Method for promoting gonad development of hippocampus kelloggi

Abstract

The invention belongs to the technical field of aquaculture, discloses a method for promoting the gonad development of hippocampus kelloggi, and particularly discloses a feed additive which comprises an antioxidant composition and Pseudodartsia andersonii. The feed additive provided by the invention can promote gonad development, shorten testis and ovary maturation time and improve sperm motility, sperm quantity and ovum quantity for male and female sea horses which are not sexually mature; for sexual maturity male hippocampus, sperm recovery can be accelerated, sperm motility is improved, and sperm quantity is increased. By feeding a feed containing the feed additive, the egg laying amount and sperm motility of the hippocampus can be remarkably improved, and the reproduction cycle is shortened, so that the reproduction rate of the hippocampus is effectively improved, population propagation is promoted, and a solid foundation is laid for large-scale breeding of the hippocampus.

Description

Technical Field

[0001] This invention belongs to the field of aquaculture technology, specifically relating to a method for promoting gonadal development in the lined hippocampus. Background Technology

[0002] Seahorses, a collective term for fish of the genus *Hippophae* in the family Syngnathidae of the order Syngnathiformes, are a highly recognizable and unique group in the ocean. They have distinctive bodies, with heads curved like a horse's head, forming a pronounced angle with their torsos. Their long, slender tails are flexible and can be used to attach themselves to seaweed or corals. In terms of reproduction, seahorses are among the few examples of male-led brooding in the animal kingdom. The female seahorse lays her eggs in the male's brood pouch, where the male fertilizes, incubates, and raises the larvae. This unique reproductive strategy makes them a popular subject of biological research. Seahorses also have significant value in traditional medicine and aquariums. However, due to habitat degradation and overfishing, most seahorse species worldwide face threats to their survival, making artificial breeding a crucial means of conservation and utilization.

[0003] Lineated seahorse ( Hippocampus erectus As an important member of the seahorse family, the striped seahorse is not only a vital component of marine biodiversity but also an important source of medicinal resources in traditional Chinese medicine and modern biomedicine. Its dried body is believed to have effects such as warming the kidneys and strengthening yang, dispersing nodules and reducing swelling, thus holding a significant position in the pharmaceutical market. However, due to overfishing and marine environmental degradation, the wild population of striped seahorses has declined sharply. The maturity of artificial breeding techniques is not only directly related to the conservation of this species but also plays a decisive role in the sustainable development of related industries.

[0004] In artificial breeding practice, low reproductive efficiency is always the primary problem restricting the large-scale breeding of the zebra shark. This problem stems from its unique reproductive biology: unlike most animals, which are “maternal gestation”, the zebra shark has evolved a special mechanism of paternal care. After the female zebra shark injects the eggs into the male's brood pouch through the oviporus, the male completes the process of fertilization and embryonic development. This special reproductive mode leads to a three-week gestation period, which greatly reduces the reproductive turnover efficiency. More seriously, the sperm production capacity of the male zebra shark is extremely weak, and the sperm-egg ratio is only maintained at a very low level of 5:1 to 191:1. In contrast, the sperm-egg ratio of zebrafish can reach 48000:1. This huge gap directly leads to low egg fertilization rate, and many eggs produced by female zebra sharks are wasted because they cannot complete fertilization. The combined effect of long gestation period and low fertilization rate makes it difficult for the zebra shark population to achieve a quantitative breakthrough, which seriously restricts the large-scale expansion of the breeding industry. Therefore, understanding the gonadal development of the zebra shark is an important prerequisite for breaking the reproductive bottleneck. Studies have shown that the gonadal development of the zebra shark can be clearly divided into six stages, each corresponding to specific physiological changes and developmental characteristics. When the zebra shark grows to 40 days, the gonadal differences between male and female individuals can be accurately distinguished through histological sectioning, which provides an important basis for early gender identification and breeding planning. When the individual grows to about 12 cm, the zebra shark reaches sexual maturity and has the ability to reproduce. It is worth noting that the zebra shark has the biological characteristics of multiple reproduction. Under suitable environmental conditions, sexually mature individuals can mate and lay eggs multiple times a year, and a single reproduction can produce 50-200 juvenile zebra sharks. This reproductive potential shows that as long as the gonadal development can be effectively promoted, the number of eggs and the quality of sperm can be improved, and the breeding interval period can be shortened, the overall reproductive efficiency can be significantly improved, and the population expansion can be effectively guaranteed.

