Cultivation method for regulating and controlling body type of rotifer

Abstract

The invention relates to the technical field of rotifer culture, and discloses a culture method for regulating and controlling the body type of rotifers. According to the method, factors such as dissolved oxygen, temperature, salinity and bait for culturing the seawater rotifers are regulated and controlled, so that the purpose of regulating and controlling the body sizes of the seawater rotifers is successfully achieved; wherein the large-scale culture condition of the rotifer has relatively low dissolved oxygen and temperature and relatively large bait, and the small-scale culture condition of the rotifer has relatively high dissolved oxygen and temperature and relatively small bait, and meanwhile, some blue-green algae can be assisted. According to the application, the environmental condition of the body type size of the seawater rotifer is stably controlled, and the bait for feeding the seawater rotifer is selectively changed, so that the purpose of selectively regulating and controlling the body type of the seawater rotifer is achieved.

Description

Technical Field

[0001] The present application relates to the technical field of rotifer cultivation, and in particular to a cultivation method for regulating the body shape of rotifers. Background Art

[0002] In existing marine fish fry farming, fry typically undergo the following developmental stages: hatching, hatching (feeding rotifers), blooming (feeding Artemia), and juvenile (feeding formulated feed). The hatching stage is the most critical: since the digestive tract of hatched fry is not yet fully developed, they require exogenous nutrients to stimulate digestive tract development. At the same time, due to insufficient egg-derived nutrients, they require energy from exogenous nutrients to sustain life. The success of this hatching stage directly determines the survival rate of the fry.

[0003] In the existing marine rotifer farming, the main farming species are Brachionus rotifers (commonly known as SS-type rotifers) and Brachionus plicatilis (commonly known as L-type rotifers). Among them, the body size of Brachionus rotifers in the gestational stage is about 120 to 220 μm, which is mainly used in the early stage of fry opening and is the first exogenous nutrient ingested by fry; the body size of Brachionus plicatilis in the gestational stage is about 240 to 320 μm, which is mainly used in the late stage of fry opening to transition to the stage of feeding brine shrimp.

[0004] The mouth size of fish fry determines the size of bait they can ingest, and an appropriate bait size is beneficial for fry development. The mouth size of fish fry during the hatching phase depends on the species. Some species with smaller mouth sizes, such as grouper, typically have mouths between 80 and 140 μm when they first hatch, making them suitable for consuming Brachionus rotifer larvae. However, these larvae are often in short supply, and weaker grouper fry often have even smaller mouths, making it more difficult for them to ingest rotifers of the appropriate size. This results in high hatching rates and low survival rates for grouper fry. Furthermore, because Artemia spp. are over 500 μm in size upon hatching, fry should only be fed Artemia when they have a larger mouth size. Brachionus plicatilis, which measures 240 to 320 μm, is too small for fry in the late hatching phase, and Brachionus plicatilis itself shrinks during the culture process. Improper bait size hinders energy accumulation in the fry, leading to decreased growth rate and reduced quality. Summary of the Invention

[0005] In view of this, the purpose of this application is to provide a culture method for regulating the body size of rotifers, so that the culture method can regulate the body size of rotifers to match the size required by fry and adapt to the bait required at different stages of the fry's opening period.

[0006] In order to solve the above technical problems / achieve the above objectives or at least partially solve the above technical problems / achieve the above objectives, the present application provides a culture method for regulating the body size of rotifers, including a rotifer miniaturization culture method and / or a rotifer enlargement culture method;

[0007] The rotifer miniaturization culture method comprises:

[0008] Providing first seawater, adjusting the salinity of the first seawater to 24‰-50‰, maintaining the dissolved oxygen content above 6 mg / L, and adjusting the temperature to 30°C-40°C;

[0009] inoculating rotifers in the first seawater for cultivation, feeding the first algae feed and maintaining a first algae density, increasing the salinity of the first seawater by no more than 1‰ and / or increasing the temperature of the first seawater by no more than 1°C daily to a desired first seawater salinity and first seawater temperature, and culturing the rotifers; wherein the salinity of the first seawater does not exceed 50‰, and the temperature does not exceed 40°C;

[0010] The large-scale culturing method of rotifers comprises:

