Combined fungicide and application thereof in preparation of medicine for preventing and treating red bottom of babylonia lutosa in seedling raising process

Abstract

The invention discloses a combined fungicide and application thereof in preparation of a medicine for preventing and treating red bottom of babylonia lutosa in a seedling raising process. The combined fungicide comprises a spore fungicide and a bacteriophage; the spore microbial agent comprises bacillus subtilis, bacillus velezensis, bacillus licheniformis, bacillus pumilus, bacillus coagulans, bacillus megaterium and bacillus circulans; the bacteriophages comprise vibrio parahaemolyticus bacteriophage, vibrio harveyi bacteriophage, vibrio alginolyticus bacteriophage and bdellovibrio The combined fungicide can effectively restrain and solve the red bottom phenomenon appearing at the pond bottom in the babylonia lutosa seedling raising process, the babylonia lutosa seedling quantity can be remarkably increased, and the babylonia lutosa seedling raising success rate can be increased.

Description

Technical Field

[0001] The invention belongs to the technical field of snail breeding, and more specifically, relates to a combined bacterial agent and an application thereof in preparing a medicine for preventing and controlling red bottom in the process of raising seedlings of baby snails. Background Art

[0002] Dongfeng conch, also known as sea pig conch, sweet conch, southern conch, yellow conch, and flower conch, belongs to the phylum Mollusca, class Gastropoda, subclass Prosobranchia, order Neogastropoda, family Buccinidae, and genus Babylonia. It is the main cultured shellfish species in my country. my country is the world's leading country for Dongfeng conch culture. At present, the culture of Dongfeng conch in China is mainly distributed in the coastal areas south of Fujian. Among them, Hainan has become the most suitable area for the cultivation and breeding of Dongfeng conch seedlings in my country due to its unique geographical environment and climatic conditions.

[0003] However, in recent years, bacterial and fungal diseases have frequently occurred in large-scale and high-density intensive breeding of baby snails, which has seriously restricted the development of the factory breeding industry of baby snails. Especially in the middle and late stages of baby snail breeding, multiple red circles composed of harmful fungi and bacterial colonies often appear at the bottom of the breeding pond, which is called the "red bottom" phenomenon. The "red bottom" phenomenon greatly affects the survival rate and growth rate of baby snail planktonic larvae breeding; when baby snail planktonic larvae metamorphose and fall to the bottom in the late stage of the breeding process, the mortality rate exceeds 95%.

[0004] Patent Publication No. CN104531563A discloses a powdered preparation, which contains Bacillus brevis and Bdellovibrio. The powdered preparation can effectively eliminate diseases caused by Vibrio in a broad spectrum. Adding it to shrimp feed can not only inhibit the growth of pathogenic Vibrio in the shrimp intestine and reduce the occurrence of Vibrio disease in shrimp, but also promote the growth of shrimp and reduce the breeding cost. It can be widely used in aquaculture. However, this powdered preparation can only eliminate diseases caused by Vibrio to a certain extent, but it has limited improvement on the "red bottom phenomenon" that occurs during the breeding process of Babylonia snails, and this powdered preparation is suitable for the breeding process of shrimp, but not for the breeding of Babylonia snails.

[0005] Therefore, finding safer and more environmentally friendly disease prevention and control methods, providing a product that can effectively prevent the formation of red bottom in the Dongfeng snail seedling cultivation process, and improving the success rate of Dongfeng snail seedling cultivation have become technical issues that need to be urgently addressed. Summary of the invention

[0006] In view of the above-mentioned existing technical problems, the primary purpose of the present invention is to provide a combined bacterial agent, which can effectively inhibit and solve the red bottom phenomenon that occurs at the bottom of the pond during the cultivation of baby snails, thereby improving the success rate of baby snail cultivation.

[0007] The second object of the present invention is to provide a use of a combined bacterial agent in the preparation of a drug for increasing the amount of Dongfeng snail seedlings or the success rate of seedling cultivation.

[0008] The third object of the present invention is to provide a use of a combined bacterial agent in the preparation of a drug for preventing and controlling the red bottom of Babylonia snails during seedling cultivation.

[0009] The fourth object of the present invention is to provide a method for raising seedlings of Babylonia sp.