[0005] However, the irrationality of feed supply in existing aquaculture models has become a key factor hindering the gonadal development of the lined seahorse. Currently, lined seahorse farming relies mainly on two feed sources: natural food, such as planktonic organisms like copepods and cladocerans; and general-purpose formulated feeds for marine fish. While natural food can meet the basic nutritional needs of seahorses to some extent, it suffers from unstable supply, susceptibility to pathogens, and nutritional imbalances, making it difficult to guarantee precise nutrient supply in large-scale aquaculture. General-purpose formulated feeds for marine fish are designed based on the nutritional needs of common marine fish, completely disregarding the unique physiological structure and gonadal development of the lined seahorse. The lined seahorse has a small mouth and long snout, its feeding methods and digestive physiology differ from conventional fish, and its gonadal development has specific requirements for certain nutrients (such as polyunsaturated fatty acids). General-purpose feeds not only fail to meet these precise nutritional needs but also lead to a series of reproductive development problems, such as delayed gonadal maturation, abnormal oocyte morphology, insufficient sperm motility, and even the inability to effectively activate the hypothalamic-pituitary-gonadal axis and other reproductive endocrine pathways, severely affecting reproductive performance.

[0006] In conclusion, the large-scale development of artificial breeding of the lined seahorse urgently requires overcoming the feed bottleneck. Developing a specialized feed that meets the physiological characteristics and nutritional needs of the lined seahorse and can precisely promote its gonadal development has become a core demand in the industry. This specialized feed needs to optimize physical properties based on the feeding habits of the lined seahorse, and through the scientific formulation of key nutrients, provide sufficient material basis and physiological regulatory signals for gonadal development, thereby increasing egg production, sperm quality and fertilization rate, shortening the reproductive cycle, and ultimately achieving efficient expansion of the lined seahorse population, providing strong support for resource conservation and industrial development. Summary of the Invention

[0007] The first aspect of the present invention is to provide a feed additive.

[0008] The second aspect of the present invention aims to provide the use of the feed additive of the first aspect of the present invention in promoting hippocampal gonadal development or in the preparation of feed that promotes hippocampal gonadal development.

[0009] A third aspect of the present invention is to provide a feed.

[0010] The fourth aspect of this invention aims to provide a method for promoting hippocampal gonadal development.

[0011] To achieve the above objectives, the technical solution adopted by the present invention is as follows: In a first aspect, the present invention provides a feed additive comprising an antioxidant composition and Daphnia annua.

[0012] In some embodiments of the present invention, the antioxidant combination includes at least one of astaxanthin, lutein and vitamin E.

[0013] In some embodiments of the present invention, the feed additive includes astaxanthin, lutein, vitamin E, and Daphnia argyi.

[0014] As a preferred starter food for seahorses such as the striped seahorse and the three-spotted seahorse, the Anselm's false dart flea is extremely rich in nutrients: the protein content accounts for 45% to 60% of the dry weight and is easily absorbed; it is rich in DHA (12% to 18% of total fatty acids, which promotes nerve and gonadal development) and EPA (6% to 10%, which regulates the synthesis of reproductive hormones); it is also rich in vitamins, selenium, calcium, phosphorus and other trace elements and minerals.

[0015] In some embodiments of the present invention, the *Pseudo-Dart Water Flea* is a dried powder of *Pseudo-Dart Water Flea*.

[0016] In some embodiments of the present invention, the feed additive comprises, by weight, 30 to 6000 parts astaxanthin, 10 to 2000 parts lutein, 8 to 1500 parts vitamin E and 8 to 1500 parts Daphnia anisellata.

[0017] In some embodiments of the present invention, the feed additive comprises, by weight, 50 to 5000 parts of astaxanthin, 12 to 1200 parts of lutein, 10 to 1000 parts of vitamin E and 10 to 1000 parts of Daphnia argyrophylla.

[0018] In some embodiments of the present invention, the feed additive comprises, by weight, 400-600 parts astaxanthin, 100-150 parts lutein, 80-150 parts vitamin E and 90-120 parts Daphnia anisellata.