[0011] Providing second seawater, adjusting the salinity of the second seawater to 10‰-16‰, maintaining the dissolved oxygen content at no more than 6 mg / L, and adjusting the temperature to 15°C-20°C;

[0012] The rotifers are cultured in the second seawater, fed with a second algae bait and maintained at a second algae density, and the salinity of the second seawater is reduced by no more than 1‰ and / or the temperature of the second seawater is reduced by no more than 1°C per day to the desired second seawater salinity and second seawater temperature, and culture is performed; wherein the salinity of the second seawater is not less than 10‰ and the temperature is not less than 15°C.

[0013] Optionally, the dissolved oxygen content of the first seawater is 6-16 mg / L, and the dissolved oxygen content of the second seawater is 5-6 mg / L.

[0014] Optionally, the first algae bait and the second algae bait are independently selected from one or more of Chlorella vulgaris, Chrysophyte, Chaetoceros hornwort, Pseudomonas tricornuta, and Nannochloropsis spp. Further, the first algae bait and the second algae bait are independently selected from one or more of Chlorella vulgaris and Nannochloropsis spp.

[0015] Optionally, the diameter of the first algae bait is 1-4 μm, and the diameter of the second algae bait is 6-10 μm.

[0016] Optionally, the first algae bait further comprises cyanobacteria with estrogen-like effects. Further optionally, the cyanobacteria with estrogen-like effects comprise one or more of Anabaena, Nostoc, and Microcystis. Further optionally, the algal density of the cyanobacteria with estrogen-like effects is maintained at 50×104 -100×10 4 cell / mL.

[0017] Optionally, the first algae density is 400×10 4 -1200×10 4 cell / mL, and the second algae density was 200×10 4 -400×10 4 cell / mL.

[0018] Optionally, the rotifers include Brachionus rotifer and Brachionus plicatilis rotifer.

[0019] The present application successfully achieves the purpose of regulating the size of marine rotifers by regulating factors such as dissolved oxygen, temperature, salinity, and bait in the cultivation of marine rotifers. The conditions for large-scale rotifer cultivation include relatively low dissolved oxygen, temperature, and larger bait, while the conditions for small-scale rotifer cultivation include relatively high dissolved oxygen, temperature, and smaller bait, and can also assist with the cultivation of certain cyanobacteria. The present application stably controls the environmental conditions for the size of marine rotifers and selectively changes the bait used to feed the marine rotifers, achieving the purpose of selectively regulating the size of marine rotifers. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 Shown is a microscopic image of small rotifers cultivated using Nannochloropsis in Example 2;

[0021] Figure 2 Shown is a microscopic image of small rotifers cultivated using Nannochloropsis and Anabaena in Example 3;

[0022] Figure 3 Shown is a microscopic image of large rotifers cultivated using Nannochloropsis sp. in Example 4. DETAILED DESCRIPTION

[0023] The present application discloses a culture method for regulating the body size of rotifers. Those skilled in the art can refer to the content of this article and appropriately improve the process parameters to achieve it. It is particularly important to point out that all similar replacements and modifications are obvious to those skilled in the art, and they are all deemed to be included in this application. The products, processes and applications described in this application have been described through preferred embodiments. Relevant personnel can obviously modify or appropriately change and combine the methods described herein without departing from the content, spirit and scope of this application to implement and apply the technology of this application. Obviously, the described embodiments are part of the embodiments of this application, not all of the embodiments. Based on the embodiments in this application, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of this application.

[0024] It should be noted that, in this document, if relational terms such as "first" and "second", "step 1" and "step 2", and "(1)" and "(2)" appear, they are only used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply that there is any such actual relationship or order between these entities or operations. Moreover, the terms "include", "comprise" or any other variants thereof are intended to cover non-exclusive inclusion, so that a process, method, article or device comprising a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements inherent to such process, method, article or device. In the absence of further restrictions, the elements defined by the sentence "including a..." do not exclude the presence of other identical elements in the process, method, article or device comprising the elements. At the same time, the embodiments in this application and the features in the embodiments can be combined with each other in the absence of conflict.