[0010] In order to achieve the above object, the present invention is implemented by the following technical solutions:

[0011] A combined bacterial agent comprises a spore agent and a phage agent; the spore agent comprises Bacillus subtilis, Bacillus velez, Bacillus licheniformis, Bacillus brevis, Bacillus coagulans, Bacillus megaterium and Bacillus circulans; the phage agent comprises Vibrio parahaemolyticus phage, Vibrio harveyi phage, Vibrio alginolyticus phage and Bdellovibrio phage.

[0012] The inventors have found through research that under the combined action of a specific combination of spore-forming bacteria, the red bottom phenomenon that occurs at the bottom of the pond during the cultivation of the snails and the problem of the larvae of the snails falling to the bottom due to metamorphosis and death can be significantly solved, with better prevention and control effects, and the success rate of the cultivation of the snails can be significantly improved. In the present invention, the spore-forming bacteria can rapidly and massively multiply and colonize in the breeding pond, effectively preventing the reproduction of pathogenic microorganisms, interfering with the infection of pathogenic microorganisms on the planktonic larvae of the snails, and destroying the colonization of pathogenic microorganisms on the snail larvae. In addition, the spore-forming bacteria can also produce bacteriolytic substances and antibacterial substances, further inhibiting the reproduction of harmful bacteria.

[0013] Furthermore, the inventors have found through research that the red bottom phenomenon that occurs at the bottom of the pond during the breeding process of baby snails can be effectively suppressed and resolved under the combined action of a specific combination of bacteriophages. The bacteriophages in the present invention can grow and reproduce in the breeding pond, can cause the lysis of pathogenic bacteria, and reduce the density of pathogenic bacteria, thereby reducing or avoiding the chance of infection or disease caused by pathogenic bacteria of baby snails, and achieving the effect of preventing and treating baby snails.

[0014] The present invention adopts the above-mentioned specific combination of spore-forming agents and phage agents to form a combined bacterial agent, which has excellent synergistic effects, can more effectively inhibit and solve the red bottom phenomenon that appears on the bottom of the pond during the cultivation of baby snails, and the problem of baby snails' planktonic larvae metamorphosing and falling to the bottom and dying; in addition, it can significantly improve the body immunity of snail seedlings, and further improve the amount of snail seedlings and the success rate of seedling cultivation. The combined bacterial agent provided by the present invention has the advantages of simple application operation, low cost, and economical and practical, and can be promoted on a large scale.

[0015] Preferably, the mass ratio of the spore-forming agent to the bacteriophore is 5 to 8: 1 to 3. More preferably, the mass ratio of the spore-forming agent to the bacteriophore is 6 to 7: 2 to 3.

[0016] Preferably, the mass ratio of Bacillus subtilis, Bacillus Velez, Bacillus licheniformis, Bacillus pumilus, Bacillus coagulans, Bacillus megaterium or Bacillus circulans is 3-4: 0.8-1.5: 3-4: 2-3: 1.5-2: 1-2: 1. More preferably, the mass ratio of Bacillus subtilis, Bacillus Velez, Bacillus licheniformis, Bacillus pumilus, Bacillus coagulans, Bacillus megaterium or Bacillus circulans is 4: 1: 4: 3: 2: 1.5: 1.

[0017] Preferably, the mass ratio of Vibrio parahaemolyticus phage, Vibrio harveyi phage, Vibrio alginolyticus phage and Bdellovibrio phage is 1 to 2: 1 to 2: 1 to 2: 3 to 5. More preferably, the mass ratio of Vibrio parahaemolyticus phage, Vibrio harveyi phage, Vibrio alginolyticus phage and Bdellovibrio phage is 1: 1: 1: 4.

[0018] Furthermore, the combined bacterial agent provided by the present invention also includes a synergist, and the synergist is a carbon source and / or a nitrogen source.

[0019] The carbon source and / or nitrogen source in the present invention can be used as a synergist and as a nutrient source for the combined bacterial agent, thereby promoting the growth and metabolism of microorganisms in the combined bacterial agent, providing the energy required for the life activities of the baby snails, providing a material basis for the normal growth and division of microbial living cells, and ultimately improving the success rate of baby snail seedling cultivation.

[0020] Preferably, the synergist comprises a carbon source and a carbon source.

[0021] Preferably, the mass ratio of the carbon source to the nitrogen source is 1-2:1-1.5.

