Method for regulating inorganic nitrogen in water in land-based round pond tilapia breeding system by microbial preparation

Abstract

The application provides a kind of land-based round pool tilapia breeding system water inorganic nitrogen microbial preparation regulation method, comprising the following steps: step S1, take Candida utilis culture to obtain activated strain one;Step S2, activated strain one expansion culture obtains Candida utilis bacterial liquid;Step S3, take Bacillus subtilis culture to obtain activated strain two;Step S4, activated strain two expansion culture obtains Bacillus subtilis bacterial liquid;Step S5, take Rhodopseudomonas palustris culture to obtain activated strain three;Step S6, activated strain three expansion culture obtains Rhodopseudomonas palustris bacterial liquid;Step S7, Candida utilis bacterial liquid, Bacillus subtilis bacterial liquid, Rhodopseudomonas palustris bacterial liquid are mixed to obtain composite microbial preparation;Step S8, round pool is raised to tilapia fry, and granular compound feed is fed;Step S9, after 20-30 days of breeding, composite microbial preparation is splashed in round pool.The application effectively reduces the concentration of ammonia nitrogen, nitrite nitrogen and nitrate nitrogen in the round pool, and reduces the tail water treatment load.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of aquaculture, in particular to a method for regulating inorganic nitrogen in water in a land-based round pond tilapia farming system. BACKGROUND

[0002] The land-based round pond farming mode is a rapidly developing aquaculture mode in China at present, among which the land-based round pond tilapia farming is developing particularly rapidly. In this process, the perfection and development of supporting technologies including disease prevention and control, water quality regulation, tail water treatment, etc. are of great importance. Among them, the regulation of water quality in the round pond is the key to the farming effect. The inorganic nitrogen, especially ammonia nitrogen and nitrite nitrogen, has a potential impact on the growth and health of tilapia. Although daily water exchange is also an important means of water quality regulation in land-based round pond farming, the water quality regulation technology in the round pond is still an important part of water quality control. In addition to ensuring the growth and health of tilapia, it also plays an important fundamental role in water resource conservation and reducing tail water treatment load. Therefore, we propose a method for regulating microorganism preparations in a land-based round pond tilapia farming system. Through the use of composite microorganism preparations, the concentrations of ammonia nitrogen, nitrite nitrogen and nitrate nitrogen in the water of the round pond are effectively regulated, the impact on the growth and health of tilapia is reduced, and the tail water treatment load is reduced. SUMMARY

[0003] The purpose of the present application is to overcome and supplement the deficiencies in the prior art, and to provide a method for regulating microorganism preparations in water in a land-based round pond tilapia farming system. Through the preparation and use of composite microorganism preparations composed of Candida utilis, Bacillus subtilis and Rhodopseudomonas palustris, the concentrations of ammonia nitrogen, nitrite nitrogen and nitrate nitrogen in the water of the round pond are reduced, the impact on the growth and health of tilapia is reduced, and the tail water treatment load is reduced.

[0004] The technical solution adopted by the present application is as follows:

[0005] A method for regulating microorganism preparations in water in a land-based round pond tilapia farming system, comprising the following steps:

[0006] Step S1, 2-3 loops of Candida utilis preserved on the slope are taken with an inoculation loop one, and the inoculation loop one is placed in the yeast paste malt juice medium. Then the medium is placed in a shaking bed for culture for at least 12h to obtain an activated strain one;

[0007] Step S2, the activated strain one is transferred to the yeast paste malt juice medium for expansion culture for at least 12h, until the concentration of the bacterial solution is >10 9 CFU / ml to obtain a Candida utilis bacterial solution;

[0008] Step S3, take 2-3 circles of Bacillus subtilis preserved in the slant with inoculation loop, and place the inoculation loop in LB medium, then place the medium in the shaker for culture for at least 10 h to obtain activated strain two;

[0009] Step S4, transfer the activated strain two to LB medium for expansion culture for at least 10 h until the concentration of bacterial liquid is >10 9 CFU / ml to obtain Bacillus subtilis bacterial liquid;

[0010] Step S5, take 2-3 circles of Rhodopseudomonas palustris preserved in the slant with inoculation loop three, and place the inoculation loop in light and bacterial medium for static culture for at least 60 h to obtain activated strain three;