[0025] The present application provides a cultivation method for regulating the body size of marine rotifers, aiming to solve two major problems currently faced in this field: one is to solve the problem that in the cultivation process of some marine fish fry, due to the characteristics of the fish species themselves, the fry's mouth is too small and the rotifer's body size is too large at the opening stage after the fry is hatched, resulting in difficulty in the fry's opening, resulting in the death of the fry and low breeding success rate; the other is to solve the problem that the size of medium and large rotifers becomes smaller during the breeding process, which does not match the size required by the fry, resulting in a decrease in the growth rate of the fry and a decrease in quality.

[0026] Previous investigations and studies conducted by the present applicant have shown that the size of marine rotifers is affected by dissolved oxygen, temperature, bait type, and bait size: higher dissolved oxygen, higher temperature, smaller bait size, and certain cyanobacteria can reduce the size of rotifers. The present applicant believes that this may be because higher dissolved oxygen and smaller bait reduce the accumulation of effective energy in rotifers, thereby forcing rotifers to become smaller, and certain cyanobacteria (such as Anabaena) may contain certain regulatory substances, thereby reducing the size of rotifers. Lower dissolved oxygen, lower temperature, and larger bait size can increase the size of rotifers. The present applicant believes that this may be because, compared with the reduction in size of rotifers under high dissolved oxygen, lower dissolved oxygen may cause less oxidative stress on rotifers, resulting in more energy being stored rather than oxidized, thereby increasing the size of rotifers. A larger body size will reduce the relative surface area, and a smaller relative surface area can effectively reduce heat dissipation, so rotifers tend to increase in size under low temperature environments. A larger bait suitable for the caliber of rotifers may reduce the energy consumed when rotifers ingest food, thereby storing more energy for body growth.

[0027] In the actual seawater rotifer cultivation process, smaller rotifers are usually miniaturized, such as the miniaturization of round Brachionus rotifers, and larger rotifers are usually enlarged, such as the enlargement of plicatilis Brachionus rotifers, so as to improve the cultivation efficiency. In certain embodiments of the present application, the initial density of the inoculated round Brachionus rotundiformis is 0.1 to 500 int / mL; the initial density of the plicatilis Brachionus rotundiformis is 0.1 to 200 int / mL; using the cultivation method of the present application, the size of the round Brachionus rotifer can be stably reduced from the conventional 120-220 μm to 60-120 μm, or stably increased to about 260 μm; the size of the plicatilis Brachionus rotifer can be stably increased from the conventional 240-320 μm to 280 to 400 μm, or stably reduced to about 160 μm.

[0028] The culture method of the present application includes a rotifer miniaturization culture method and / or a rotifer large-scale culture method, and the two culture methods can be carried out separately or simultaneously according to actual needs;

[0029] In a first aspect, the method for miniaturizing rotifer cultivation comprises:

[0030] The rotifer miniaturization culture method comprises:

[0031] Providing first seawater, adjusting the salinity of the first seawater to 24‰-50‰, maintaining the dissolved oxygen content above 6 mg / L, and adjusting the temperature to 30°C-40°C;

[0032] inoculating rotifers in the first seawater for cultivation, feeding the first algae feed and maintaining a first algae density, increasing the salinity of the first seawater by no more than 1‰ and / or increasing the temperature of the first seawater by no more than 1°C daily to a desired first seawater salinity and first seawater temperature, and culturing the rotifers; wherein the salinity of the first seawater does not exceed 50‰, and the temperature does not exceed 40°C;

[0033] During the rotifer miniaturization process, the primary seawater salinity and temperature can be increased simultaneously daily to rapidly reduce the rotifer's size. Daily changes exceeding 1‰ of salinity and 1°C will cause these changes to occur too rapidly, affecting the rotifer's health, hindering its growth, and even causing its death, leading to acclimation failure. Increasing the parameters of either factor alone can also achieve miniaturization, but the process will take longer. The impact of temperature increases is greater than that of salinity increases. Salinity exceeding 50‰ and temperature exceeding 40°C are detrimental to rotifer growth.

