% of Isochrysis zhanjiangensis or marine photosynthetic bacteria or Chaetoceros muelleri.

4. The breeding method according to claim 3, characterized in that: The feed comprises the following components by mass fraction: 60% to 80% of concentrated Nannochloropsis, 15% to 35% of Isochrysis Zhanjiangensis or marine photosynthetic bacteria or Chaetoceros muelleri, and 3% to 7% of hydrolyzed fish oil or Schizochytrium powder; Preferably, the feed comprises the following components in mass fractions: 65% to 75% concentrated Nannochloropsis, 20% to 30% Isochrysis zhanjiangensis or marine photosynthetic bacteria or Chaetoceros muelleri, and 4% to 6% hydrolyzed fish oil or Schizochytrium powder.

5. The breeding method according to claim 1, characterized in that: The Daphnia sulawesiensis inoculated in step S3 is obtained by breeding through the following breeding method: (1) Sampling: ① Collect water samples from the sea and place them in a sterilized container containing ultrafiltered seawater; ② Add mixed algae to the container so that the concentration of algae in the container is 1×10 5 / L~5×10 5 / L; the mixed algae liquid contains Chlorella, Nannochloropsis, and Thalassiosira, and the mass ratio of Chlorella, Nannochloropsis, and Thalassiosira is (1-2): (1-2): (1-2); ③ an air stone is placed in the container for micro-aeration culture; ④ the water is changed every day and the mixed algae is added to allow Daphnia sulawesiensis to develop into a dominant species; (2) Screening: After Daphnia sulawesiensis develops into a dominant species, adult Daphnia gracilis are screened from it; (3) Breeding and domestication: ① Domestication with feed: the adult Daphnia sulawesi obtained by screening in step (2) is domesticated to accept concentrated Nannochloropsis sulawesi; ② Selective breeding: the Daphnia sulawesi domesticated with feed containing a large amount of Daphnia sulawesi is selected and cultivated to obtain Daphnia sulawesi for inoculation.

6. The breeding method according to claim 1, characterized in that: The salinity of the mixed water in step S2 is 20 ppt to 25 ppt; and / or, The total amount of feed fed into the aquaculture device per day is 1 to 3 mg of dry weight feed per liter of water; preferably, the feed is fed in 2 to 3 times; and / or, The volume of water replaced every day in step S5 is 15% to 25% of the total volume of water in the breeding device.

7. The breeding method according to claim 1, characterized in that: The ammonia nitrogen concentration of the water in the aquaculture device is less than 0.5 mg / L, the pH value is 7.5 to 8.5, and the transparency is greater than 50 cm; and / or, The water level of the aquaculture device is 60 cm to 80 cm; and / or, The temperature of the water in the aquaculture device is 30°C ± 3°C; and / or, In step S3, Daphnia sulawesiensis is inoculated after the circulating water of the aquaculture device has been running for 3 to 5 days.

8. Use of the aquaculture method according to any one of claims 1 to 7 in preparing fish or crustacean bait.

9. A feed for Daphnia sulawesiensis, characterized in that: The feed comprises the following components in mass fractions: 60% to 80% of concentrated Nannochloropsis edulis, 15% to 35% of Isochrysis zhanjiangensis or marine photosynthetic bacteria.

10. The feed for Daphnia sulawesiensis according to claim 9, characterized in that The feed comprises the following components by mass fraction: 60% to 80% of concentrated Nannochloropsis, 15% to 35% of Isochrysis Zhanjiangensis or marine photosynthetic bacteria or Chaetoceros muelleri, and 3% to 7% of hydrolyzed fish oil or Schizochytrium powder; Preferably, the feed comprises the following components in mass fractions: 65% to 75% concentrated Nannochloropsis, 20% to 30% Isochrysis zhanjiangensis or marine photosynthetic bacteria or Chaetoceros muelleri, and 4% to 6% hydrolyzed fish oil or Schizochytrium powder.

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