[0011] Step S6, transfer the activated strain three to light and bacterial medium for expansion culture for at least 60 h until the concentration of bacterial liquid is >10 9 CFU / ml to obtain Rhodopseudomonas palustris bacterial liquid;

[0012] Step S7, mix the Candida utilis bacterial liquid, Bacillus subtilis bacterial liquid and Rhodopseudomonas palustris bacterial liquid according to 40-45%, 40-45% and 10-20% of the total volume to obtain a composite microbial preparation;

[0013] Step S8, provide a plurality of small land-based circular pools with a volume of 1m 3 In late May, 40-50 healthy Oreochromis aureus fry with an average body length of 4-6 cm are placed in each circular pool, and during the breeding period, the fish are fed with granular compound feed, and the dissolved oxygen concentration in the water is controlled to be >4 mg / L by an oxygenator;

[0014] Step S9, after 20-30 days of breeding, the composite microbial preparation in step S7 is sprayed into the circular pool at 90-110 ml every 7-10 days until the end of breeding.

[0015] Preferably, the method for regulating the inorganic nitrogen microbial preparation in the water of the land-based circular pool tilapia breeding system, wherein: the rotation speed of the shaker in step S1 is 130-180 rpm, and the temperature is 28-32℃, the rotation speed of the shaker in step S2 is 130-180 rpm, the pH is 7-8, and the temperature is 28-32℃, and the inoculation amount of the activated strain one is 2%-3%.

[0016] Preferably, the method for regulating the inorganic nitrogen microbial preparation in the water of the land-based circular pool tilapia breeding system, wherein: the rotation speed of the shaker in step S3 is 130-180 rpm, and the temperature is 30-40℃, the rotation speed of the shaker in step S4 is 130-180 rpm, the pH is 6-8, and the temperature is 30-40℃, and the inoculation amount of the activated strain two is 2%-3%.

[0017] Preferably, the inorganic nitrogen microbial preparation regulation method of the water in the land-based round pond tilapia breeding system, wherein: the light and bacterial culture medium in step S5 and step S6 comprises 2.0 g / L of ammonium chloride, 3.0 g / L of sodium acetate, 0.5 g / L of magnesium sulfate, 0.1 g / L of calcium chloride, 0.5 g / L of potassium dihydrogen phosphate, 0.5 g / L of dipotassium hydrogen phosphate, and 1.0 g / L of yeast extract.

[0018] Preferably, the inorganic nitrogen microbial preparation regulation method of the water in the land-based round pond tilapia breeding system, wherein: the light and bacterial culture medium in step S5 has a light intensity of 3500-5500 Lx and a temperature of 30-35℃; the light intensity in step S6 is 3500-5500 Lx, the pH is 6-8, and the temperature is 30-35℃, and the inoculation amount of the activated strain three is 10-20%.

[0019] Preferably, the inorganic nitrogen microbial preparation regulation method of the water in the land-based round pond tilapia breeding system, wherein: the feeding in step S8 is specifically as follows: 3-4 times of feeding per day with a feeding amount of 1%-3% of the weight of the fish in the first 1-2 months; and then 2-3 times of feeding per day with a feeding amount of 4%-6%.

[0020] Preferably, the inorganic nitrogen microbial preparation regulation method of the water in the land-based round pond tilapia breeding system, wherein: 10-15% of water needs to be changed every day during the breeding in step S8.

[0021] Advantages of the present application:

[0022] (1) The inorganic nitrogen microbial preparation regulation method of the water in the land-based round pond tilapia breeding system can effectively reduce the concentrations of ammonia nitrogen, nitrite nitrogen, and nitrate nitrogen in the water in the round pond, reduce the influence of the three on the growth and health of tilapia, and reduce the tail water treatment load.

[0023] (2) The inorganic nitrogen microbial preparation regulation method of the land-based round pond tilapia breeding system can reduce the water changing frequency, thereby reducing the cost caused by energy consumption; and can also promote the growth of tilapia. DETAILED DESCRIPTION

[0024] The present application will be further described below in combination with specific examples.

[0025] The Candida utilis is preserved in the Guangdong Microbial Digital Culture Collection Center with a preservation number of GDMCC 2.148; the Bacillus subtilis is preserved in the Guangdong Microbial Digital Culture Collection Center with a preservation number of GDMCC 1.372; and the Rhodopseudomonas palustris is preserved in the Guangdong Microbial Digital Culture Collection Center with a preservation number of GDMCC 1.167. The above strains are all purchased from the Guangdong Microbial Digital Culture Collection Center.