[0022] Preferably, the mass ratio of the total amount of the spore-forming agent and the phage agent to the synergist is 4 to 14: 1. More preferably, the mass ratio is 8 to 12: 1; further preferably, the mass ratio is 10: 1.

[0023] Preferably, the carbon source includes but is not limited to non-toxic and harmless biological products produced by fermenting some sugars and agricultural wastes through bioengineering principles, and the main components of the products are small molecule organic acids or salts, alcohols and sugars.

[0024] More specifically, the carbon source is selected from one or more of bicarbonate, glucose or acetate. More specifically, the bicarbonate includes but is not limited to sodium bicarbonate, etc.; the acetate includes but is not limited to sodium acetate, etc.

[0025] Preferably, the nitrogen source is selected from one or more of ammonia water, ammonium salts, nitrates or urea. More specifically, the ammonium salts include but are not limited to ammonium sulfate, ammonium chloride and the like.

[0026] Furthermore, the present invention claims a method for preparing a combined bacterial agent, comprising the following steps:

[0027] In a sterile environment, Bacillus subtilis, Bacillus velez, Bacillus licheniformis, Bacillus pumilus, Bacillus coagulans, Bacillus megaterium and Bacillus circulans are mixed to prepare a spore-forming agent; more specifically, the temperature of the sterile environment is 25-30°C.

[0028] Under normal temperature and aseptic conditions, Vibrio parahaemolyticus phage, Vibrio harveyi phage, Vibrio alginolyticus phage and Bdellovibrio phage are mixed to prepare a phage agent;

[0029] The spore-forming agent and the bacteriophagic agent are mixed to prepare a combined bacterial agent.

[0030] In some other embodiments, the preparation of a synergist is also included: a carbon source and a nitrogen source are mixed at normal temperature and pressure to prepare the synergist.

[0031] In some other embodiments, a spore-forming agent, a bacteriophore, and a synergist are mixed to prepare a combined bacterial agent.

[0032] Furthermore, the present invention seeks to protect the use of the combined bacterial agent in the preparation of a drug for increasing the amount of Dongfeng snail seedlings or the success rate of seedling cultivation.

[0033] Specifically, in the present invention, the success rate of raising baby snails refers to: the ratio of the amount of baby snails to the total number of planktonic larvae of baby snails.

[0034] Furthermore, the present invention seeks to protect the use of the combined bacterial agent in the preparation of a drug for preventing and controlling the red bottom of the baby snail during the seedling cultivation process.

[0035] Specifically, the red bottom refers to: a red circle composed of multiple harmful fungi and / or bacterial colonies appearing at the bottom of the snail nursery pond.

[0036] Furthermore, the present invention seeks to protect a method for raising seedlings of Babylonia snails, which employs the above-mentioned combined bacterial agent to raise seedlings of Babylonia snails.

[0037] Preferably, during the seedling raising process, the water temperature is maintained at 20-30°C, the dissolved oxygen is 5-7 mg / L, and the pH is 8.2-8.5. During the seedling raising process, the total alkali in the Dongfeng snail pond is controlled to be 120-200 g / t, the ammonia nitrogen is ≤0.60 g / t, and the nitrite is ≤0.05 g / t.

[0038] In some specific embodiments, the above-mentioned combined bacterial agent can be used for preventive seedling cultivation immediately after the hatching of the Dongfeng snail egg sacs, and can also be used for therapeutic seedling cultivation of the Dongfeng snail planktonic larvae that have already shown the red bottom phenomenon.

[0039] In some embodiments, when prevention is performed, the dosage of the combined bacterial agent is 0.5 to 1.5 ppm with a cycle of 1 to 5 days. More preferably, the dosage of the combined bacterial agent is 0.6 to 1.2 ppm.

[0040] In some embodiments, when the treatment is performed, the combined bacterial agent is used once a day for 2 to 4 consecutive days, with a dosage of 3 to 15 g / m 3 More preferably, the dosage is 5 to 10 g / m 3 .

[0041] Compared with the prior art, the present invention has the following beneficial effects:

[0042] The present invention adopts a specific combination of spore-forming agents and phage agents to form a combined bacterial agent, which has excellent synergistic effects and can more effectively inhibit and solve the red bottom phenomenon that appears on the bottom of the pond during the breeding of Dongfeng snails, as well as the problem of Dongfeng snail planktonic larvae metamorphosing and falling to the bottom and dying; it can significantly improve the body immunity of snail seedlings, and further increase the amount of snail seedlings and the success rate of breeding. BRIEF DESCRIPTION OF THE DRAWINGS

[0043] Figure 1 This is a schematic diagram of a red bottom appearing in the seedling raising pond during the seedling raising process of Example 4.