[0019] The feed additive provided by this invention can promote gonadal development, shorten the maturation time of testes and ovaries, and increase sperm motility, sperm count, and egg count in immature male and female seahorses. For sexually mature male seahorses, it can accelerate testicular recovery and increase sperm motility and sperm count. Feeding seahorses with this additive can significantly increase egg production and sperm motility, shorten the reproductive cycle, and thus effectively improve seahorse fertility, promote population expansion, and lay a solid foundation for large-scale seahorse breeding.

[0020] A second aspect of the present invention provides the use of the feed additive of the first aspect of the present invention in promoting hippocampal gonadal development or in the preparation of feed that promotes hippocampal gonadal development.

[0021] In some embodiments of the present invention, the seahorse includes at least one of the following: striped seahorse, spiny seahorse, three-spotted seahorse, large seahorse, crowned seahorse, Japanese seahorse, tube seahorse, Kreutz seahorse, small seahorse, high-crowned seahorse, tiger-tailed seahorse, Pacific seahorse, and short-snout seahorse; preferably striped seahorse.

[0022] A third aspect of the present invention provides a feed comprising the feed additive of the first aspect of the present invention.

[0023] In some embodiments of the present invention, the feed also includes mysid shrimp.

[0024] In some embodiments of the present invention, the mass ratio of the mysid shrimp to the feed additive is 1,000,000:1.

[0025] In some embodiments of the present invention, the mysids include thawed mysids.

[0026] A fourth aspect of the present invention provides a method for promoting hippocampal gonadal development, comprising feeding a hippocampus with a feed additive of the first aspect of the present invention or a feed of the third aspect of the present invention.

[0027] In some embodiments of the present invention, when feeding feed additives or feed, for seahorses in the developmental stage, the feed of the third aspect of the present invention is fed 1 to 3 times a day, each time at 4% to 7% of the seahorse's body weight (to promote the differentiation of gonadal primordia); for seahorses in the reproductive stage, the feed of the third aspect of the present invention is fed 2 to 4 times a day, each time at 1% to 6% of the seahorse's body weight (to enhance antioxidant and fatty acid supply).

[0028] In some embodiments of the present invention, the seahorse includes at least one of the following: striped seahorse, spiny seahorse, three-spotted seahorse, large seahorse, crowned seahorse, Japanese seahorse, tube seahorse, Kreutz seahorse, small seahorse, high-crowned seahorse, tiger-tailed seahorse, Pacific seahorse, and short-snout seahorse; preferably striped seahorse.

[0029] The beneficial effects of this invention are: The feed additive provided by this invention comprehensively enhances reproductive performance by addressing the reproductive needs of seahorses at different growth stages, offering an innovative solution to overcome the bottleneck in seahorse breeding. Specifically, for immature male and female seahorses, gonadal development is a crucial physiological process for entering the reproductive period. The feed additive provided by this invention, through a precisely formulated combination of nutrients, provides sufficient material basis and physiological regulatory signals for seahorse gonadal development. Experimental data shows that feeding immature female seahorses with feed containing this additive significantly shortens the ovarian development cycle and significantly increases the number of mature eggs in the ovary; the testis development process of immature male seahorses is simultaneously accelerated, and the rate of spermatogonial cell division is increased, reserving sufficient reproductive cell resources for subsequent sperm production. This targeted developmental promotion effect allows both male and female seahorses to reach sexual maturity earlier and enter the reproductive cycle sooner. Even after seahorses reach sexual maturity, this feed additive continues to play an important role in enhancing reproductive performance. For sexually mature male seahorses, the testis is heavily depleted during reproduction, and the long recovery period under traditional feeding methods severely affects reproductive efficiency. The special nutrients in this additive can quickly replenish the energy substances needed for testicular development, shorten the testicular recovery time, and improve sperm motility and sperm count.

[0030] In practical aquaculture applications, feeding seahorses with this feed additive resulted in a significant leap in overall reproductive performance. The spawning interval of female seahorses was significantly shortened, and the plumpness and viability of the eggs were markedly improved; the brood pouch capacity and embryo hatching capacity of male seahorses were enhanced. By shortening the reproductive cycle and improving reproductive efficiency, the seahorse fertility rate was effectively increased, promoting population expansion. This feed additive successfully solved a core problem in large-scale seahorse breeding, providing solid technical support for the sustainable development of the seahorse aquaculture industry. Detailed Implementation

[0031] The present invention will be further described in detail below through specific embodiments.

[0032] It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention.

[0033] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer shall apply. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased commercially.