[0034] In certain embodiments of the present application, the first seawater salinity can be adjusted to any one of 24‰, 25‰, 26‰, 27‰, 28‰, 29‰, 30‰, 31‰, 32‰, 33‰, 34‰, 35‰, 36‰, 37‰, 38‰, 39‰, 40‰, 41‰, 42‰, 43‰, 44‰, 45‰, 46‰, 47‰, 48‰, 49‰, 50‰, or any value therebetween. The first seawater temperature can be adjusted to any one of 30°C, 31°C, 32°C, 33°C, 34°C, 35°C, 36°C, 37°C, 38°C, 39°C, or 40°C, or any value therebetween.

[0035] In certain embodiments of the present application, the dissolved oxygen content of the first seawater can be regulated by using nanostones or introducing pure oxygen. Higher dissolved oxygen will promote further miniaturization of rotifers, but dissolved oxygen exceeding 16 mg / L begins to inhibit the growth of rotifers and affect the final rotifer yield. Therefore, the present application controls the dissolved oxygen content of the first seawater to 6-16 mg / L, for example, any one of 6 mg / L, 7 mg / L, 8 mg / L, 9 mg / L, 10 mg / L, 11 mg / L, 12 mg / L, 13 mg / L, 14 mg / L, 15 mg / L, 16 mg / L, or any point value therebetween.

[0036] In certain embodiments of the present application, the first algae bait can be selected from one or more of Chlorella vulgaris, Chrysophyte, Chaetoceros hornwort, Pseudomonas tricornutum, and Nannochloropsis spp. In other embodiments of the present application, normal culture conditions (without controlling temperature, salinity, or dissolved oxygen) are used, and only the growth of rotifers is considered, with the final density of rotifers under the same culture conditions being used to reflect the nutritional differences between the various algae. The results showed that Nannochloropsis spp. and Chlorella vulgaris have higher nutritional value for rotifers and can be used as preferred microalgae baits for rotifer cultivation. Therefore, the first algae bait is one or more of Chlorella vulgaris and Nannochloropsis spp.

[0037] In certain embodiments of the present application, the diameter of the first algae bait is 1-4 μm, for example, any one of 1 μm, 2 μm, 3 μm, and 4 μm, or any value therebetween. A first bait diameter that is too low is not conducive to rotifer feeding, resulting in a low bait conversion rate; a first bait diameter that is too high can prevent rotifers from ingesting the bait, leading to rotifer death.

[0038] In certain embodiments of the present application, when feeding the first algae bait, the initial algae density is controlled at 200×10 4 -800×10 4 cell / mL, and then maintained at 400×10 4 -1200×10 4cell / mL, which is the initial algal density to be maintained. Algal densities below the initial and maintenance levels are not conducive to rotifer growth, resulting in slow growth. Algal densities above the initial and maintenance levels will inhibit rotifer growth and may even cause death.

[0039] In certain embodiments of the present application, the first algae bait further comprises cyanobacteria with estrogen-like effects. The cyanobacteria with estrogen-like effects can shorten the rotifer egg-holding period, reduce the diameter of rotifer eggs, and thereby reduce the size of the new generation of rotifers.

[0040] In certain embodiments of the present application, the cyanobacteria with estrogen-like effects include one or more of Anabaena, Nostoc, and Microcystis. In other embodiments of the present application, using Chlorella vulgaris as microalgae feed, under normal culture conditions (without controlling temperature, salinity, or dissolved oxygen), the effects of various cyanobacteria on the growth and size of rotifers were investigated. The effect of each alga on improving rotifer size was measured by the final size and density of rotifers under the same culture conditions. The results showed that Anabaena was the optimal alga for improving rotifer size (taking into account both the final rotifer density and the control of body size).

[0041] In certain embodiments of the present application, the initial algal density and the maintenance algal density of the cyanobacteria with estrogen-like effect are both controlled at 50×10 4 -100×10 4 cell / mL. A density lower than this is not conducive to the reduction of rotifer body size, while a density higher than this inhibits the growth of rotifer population and may lead to an increase in the number of sexual generations of rotifers.