[0026] Embodiment 1

[0027] A method for regulating microorganism preparation of inorganic nitrogen in water of a land-based round pond tilapia breeding system, comprising the following steps:

[0028] Step S1, taking 3 loops of Candida utilis preserved in a slant with a loop one, and placing the loop one in a yeast extract malt extract medium, and then placing the medium in a shaker for culture for 12 hours to obtain an activated strain one, the rotation speed of the shaker being 150 rpm and the temperature being 30°C;

[0029] Step S2, transferring the activated strain one to the yeast extract malt extract medium for expansion culture for 16 hours until the concentration of the bacterial liquid is >10 9 CFU / ml, obtaining 10 9 CFU / ml of Candida utilis bacterial liquid by dilution, the rotation speed of the shaker being 150 rpm, the pH being 7, the temperature being 30°C, and the inoculation amount of the activated strain one being 2%, to obtain the Candida utilis bacterial liquid;

[0030] Step S3, taking 3 loops of Bacillus subtilis preserved in a slant with a loop two, and placing the loop in an LB medium, and then placing the medium in a shaker for culture for 10 hours until the bacterial liquid is turbid with obvious bacterial bodies to obtain an activated strain two, the rotation speed of the shaker being 150 rpm and the temperature being 40°C;

[0031] Step S4, transferring the activated strain two to the LB medium for expansion culture for 12 hours until the concentration of the bacterial liquid is >10 9 CFU / ml, obtaining 10 9 CFU / ml of Bacillus subtilis bacterial liquid by dilution, the rotation speed of the shaker being 150 rpm, the pH being 7, the temperature being 40°C, and the inoculation amount of the activated strain two being 2%;

[0032] Step S5, taking 3 loops of Rhodopseudomonas palustris preserved in a slant with a loop three, and placing the loop in a light and bacterial medium for static culture for 60 hours until the bacterial liquid is obviously red to obtain an activated strain three, the light intensity being 4000 Lx and the temperature being 30°C;

[0033] The light and bacterial medium comprises ammonium chloride 2.0 g / L, sodium acetate 3.0 g / L, magnesium sulfate 0.5 g / L, calcium chloride 0.1 g / L, potassium dihydrogen phosphate 0.5 g / L, dipotassium hydrogen phosphate 0.5 g / L, and yeast extract 1.0 g / L.

[0034] Step S6, transferring the activated strain three to the light and bacterial medium for expansion culture for 65 hours until the concentration of the bacterial liquid is >10 9 CFU / ml, obtaining 10 9CFU / ml of Rhodopseudomonas palustris bacteria liquid, light intensity of 4000Lx, pH of 7, temperature of 30℃, and inoculation amount of 10% of the activated strain three;

[0035] Step S7, the Candida utilis bacteria liquid, Bacillus subtilis bacteria liquid and Rhodopseudomonas palustris bacteria liquid are mixed in a ratio of 45%, 45% and 10% of the total volume to obtain the compound microbial preparation;

[0036] Step S8, a plurality of small land-based circular pools with a volume of 1m 3 are provided, and in late May, 50 healthy Oreochromis aureus fry with an average body length of 4-6cm are raised in each circular pool, and during the breeding period, the fry are fed with granular compound feed, the feed is fed 3 times a day within the first month, and the feeding amount is 2% of the body weight of the fish; then the feed is fed 2 times a day, and the feeding amount is increased to 4%, and the dissolved oxygen concentration in the water is controlled to be greater than 4mg / L by an oxygenator, and the water needs to be replaced by 10% every day during the breeding period;

[0037] Step S9, after 20 days of breeding, the compound microbial preparation in step S7 is sprayed into the circular pool at 110ml every 7 days, and the spraying time is preferably selected in the morning on a sunny day until the end of breeding.

[0038] Effect of the compound microbial preparation in Example 1 on inorganic nitrogen in water: after 6 days of using the compound microbial preparation for the first time, the ammonia nitrogen, nitrite and nitrate nitrogen in the water in the circular pool are decreased by 37.32%, 41.15% and 39.27% respectively compared with not using the compound microbial preparation; effect on the growth of tilapia: after 60 days of using the compound microbial preparation for the first time, the average body length and average body weight of tilapia in the circular pool are increased by 12.1% and 32.1% respectively compared with not using the compound microbial preparation.