[0044] Figure 2 Schematic diagram of emptying the nursery pond after successful seedling cultivation in order to solve the red bottom problem in Example 4.

[0045] Figure 3 Schematic diagram of lowering the water level of the nursery pond for comparative example 1, finding a red bottom and the death of the Babylonia snail larvae sinking to the bottom of the pond.

[0046] Figure 4 This is a schematic diagram of emptying the seedling pond with a red bottom during the seedling raising process in Comparative Example 1.

[0047] Figure 5 This is a schematic diagram of the death of the Babylonia snail larvae in comparative example 1 when the red bottom appeared in the nursery pond and no treatment was given. DETAILED DESCRIPTION

[0048] The present invention is further described below in conjunction with the accompanying drawings and specific examples, but the examples do not limit the present invention in any form. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0049] The phrase "early veliger larval stage" refers to the developmental period when two residual yolk masses are clearly visible in the body of the Babylonia snail larvae, and there are two oval veligers that are symmetrical on both sides and have densely ciliated edges.

[0050] The phrase "mid-veliger larvae" means that the yolk mass of the Babylonia snail larvae disappears, and it has distinct features such as eyes, forelegs, primary shells, and a pair of antennae.

[0051] The phrase "late veliger larvae" means that the larvae of the Dongfeng snail have a proboscis on their heads, water tubes, clearly differentiated gill filaments, and 1 to 2 rows of reddish-brown pigment spots on the whorls.

[0052] Example 1 Preparation of combined bacterial agent

[0053] (1) Under sterile environmental conditions at 25-30° C., Bacillus subtilis BS-01, Bacillus velezensis YFI-4, Bacillus licheniformis JHD-Z2, Bacillus pumilus DS-3, Bacillus coagulans NR372, Bacillus megaterium NCT-2, and Bacillus circulans L38 (all of the above strains are from the China Center for the Collection of Microbiological Cultures) are mixed in a mass ratio of 4:1:4:3:2:1.5:1 to prepare a spore-forming agent.

[0054] (2) Under sterile conditions at room temperature, Vibrio parahaemolyticus phage A3S, Vibrio harveyi phage Viha10, Vibrio alginolyticus phage HH109, ​​and Bdellovibrio phage NA7 (all of the above strains are from Shanghai Xinweilai Biotechnology Co., Ltd.) are mixed in a mass ratio of 1:1:1:4 to prepare a phage agent.

[0055] (3) At room temperature and pressure, sodium bicarbonate and ammonium chloride are mixed in a mass ratio of 2:1 to prepare a synergist.

[0056] (4) At room temperature and pressure, the spore-forming agent and the bacteriophore are mixed at a mass ratio of 7:2 under normal temperature and sterile conditions to obtain a mixture; and the mixture and the enhancer are then mixed at a mass ratio of 10:1 under normal temperature and sterile conditions to obtain a combined bacterial agent.

[0057] Example 2 Preparation of combined bacterial agents

[0058] The difference between this embodiment and embodiment 1 is that:

[0059] The mass ratio of Bacillus subtilis, Bacillus velez, Bacillus licheniformis, Bacillus brevis, Bacillus coagulans, Bacillus megaterium, and Bacillus circulans is 3:1:3:2:2:2:1;

[0060] The mass ratio of Vibrio parahaemolyticus phage, Vibrio harveyi phage, Vibrio alginolyticus phage, and Bdellovibrio phage was 2:2:1:3;

[0061] The mass ratio of spore-forming agent to bacteriostatic agent is 6:3.

[0062] Example 3 Preparation of combined bacterial agent

[0063] The difference between this embodiment and embodiment 1 is that:

[0064] The mass ratio of Bacillus subtilis, Bacillus velez, Bacillus licheniformis, Bacillus brevis, Bacillus coagulans, Bacillus megaterium, and Bacillus circulans is 4:1:3.5:2.5:2:1.5:1;

[0065] The mass ratio of Vibrio parahaemolyticus phage, Vibrio harveyi phage, Vibrio alginolyticus phage, and Bdellovibrio phage was 1.5:2:1.5:4;

[0066] The mass ratio of spore-forming agent to bacteriostatic agent is 8:2.