[0042] In a second aspect, the large-scale culture method of rotifers comprises:

[0043] The large-scale culturing method of rotifers comprises:

[0044] Providing second seawater, adjusting the salinity of the second seawater to 10‰-16‰, maintaining the dissolved oxygen content at no more than 6 mg / L, and adjusting the temperature to 15°C-20°C;

[0045] The rotifers are cultured in the second seawater, fed with a second algae bait and maintained at a second algae density, and the salinity of the second seawater is reduced by no more than 1‰ and / or the temperature of the second seawater is reduced by no more than 1°C per day to the desired second seawater salinity and second seawater temperature, and culture is performed; wherein the salinity of the second seawater is not less than 10‰ and the temperature is not less than 15°C.

[0046] During the rotifer enlargement process, the salinity and temperature of the secondary seawater can be reduced simultaneously daily to rapidly increase the size of the rotifers. Daily changes exceeding 1‰ of salinity and 1°C will cause the salinity and temperature to change too rapidly, affecting the health of the rotifers, hindering their growth, and even causing their death, leading to acclimation failure. Reducing the parameters of only one of these influencing factors can also achieve the goal of enlargement, but it will take longer, and the impact of lowering temperature is greater than that of lowering salinity. Salins between 0-10‰ are detrimental to rotifer growth, especially at salinities below 5‰, which can kill rotifers within a few days. Therefore, the salinity of the secondary seawater should not be lower than 10‰. At temperatures below 15°C, rotifers still tend to increase in size, but the population does not grow, meaning that the density of the rotifers no longer increases.

[0047] In certain embodiments of the present application, the second seawater salinity can be adjusted to any one of 10‰, 11‰, 12‰, 13‰, 14‰, 15‰, 16‰, or any value therebetween. The second seawater temperature can be adjusted to any one of 15°C, 16°C, 17°C, 18°C, 19°C, or 20°C, or any value therebetween.

[0048] In certain embodiments of the present application, the dissolved oxygen content of the second seawater can be regulated by using nanostones. Lower dissolved oxygen will promote further growth of rotifers. Introducing pure oxygen will cause the dissolved oxygen to exceed 6 mg / L, which will cause the rotifers to shrink in size, which is not conducive to the growth of rotifers. Dissolved oxygen below 5 mg / L begins to inhibit the growth of rotifers. Therefore, the present application controls the dissolved oxygen content of the second seawater at 5-6 mg / L.

[0049] In certain embodiments of the present application, the second algae bait may also be selected from one or more of Chlorella vulgaris, Chrysophyte, Chaetoceros, Tripterocarpus tricornutus, and Nannochloropsis. Based on the results of the effects of the aforementioned algae baits on the final density of rotifers, the second algae bait may also be one or more of Chlorella vulgaris and Nannochloropsis.

[0050] In certain embodiments of the present application, the diameter of the second algae bait is 6-10 μm, for example, any one of 6 μm, 7 μm, 8 μm, 9 μm, 10 μm, or any value therebetween. A second bait diameter that is too low will cause the rotifers to shrink, hindering their growth. A second bait diameter that is too high will prevent the rotifers from ingesting the bait, leading to their death.

[0051] In certain embodiments of the present application, when feeding the second algae bait, the initial algae density is controlled at 100×10 4 -300×10 4 cell / mL, and then maintained at 200×104 -400×10 4 cell / mL, which is the second maintained algal density. Algal densities below the initial and maintained values ​​are not conducive to rotifer growth, resulting in slow growth. Algal densities above the initial and maintained values ​​will inhibit rotifer growth and even cause death.

[0052] In the study of the enlargement and miniaturization of rotifers in this application, the difference caused by the change in rotifer body size was not significant when the seawater salinity was 16‰ < 24‰; and the body size tended to miniaturize as the temperature increased from 20℃ < 30℃, but the trend was slow.

[0053] In the comparative experiments provided in this application, unless otherwise specified, all experimental conditions and materials, except for the differences noted in each group, were kept consistent to ensure comparability. In addition, all materials used in this application can be purchased from commercial sources.

[0054] The following further describes a culture method for regulating the body size of rotifers provided in this application.