[0039] Example 2

[0040] A microbial preparation regulation method for inorganic nitrogen in water of a land-based circular pool tilapia breeding system, wherein the method comprises the following steps:

[0041] Step S1, 3 loops of Candida utilis preserved on an inclined plane are taken by a loop inoculator, and the loop inoculator is placed in a yeast extract malt extract medium, and then the medium is placed in a shaker for culture for 12h, and the bacteria liquid is turbid with obvious bacteria, to obtain an activated strain one, the rotation speed of the shaker is 150rpm, and the temperature is 30℃;

[0042] Step S2, the activated strain one is transferred to the yeast extract malt extract medium for expansion culture for 16h, until the bacteria liquid concentration is greater than 10 9 CFU / ml, and 10 9Candida utilis bacteria liquid, the rotation speed of the shaker was 150 rpm, the pH was 7, the temperature was 30 DEG C, the inoculation amount of the activated strain one was 2%, and Candida utilis bacteria liquid was obtained;

[0043] Step S3, the Bacillus subtilis 3 ring preserved in the slope was taken with the inoculation ring, and the inoculation ring was placed in the LB culture medium, and then the culture medium was placed in the shaker for culture for 10 hours, the bacteria liquid was turbid with obvious bacteria, and the activated strain two was obtained, the rotation speed of the shaker was 150 rpm, and the temperature was 40 DEG C;

[0044] Step S4, the activated strain two was transferred to the LB culture medium for expansion culture for 12 hours, and the bacteria liquid concentration was greater than 10 9 CFU / ml, 10 9 CFU / ml Bacillus subtilis bacteria liquid, the rotation speed of the shaker was 150 rpm, the pH was 7, the temperature was 40 DEG C, and the inoculation amount of the activated strain two was 2%;

[0045] Step S5, the Rhodopseudomonas palustris 3 ring preserved in the slope was taken with the inoculation ring three, and the inoculation ring was placed in the light and bacteria culture medium for static culture for 60 hours, the bacteria liquid was obviously red, and the activated strain three was obtained, the light intensity was 4000 Lx, and the temperature was 30 DEG C;

[0046] The light and bacteria culture medium comprises ammonium chloride 2.0 g / L, sodium acetate 3.0 g / L, magnesium sulfate 0.5 g / L, calcium chloride 0.1 g / L, potassium dihydrogen phosphate 0.5 g / L, dipotassium hydrogen phosphate 0.5 g / L and yeast extract 1.0 g / L.

[0047] Step S6, the activated strain three was transferred to the light and bacteria culture medium for expansion culture for 65 hours, and the bacteria liquid concentration was greater than 10 9 CFU / ml, 10 9 CFU / ml Rhodopseudomonas palustris bacteria liquid, the light intensity was 4000 Lx, the pH was 7, the temperature was 30 DEG C, and the inoculation amount of the activated strain three was 10%;

[0048] Step S7, Candida utilis bacteria liquid, Bacillus subtilis bacteria liquid and Rhodopseudomonas palustris bacteria liquid were mixed according to 40%, 40% and 20% of the total volume, and the composite microbial preparation was obtained;

[0049] Step S8, a container with a volume of 1 m 3a plurality of small land-based circular ponds, 50 healthy Oreochromis aureus fry with an average body length of 4-6 cm will be raised in each circular pond in late May, and granular compound feed will be used for feeding during the breeding period. The initial feeding is 3 times a day for 1 month, and the feeding amount is 2% of the fish body weight; then the feeding is 2 times a day, and the feeding amount is increased to 4%, and the dissolved oxygen concentration in the water is controlled to be >4 mg / L by an oxygenator. The water needs to be changed by 10% every day during the breeding period;

[0050] Step S9, 20 days after the start of breeding, 110 ml of the compound microbial preparation in step S7 is sprayed into the circular pond every 7 days, and the spraying time is preferably selected in the morning on sunny days until the end of breeding.