[0067] Example 4 Treatment test of red bottom in the process of raising baby snails

[0068] Purchase the egg capsules of Dongfeng Snail from Wenchang, Hainan. Use seawater to wash the attachments and sludge on the surface of Dongfeng Snail egg capsules, remove the whitish and reddish infected egg capsules, put them in the hatching basket, and put 4 jin of egg capsules in the nursery pond for each pond. The nursery pond is 3m×4m×1m in size, with a water depth of 0.8m. The water temperature is 28℃ when entering the pond. The pond is turned over once a day before hatching. When hatching is approaching, it is no longer moved. The hatching density is maintained at 0.1 / ml. Continuous aeration during the cultivation period, 1 / 3 of the water is changed every day during the early face plate larvae stage, 1 / 2 of the water is changed every day during the middle face plate larvae stage, and 3 / 4 of the water is changed every day during the late face plate larvae stage. In the early face plate larvae stage, all golden algae are fed, and in the middle face plate larvae stage, golden algae are mainly fed, supplemented by horn algae. In the late face plate larvae stage, golden algae, horn algae, and flat algae are mixed according to the bait situation. The feeding of golden algae is 3,000 per ml per day at the beginning, and gradually increases to 17,000 per ml per day in the later stage of the larvae; the feeding of horn algae and flat algae is based on the amount of stomach contents of the planktonic larvae of the baby snails. Feeding is increased from 2 times a day at the beginning to 4 times a day in the later stage. At the same time, spirulina powder is fed as an auxiliary, 2-3 grams / m 3 , feed 2 to 3 times a day.

[0069] On the 10th day of cultivation, the bottom of the nursery pond was covered with red circles (i.e., the "red bottom" phenomenon appeared, such as Figure 1 As shown), the combined bacterial agent prepared in Example 1 was used at 5 g / m 3 Spray the entire pond (based on the volume of the aquaculture water body) once a day for 4 consecutive days until the red circle on the bottom of the pond disappears (such as Figure 2 shown).

[0070] Example 5 Treatment test of red bottom in the process of raising baby snails

[0071] The difference between this example and example 4 is that: on the 11th day of cultivation, the nursery pool was covered with red circles, and the combined bacterial agent prepared in example 2 was used. The dosage, frequency and time of the combined bacterial agent were the same as those in example 4.

[0072] Example 6 Prevention test of red bottom in the process of raising baby snails

[0073] The difference between this embodiment and embodiment 4 is that when the planktonic larvae of Babylonia snails hatch from the egg capsules, the combined bacterial agent prepared in embodiment 1 is added once every 2 days, with 1 ppm added each time (based on the volume of the aquaculture water body) until the last day of seedling cultivation.

[0074] Example 7 Prevention test of red bottom in the process of raising baby snails

[0075] The difference between this embodiment and embodiment 4 is that the combined bacterial agent only contains spore-forming agent and bacteriophagic agent, and does not contain synergist.

[0076] Comparative Example 1

[0077] The difference between this comparative example and Example 4 is that no treatment measures are taken, that is, the combined bacterial agent prepared in Example 1 is not used.

[0078] like Figures 3 to 5 As shown in the figure, the nursery pond had a red bottom and the larvae of the baby snails died and sank to the bottom of the pond. Starting from the 11th day, the density of the floating larvae of the baby snails in the nursery pond water body gradually decreased. By the 13th day, there were almost no floating larvae of the baby snails in the nursery pond. The planktonic larvae of the baby snails died and sank to the bottom of the pond, and the nursery cultivation failed.

[0079] Comparative Example 2

[0080] The difference between this comparative example and Example 4 is that the combined bacterial agent only contains spore-forming agents.

[0081] Comparative Example 3

[0082] The difference between this comparative example and Example 4 is that the combined bacterial agent only contains bacteriophagy.

[0083] Comparative Example 4

[0084] The difference between this comparative example and Example 4 is that:

[0085] The spore-forming agent in the combined inoculant contains only Bacillus subtilis.

[0086] Comparative Example 5

[0087] The difference between this comparative example and Example 4 is that:

[0088] The phage agent in the combination agent contains only Vibrio parahaemolyticus phage.