[0055] Example 1: Algae bait screening

[0056] (1) Screening of bait microalgae

[0057] Chlorella vulgaris, Chrysophyte, Chaetoceros, Tricornutum, and Nannochloropsis were selected as rotifer nutrition sources. Culture conditions were normal (no temperature, salinity, or dissolved oxygen control, using natural seawater culture). The focus was on rotifer growth, with the final rotifer density under the same culture conditions reflecting the nutritional differences among the algae. The results are shown in Table 1 below.

[0058] Table 1

[0059]

[0060]

[0061] According to the results in Table 1, Nannochloropsis and Chlorella vulgaris have higher nutritional value for rotifers and can be used as bait microalgae for rotifer cultivation.

[0062] (2) Screening of improved algae

[0063] Anabaena, Spirulina, Nostoc, and Microcystis, all of which have estrogen-like effects, were selected as rotifer size-modifying algae. Using Chlorella vulgaris as bait, the effects of the size-modifying algae on rotifer growth and size were investigated under normal culture conditions (no manipulation of temperature, salinity, or dissolved oxygen, using natural seawater culture). The effects of each alga on rotifer size modification were measured using the final size and density of the rotifers under the same culture conditions. The results are shown in Table 2 below.

[0064] Table 2

[0065]

[0066] According to the results in Table 2, although Microcystis can reduce the size of rotifers to a lower size of 116 μm among the four algae, the final density of rotifers is significantly lower than that of the other three algae, so it is not given priority consideration; Spirulina cannot miniaturize rotifers, so it is not considered; both Nostoc and Anabaena can reduce the size of rotifers and maintain a certain density, among which Anabaena is the most suitable alga for improving the size of rotifers.

[0067] Example 2: Cultivation of small rotifers using Nannochloropsis

[0068] Take a portion of natural seawater with a salinity of 30‰ and place it in a 300L tendon water storage bucket (76cm top, 65cm bottom, and 85cm height). Maintain the water level at approximately 60cm, which is the ideal salinity. Without adjusting the water level, inoculate 120-220μm round-shaped rotifers (Brachionus rotundiformis) to a density of 122 int / mL. Maintain the water level at approximately 60cm throughout the entire process, increasing the salinity by 1‰ daily until it reaches 40‰, where it is maintained.

[0069] Use a nano air stone to introduce air, adjust the air output to a slightly boiling state, and intermittently introduce pure oxygen to maintain the dissolved oxygen content in the water above 8mg / L;

[0070] Inoculate Nannochloropsis sp. with a diameter of 1-4 μm and mix well to make the algae density reach 360×10 4 cell / mL, and in the subsequent cultivation process, the algal liquid density was maintained at 400×10 4 -1200×10 4 cell / mL;

[0071] Add a heating rod and maintain the water temperature at 30°C. Then increase it by 1°C every day until the water temperature reaches 38°C. Maintain the water temperature at 38°C for 28 days.

[0072] Since the body size control period is too long, the water needs to be changed every 7 days. The specific method of changing the water is: put the new water into a large new bucket of the same size, adjust the temperature to the corresponding temperature of the original breeding bucket, stop the gas in the original breeding bucket for 2 hours, remove the live rotifers on the upper layer, and add them to the new bucket. The rest of the steps are carried out normally.

[0073] Rotifer size measurement method: Take 1 mL of rotifers and place them in a 2 mL centrifuge tube. Add 30-50 μL of Luger's reagent to fix the rotifers. Wait 10 minutes for the rotifers to settle. Place the rotifers under an Osparin microscope. Count the rotifers whose length is greater than or equal to the length of the smallest rotifer with eggs in the group as adults. Measure and record the length of adult rotifers to determine the size range of rotifers. Figure 1 Finally, the adult round-bodied Brachionus rotifer with a body size of 75-108 μm was obtained.

[0074] Example 3: Cultivation of small rotifers using Nannochloropsis and Anabaena

[0075] Take a portion of natural seawater with a salinity of 30‰ and place it in a 300L tendon water storage bucket (76cm top, 65cm bottom, and 85cm height). Maintain the water level at approximately 60cm, which is the ideal salinity. Without adjusting the water level, inoculate 120-220μm round-shaped rotifers (Brachionus rotundiformis) to a density of 114 int / ml. Maintain the water level at approximately 60cm throughout the entire process, increasing the salinity by 1‰ daily until it reaches 40‰, where it is maintained.