[0034] In the description of this invention, "several" means one or more, "multiple" means two or more, "greater than," "less than," and "exceeding" are understood to exclude the stated number, while "above," "below," and "within" are understood to include the stated number. The use of "first" and "second" in the description is merely for distinguishing technical features and should not be construed as indicating or implying relative importance, or implicitly indicating the number of indicated technical features, or implicitly indicating the order of the indicated technical features.

[0035] In the description of this invention, the terms "one embodiment," "some embodiments," "illustrative embodiment," "example," "specific example," or "some examples," etc., refer to specific features, structures, materials, or characteristics described in connection with that embodiment or example, which are included in at least one embodiment or example of the invention. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.

[0036] The features and performance of the present invention will be further described in detail below with reference to embodiments.

[0037] Example 1 A feed that promotes gonadal development in linen hippocampus includes the following ingredients, found in mysids ( Mysis relicta Add 500 mg / kg 10% astaxanthin (Tianli Pink), 120 mg / kg lutein (Tianli Yellow), 100 mg / kg vitamin E and 100 g / kg Anseloidopterus powder (Langnuo Seedling Treasure - SPF Copepods).

[0038] The above feed is prepared as follows: Dissolve the above ingredients in sterile seawater (salinity 28-30‰) in proportion, mix with mysids, and let stand in the dark for 5 minutes to ensure the effective ingredients are absorbed, thus obtaining the feed.

[0039] Comparative Example 1 A feed that promotes gonadal development in linen hippocampus includes the following ingredients, found in mysids ( Mysis relicta Add 120 mg / kg lutein (Tianlihuang), 100 mg / kg vitamin E and 100 g / kg Anselma pseudophalloides dry powder to the product.

[0040] The above feed is prepared as follows: Dissolve the above ingredients in sterile seawater (salinity 28-30‰) in proportion, mix with mysids, and let stand in the dark for 5 minutes to ensure the effective ingredients are absorbed, thus obtaining the feed.

[0041] Comparative Example 2 A feed that promotes gonadal development in linen hippocampus includes the following ingredients, found in mysids ( Mysis relicta Mysis relicta Mysis relicta Mysis relicta Mysis relicta Mysis relictaAdd 500 mg / kg of 10% astaxanthin (Tianli Pink), 12 mg / kg of lutein (Tianli Yellow), and 100 mg / kg of vitamin E.

[0042] The above feed is prepared as follows: Dissolve the above ingredients in sterile seawater (salinity 28-30‰) in proportion, mix with mysids, and let stand in the dark for 5 minutes to ensure the effective ingredients are absorbed, thus obtaining the feed.

[0043] Comparative Example 3 Thaw the mysid shrimp without any additives, and let them stand in the dark for 5 minutes before feeding.

[0044] Example 1 Location: Zhanjiang Jinma Biotechnology Co., Ltd., water temperature 26-28℃, natural photoperiod, salinity 30‰-32‰.

[0045] (1) Six hundred healthy male striped seahorses (8-10cm in length) aged 3 months were selected and randomly divided into an experimental group (n=300), with three parallel groups of 100 seahorses each; and a control group (n=300), with three parallel groups of 100 seahorses each. The control group was fed thawed mysids according to Comparative Example 3, while the experimental group was fed according to Feeding Example 1.

[0046] (2) The experiment lasted for 60 days. For the first 30 days, the animals were fed twice a day (each feeding was 5% to 6% of their body weight), and for the next 30 days, they were fed three times a day (each feeding was 3% to 5% of their body weight). Every Wednesday and Sunday, 6 male seahorses were randomly selected for testicular histology examination, and the time to sexual maturity, sperm count, and sperm motility were recorded.

[0047] The results showed that the maturation time of male hippocampal testes (stage VI) in the experimental group was 45±4 days, while that in the control group was 56±4 days, representing a 19.6% reduction in testicular maturation time. The experimental group had 4066±568 sperm counts and 56.2%±3.5% sperm motility, while the control group had 3183±419 mature sperm counts and 36.8%±3.4% sperm motility. The experimental group showed a 27.6% increase in mature sperm count and a 52.7% increase in sperm motility.

[0048] Example 2 Location: Zhanjiang Jinma Biotechnology Co., Ltd., water temperature 26-27℃, natural photoperiod, salinity 30‰-32‰.

[0049] (1) Six hundred healthy female striped seahorses (8-10cm in length) aged 3 months were selected and randomly divided into an experimental group (n=300) and a control group (n=300), with three parallel groups of 100 each. The control group was fed according to Example 1, and the experimental group was fed according to Example 1.