[0089] Comparative Example 6

[0090] The difference between this comparative example and Example 4 is that:

[0091] The spore-forming agent in the combined bacterial agent only contains Bacillus pumilus; the phage agent only contains Bdellovibrio phage.

[0092] Comparative Example 7

[0093] The difference between this comparative example and Example 4 is:

[0094] The disinfectant povidone-iodine is used to replace the combined bacterial agent. The specific operation is as follows:

[0095] Add 0.75 ml of povidone-iodine as disinfectant per cubic meter of water, and observe the density of planktonic larvae of Babylonia sinensis and the red bottom phenomenon of the pond several times a day.

[0096] The Dongfeng snail seedlings were not able to adapt and refused to eat the feed. Most of the Dongfeng snail planktonic larvae sank to the bottom and died that day. After the dead seedlings were cleared away, water was added and povidone iodine was continued to be used. The area of ​​the red circle did not shrink, but continued to expand.

[0097] Comparative Example 8

[0098] The difference between this comparative example and Example 4 is:

[0099] Chlorine dioxide is used to replace the combined bacterial agent. The specific operation is as follows:

[0100] Add 1ppm of chlorine dioxide, and observe the density of planktonic larvae of Babylonia sinensis and the red bottom phenomenon of the pond several times a day.

[0101] The red circles were reduced by half on the second day and disappeared on the third day, but the floating larvae of the baby snails died and sank to the bottom quickly on the first day.

[0102] Comparative Example 9

[0103] The difference between this comparative example and Example 4 is:

[0104] Florfenicol was used to replace the combined bacterial agent. The specific operation was as follows:

[0105] Add 8×10 -6 ppm of fluoroquinol, and observe the density of planktonic larvae of the baby snails and the red bottom phenomenon of the pond several times a day.

[0106] The floating larvae of Dongfeng snails still have a poor appetite. After 3 days, there are only a few scattered Dongfeng snail larvae floating in the nursery pond. Most of the Dongfeng snail larvae have sunk to the bottom and died. The area of ​​the red circle has not decreased, but continued to expand.

[0107] Comparative Example 10

[0108] The difference between this comparative example and Example 4 is:

[0109] Hydrogen peroxide is used to replace the combined bacterial agent. The specific operation is as follows:

[0110] After adding 1ppm of hydrogen peroxide (active oxygen 10%), it was found that the area of ​​the red circle was reduced by 2 / 3 on the first day, but the density of the larvae decreased by 50% on the first day of using hydrogen peroxide. On the third day, only a few larvae were left floating, and most of the snail larvae had sunk to the bottom and died.

[0111] Test Case

[0112] After the hatching of the egg sacs, each nursery pond adopts the five-point sampling method. Water samples are taken from the four corners and the middle of the nursery pond to fill a 250ml measuring cylinder. The number of planktonic larvae of Dongfeng snails in the 250ml measuring cylinder each time is counted, and the average value is converted to obtain the average number of planktonic larvae of Dongfeng snails per 250ml. The average value is multiplied by 4 and 1000 to obtain the number of planktonic larvae of Dongfeng snails per cubic meter of water. The number is then multiplied by the volume of the nursery water body, 9.6 cubic meters, which is the total number of planktonic larvae of Dongfeng snails in the nursery pond.

[0113] Amount of snail seedlings: Take the average count of 50 grams of Dongfeng snail seedlings at random three times, divide it by 50, and you will get the specification of snail seedlings (pieces / gram). Multiply it by the weight of snail seedlings (g) to get the amount of snail seedlings (pieces).

[0114] The calculation formula is as follows:

[0115] The total number of floating larvae of Babylonia sinensis = the sum of the number of larvae in 5 250ml measuring cylinders / 5*4*1000*9.6;

[0116] Snail seedling quantity = average number of 50g Dongfeng snail seedlings in 3 times / 50*number of grams;

[0117] The success rate of seedling cultivation = the number of snail seedlings / the total number of planktonic larvae of Babylonia snails*100%;

[0118] The test data of the above embodiments and comparative examples are shown in Table 1 below.