[0076] Use a nano air stone to introduce air, adjust the air output to a slightly boiling state, and intermittently introduce pure oxygen to maintain the dissolved oxygen content in the water above 8mg / L;

[0077] Inoculate Nannochloropsis sp. with a diameter of 1-4 μm and mix well to make the algae density reach 420×10 4 cell / mL, and in the subsequent cultivation process, the density of the Nannochloropsis algae liquid was maintained at 400*10 4 ~1200*10 4 cell / mL, and 73×10 4 cell / mL of Anabaena sp., and maintain the density of Anabaena liquid at 50×10 4 -100×10 4 cell / mL;

[0078] Add a heating rod and maintain the water temperature at 30°C. Then increase the temperature by 1°C every day until the water temperature reaches 38°C. Maintain the water temperature at 38°C for 28 days.

[0079] Since the body size control period is too long, the water needs to be changed every 7 days. The specific method of changing the water is: put the new water into a large new bucket of the same size, adjust the temperature to the corresponding temperature of the original breeding bucket, stop the gas in the original breeding bucket for 2 hours, remove the live rotifers on the upper layer, and add them to the new bucket. The rest of the steps are carried out normally.

[0080] Rotifer size measurement method: Take 1 mL of rotifers and place them in a 2 mL centrifuge tube. Add 30-50 μL of Luger's reagent to fix the rotifers. Wait 10 minutes for the rotifers to settle. Place the rotifers under an Osparin microscope. Count the rotifers whose length is greater than or equal to the length of the smallest rotifer with eggs in the group as adults. Measure and record the length of adult rotifers to determine the size range of rotifers. Figure 2 Finally, the adult round-bodied Brachionus rotifer with a body size of 65-98 μm was obtained.

[0081] Example 3: Cultivation of large rotifers using Nannochloropsis

[0082] Take some natural seawater with a salinity of 30‰ and place it in a 5L beaker. Adjust the salinity to 16‰ with fresh water, adjust the water level to about 20cm, and inoculate 240-320μm Brachionus plicatilis rotifers to a density of 56int / ml. Then, maintain the water level at about 20cm and reduce the salinity by 1‰ per day until the salinity drops to 10‰, whereupon the salinity is maintained at 10‰.

[0083] Use a nano air stone to introduce air, adjust the air output to a slightly boiling state, and maintain the dissolved oxygen content in the water at 5-6 mg / L;

[0084] Inoculate Nannochloropsis sp. with a diameter of 6-10 μm and mix well to make the algae density reach 280×10 4 cell / mL, and in the subsequent cultivation process, the algal liquid density was maintained at 200×10 4 -400×10 4 cell / mL;

[0085] Place it in a constant temperature box and adjust the water temperature to around 20°C. Then lower it by 1°C every day until it reaches 15°C and maintain it for 18 days.

[0086] Since the body size control period is too long, the water needs to be changed every 7 days. The specific method of changing the water is: put the new water into a first-class large beaker, adjust the temperature to the corresponding temperature of the original beaker, stop the gas in the original beaker for 2 hours, remove the live rotifers on the upper layer, and put them into a new bucket. The rest of the steps are carried out normally.

[0087] Rotifer size measurement method: Take 1 mL of rotifers and place them in a 2 mL centrifuge tube. Add 30-50 μL of Luger's reagent to fix the rotifers. Wait 10 minutes for the rotifers to settle. Place the rotifers under an Osparin microscope. Count the rotifers whose length is greater than or equal to the length of the smallest rotifer with eggs in the group as adults. Measure and record the length of adult rotifers to determine the size range of rotifers. Figure 3 , and finally obtained adult rotifers with a body size of 300-390μm.

[0088] Example 4: Effects of different seawater salinity, temperature, and dissolved oxygen on rotifer body size

[0089] (1) Miniaturization of rotifers

[0090] Brachionus rotifers (120-220 μm) were used as the acclimation object, and natural seawater (initial seawater salinity 20‰, temperature 22-24°C) was selected. The culture was carried out according to the method of Example 2, and the final temperature, salinity, dissolved oxygen, and culture time parameters were referred to in Table 3 below. The average body size of the cultured rotifers is shown in Table 3;

[0091] Table 3

[0092]

[0093] As can be seen from Table 3, the higher the temperature, salinity and dissolved oxygen, the smaller the rotifer size. The length of the culture time is usually determined by the temperature, salinity and dissolved oxygen, usually 15-30 days.