[0050] (2) The experiment lasted for 60 days. For the first 30 days, the animals were fed twice a day (each feeding was 5% to 6% of their body weight), and for the next 30 days, they were fed three times a day (each feeding was 3% to 5% of their body weight). Every Wednesday and Sunday, 6 female seahorses were randomly selected for ovarian histology examination, and the time of ovarian sexual maturity and the number of eggs were recorded.

[0051] The experimental results showed that the maturation time of the female hippocampal ovary (stage VI) in the experimental group was 42±3 days, while that in the control group was 52±4 days, with the ovarian maturation time shortened by 19.2%. The number of female hippocampal eggs in the experimental group was 385±68, while that in the control group was 290±77, with the number of mature eggs increasing by 32.9%.

[0052] Example 3 Location: Zhanjiang Jinma Biotechnology Co., Ltd., water temperature 26-27℃, natural photoperiod, salinity 30‰-32‰.

[0053] (1) Sixty male striped seahorses that had just given birth at 7 months of age were randomly divided into an experimental group (n=30) and a control group (n=30), with three parallel groups of 10 each. The control group was fed according to Example 2, while the experimental group was fed according to Example 1.

[0054] (2) The experiment lasted for 60 days. For the first 30 days, the animals were fed twice a day (each feeding was 5% to 6% of their body weight), and for the next 30 days, they were fed three times a day (each feeding was 3% to 5% of their body weight). Every Wednesday and Sunday, 6 male seahorses were randomly selected for testicular histology examination, and the time to sexual maturity, sperm count, and sperm motility were recorded.

[0055] The experimental results showed that in the experimental group, the time for the male hippocampus testes to redevelop to stage VI was 15±5 minutes, the number of mature sperm was 10348±612, and the sperm motility was 63.6%±5.1%. In the control group, the time for the male hippocampus testes to redevelop to stage VI was 25±5 minutes, the number of mature sperm was 8677±893, and the sperm motility was 51.7%±6.4%. The experimental group showed a 40% shorter testicular recovery time, a 19.3% increase in sperm count, and a 23% increase in sperm motility.

[0056] The embodiments of the present invention have been described in detail above. However, the present invention is not limited to the above embodiments. Within the scope of knowledge possessed by those skilled in the art, various changes can be made without departing from the spirit of the present invention. Furthermore, the embodiments of the present invention and the features thereof can be combined with each other unless otherwise specified.

Claims

1. A feed additive comprising an antioxidant composition and Daphnia angustifolia.

2. The feed additive according to claim 1, characterized in that, The antioxidant combination includes at least one of astaxanthin, lutein, and vitamin E.

3. The feed additive according to claim 2, characterized in that, The feed additives include astaxanthin, lutein, vitamin E, and Daphnia argyi.

4. The feed additive according to claim 3, characterized in that, The feed additives, by weight, include 30-6000 parts astaxanthin, 10-2000 parts lutein, 8-1500 parts vitamin E, and 8-1500 parts Daphnia anisellata.

5. The use of the feed additive according to any one of claims 1 to 4 in promoting hippocampal gonadal development or in preparing feed that promotes hippocampal gonadal development.

6. A feed comprising the feed additive according to any one of claims 1 to 4.

7. The feed according to claim 6, characterized in that, The feed also includes mysids and brine shrimp.

8. A method for promoting hippocampal gonadal development, comprising feeding the hippocampus with the feed additive of any one of claims 1 to 4 or the feed of claim 6 or 7.

9. The method according to claim 8, characterized in that, When feeding feed additives or feed, for seahorses in the developmental stage, feed them the feed described in claim 6 or 7 1 to 3 times a day, each time at 4% to 7% of the seahorse's body weight; for seahorses in the breeding stage, feed them the feed described in claim 6 or 7 2 to 4 times a day, each time at 1% to 6% of the seahorse's body weight.

10. The method according to claim 8 or 9, characterized in that, The seahorses include at least one of the following: the striped seahorse, the spiny seahorse, the three-spotted seahorse, the large seahorse, the crowned seahorse, the Japanese seahorse, the tube seahorse, the Kree's seahorse, the small seahorse, the veiled seahorse, the tiger-tailed seahorse, the Pacific seahorse, and the short-snouted seahorse.