[0119] Table 1

[0120]

[0121]

[0122] It can be seen from the above-mentioned Examples 4 to 7 that the combined bacterial agent provided by the present invention can more effectively inhibit and solve the red bottom phenomenon that appears at the bottom of the pond during the cultivation of Dongfeng snails, as well as the problem of the metamorphosis and death of Dongfeng snail planktonic larvae; it can improve the body immunity of snail seedlings and improve the success rate of Dongfeng snail seedling cultivation. In the above-mentioned embodiments, the amount of snail seedlings is ≥412560.2, the success rate of seedling cultivation is ≥48.4%, and the number of days for the red bottom to disappear is ≤5 days. Furthermore, the amount of snail seedlings is ≥663492.3, the success rate of seedling cultivation is ≥72.6%, and the number of days for the red bottom to disappear is ≤4 days.

[0123] It can be seen from Example 4, Example 7, Comparative Example 1, Comparative Example 2 and Comparative Example 3 that when a spore-forming agent and a phage agent are used in combination, a combined bacterial agent has an excellent synergistic effect, which can significantly increase the amount of snail seedlings, the success rate of seedling cultivation and reduce the number of days for the red bottom to disappear. However, it is difficult to achieve the technical effect of the present invention by using any one of the bacterial agents.

[0124] It can be seen from Example 4, Comparative Example 4, Comparative Example 5 and Comparative Example 6 that when a spore-forming agent or a bacteriovorus agent is used in combination with another bacteriovorus agent, or when Bacillus pumilus and Bdellovibrio phage are used in combination, it is difficult to achieve the technical effect of the present invention.

[0125] It can be seen from Example 4 and Comparative Examples 7 to 10 that when the disinfectant povidone iodine, chlorine dioxide, fluoroquinolone or hydrogen peroxide is used to replace the combined bacterial agent of the present invention, it is difficult to achieve the technical effect of the present invention.

[0126] The foregoing examples are merely illustrative and are used to explain some features of the method of the present invention. The appended claims are intended to require the widest possible range that can be imagined, and the embodiments presented herein are demonstrated by the applicant's actual test results. Therefore, the applicant's intention is that the appended claims are not limited by the selection of examples that illustrate the features of the present invention. Some numerical ranges used in the claims also include sub-ranges therein, and changes in these ranges should also be interpreted as being covered by the appended claims where possible.

Claims

1. A combined bacterial agent, characterized in that: Including spore-forming agents and bacteriophagic agents; The spore-forming agents include Bacillus subtilis, Bacillus velez, Bacillus licheniformis, Bacillus pumilus, Bacillus coagulans, Bacillus megaterium and Bacillus circulans; The bacteriolytic agents include Vibrio parahaemolyticus phage, Vibrio harveyi phage, Vibrio alginolyticus phage and Bdellovibrio phage.

2. The combined bacterial agent according to claim 1, characterized in that: The mass ratio of the spore agent to the bacteriophagy agent is 5-8:1-3.

3. The combined bacterial agent according to claim 1, characterized in that: The mass ratio of Bacillus subtilis, Bacillus Velez, Bacillus licheniformis, Bacillus brevis, Bacillus coagulans, Bacillus megaterium and Bacillus circulans is 3-4:0.8-1.5:3-4:2-3:1.5-2:1-2:

1.

4. The combined bacterial agent according to claim 1, characterized in that: The mass ratio of Vibrio parahaemolyticus phage, Vibrio harveyi phage, Vibrio alginolyticus phage and Bdellovibrio phage is 1-2:1-2:1-2:3-5.

5. The combined bacterial agent according to claim 1, characterized in that: The combined bacterial agent also includes a synergist; the synergist is a carbon source and / or a nitrogen source.

6. The combined bacterial agent according to claim 5, characterized in that: The carbon source is selected from one or more of bicarbonate, glucose or acetate; the nitrogen source is selected from one or more of ammonia water, ammonium salt, nitrate or urea.

7. The combined bacterial agent according to claim 5, characterized in that: The mass ratio of the total amount of spore-forming agent and bacteriophagy agent to the synergist is 4-14:

1.

8. Use of the combined bacterial agent according to any one of claims 1 to 7 in the preparation of a drug for increasing the amount of Babylonia snail seedlings or the success rate of seedling cultivation.

9. Use of the combined bacterial agent according to any one of claims 1 to 7 in the preparation of a drug for preventing and controlling red bottom in the process of raising seedlings of Babylonia snails.

10. A method for raising seedlings of Babylonia spp., characterized in that: The combined bacterial agent described in any one of claims 1 to 7 is used to cultivate seedlings of Babylonia snails.

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