[0094] (2) Rotifers grow larger

[0095] Brachionus plicatilis (240-320 μm) was used as the acclimation object, and natural seawater (initial seawater salinity 20‰, temperature 22-24°C) was selected for cultivation according to the method of Example 4. The final temperature, salinity, dissolved oxygen, and cultivation time parameters were referred to in Table 4 below. The average body size of the cultured rotifers is shown in Table 4.

[0096] Table 4

[0097]

[0098]

[0099] According to Table 4, it can be seen that the lower the temperature, salinity and dissolved oxygen, the larger the rotifer body size. The length of the culture time is usually determined by the temperature, salinity and dissolved oxygen levels, usually 10-30 days.

[0100] The foregoing is merely a list of specific embodiments of the present application, intended to enable those skilled in the art to understand or implement the present application. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the present application. Therefore, the present application is not limited to the embodiments shown herein, but is intended to conform to the broadest scope consistent with the principles and novel features of the present application.

Claims

1. A method for regulating the body shape of rotifers, characterized in that: The invention comprises a method for miniaturizing rotifer culture and / or a method for enlarging rotifer culture; The rotifer miniaturization culture method comprises: Providing first seawater, adjusting the salinity of the first seawater to 24‰-50‰, maintaining the dissolved oxygen content above 6 mg / L, and adjusting the temperature to 30°C-40°C; inoculating rotifers in the first seawater for cultivation, feeding the first algae feed and maintaining a first algae density, increasing the salinity of the first seawater by no more than 1‰ and / or increasing the temperature of the first seawater by no more than 1°C daily to a desired first seawater salinity and first seawater temperature, and culturing the rotifers; wherein the salinity of the first seawater does not exceed 50‰, and the temperature does not exceed 40°C; The large-scale culturing method of rotifers comprises: Providing second seawater, adjusting the salinity of the second seawater to 10‰-16‰, maintaining the dissolved oxygen content at no more than 6 mg / L, and adjusting the temperature to 15°C-20°C; The rotifers are cultured in the second seawater, fed with a second algae bait and maintained at a second algae density, and the salinity of the second seawater is reduced by no more than 1‰ and / or the temperature of the second seawater is reduced by no more than 1°C per day to the desired second seawater salinity and second seawater temperature, and culture is performed; wherein the salinity of the second seawater is not less than 10‰ and the temperature is not less than 15°C.

2. The culture method according to claim 1, wherein The dissolved oxygen content of the first seawater is 6-16 mg / L, and the dissolved oxygen content of the second seawater is 5-6 mg / L.

3. The culture method according to claim 1, wherein The first algae bait and the second algae bait are independently selected from one or more of Chlorella vulgaris, Chrysophyte, Chaetoceros, Pseudomonas tricornutum, and Nannochloropsis spp.

4. The culture method according to claim 3, wherein The first algae bait and the second algae bait are independently selected from one or both of Chlorella vulgaris and Nannochloropsis salsa.

5. The culture method according to claim 1, wherein The diameter of the first algae bait is 1-4 μm, and the diameter of the second algae bait is 6-10 μm.

6. The culture method according to any one of claims 1, 3, 4, and 5, characterized in that: The first algae bait also includes cyanobacteria with estrogen-like effects.

7. The culture method according to claim 6, characterized in that The cyanobacteria with estrogen-like effects include one or more of Anabaena, Nostoc, and Microcystis.

8. The culture method according to claim 6, characterized in that The algae density of the cyanobacteria with estrogen-like effect was maintained at 50×10 4 -100×10 4 cell / mL.

9. The culture method according to claim 1, wherein The first algae density is 400×10 4 -1200×10 4 cell / mL, and the second algae density was 200×10 4 -400×10 4 cell / mL.

10. The culture method according to claim 1, wherein The rotifers include Brachionus rotifer and Brachionus plicatilis rotifer.

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