[0051] The influence of the compound microbial preparation on inorganic nitrogen in the water in Example 2: 6 days after the first use of the compound microbial preparation, the ammonia nitrogen, nitrite and nitrate nitrogen in the water in the circular pond decreased by 25.17%, 29.42% and 31.52% respectively compared with the case where the compound microbial preparation is not used; and the influence on the growth of tilapia: 60 days after the first use of the compound microbial preparation, the average body length and average body weight of tilapia in the circular pond increased by 8.9% and 19.7% respectively compared with the case where the compound microbial preparation is not used.

[0052] Example 3

[0053] A microbial preparation regulation method for inorganic nitrogen in the water of a land-based circular pond tilapia breeding system, wherein the method comprises the following steps:

[0054] Step S1, 3 loops of Candida utilis preserved on an inclined plane are taken with an inoculation loop one, and the inoculation loop one is placed in a yeast extract malt extract medium, and then the medium is placed in a shaker for culture for 12 h. The bacterial liquid is turbid with obvious bacteria, and an activated strain one is obtained. The rotation speed of the shaker is 150 rpm, and the temperature is 30°C;

[0055] Step S2, the activated strain one is transferred to the yeast extract malt extract medium for expansion culture for 16 h, until the bacterial liquid concentration is >10 9 CFU / ml, 10 9 CFU / ml of Candida utilis bacterial liquid is obtained by dilution, the rotation speed of the shaker is 150 rpm, the pH is 7, the temperature is 30°C, and the inoculation amount of the activated strain one is 2%, and a Candida utilis bacterial liquid is obtained.

[0056] Step S3, 3 loops of Bacillus subtilis preserved on an inclined plane are taken with an inoculation loop two, and the inoculation loop is placed in an LB medium, and then the medium is placed in a shaker for culture for 10 h. The bacterial liquid is turbid with obvious bacteria, and an activated strain two is obtained. The rotation speed of the shaker is 150 rpm, and the temperature is 40°C;

[0057] Step S4, the activated strain two is transferred to the LB medium for expansion culture for 12 h, until the bacterial liquid concentration is >109 CFU / ml, obtained by dilution 10 9 CFU / ml Bacillus subtilis bacterial liquid, the rotation speed of the shaker was 150 rpm, the pH was 7, the temperature was 40℃, and the inoculation amount of the activated strain two was 2%;

[0058] Step S5, three Rhodopseudomonas palustris 3 rings preserved in the slant were taken with the inoculation ring, and the inoculation ring was placed in the light and bacterial culture medium for stationary culture for 60h. The bacterial liquid was obviously red, and the activated strain three was obtained. The light intensity was 4000Lx, and the temperature was 30℃.

[0059] The light and bacterial culture medium comprises ammonium chloride 2.0g / L, sodium acetate 3.0g / L, magnesium sulfate 0.5g / L, calcium chloride 0.1g / L, potassium dihydrogen phosphate 0.5g / L, dipotassium hydrogen phosphate 0.5g / L, and yeast extract 1.0g / L.

[0060] Step S6, the activated strain three was transferred to the light and bacterial culture medium for expansion culture for 65h, and the bacterial liquid concentration was >10 9 CFU / ml, obtained by dilution 10 9 CFU / ml Rhodopseudomonas palustris bacterial liquid, the light intensity was 4000Lx, the pH was 7, the temperature was 30℃, and the inoculation amount of the activated strain three was 10%;

[0061] Step S7, the Candida utilis bacterial liquid, the Bacillus subtilis bacterial liquid and the Rhodopseudomonas palustris bacterial liquid were mixed in the proportions of 45%, 45% and 10% of the total volume, respectively, to obtain a composite microbial preparation;

[0062] Step S8, a plurality of small land-based circular pools with a volume of 1m 3 In late May, 50 healthy Oreochromis aureus fry with an average body length of 4-6cm were raised in each circular pool, and during the breeding period, the fry were fed with granular compound feed. During the initial 1 month, the fry were fed 3 times a day, and the feeding amount was 2% of the body weight of the fish. Subsequently, the fry were fed 2 times a day, and the feeding amount was increased to 4%. The dissolved oxygen concentration in the water was controlled to be >4mg / L by an oxygenator, and the water was changed by 10% every day during the breeding period.

[0063] Step S9, after 20 days of breeding, the composite microbial preparation in step S7 was sprayed into the circular pool at an interval of 7 days. The spraying time was preferably selected in the morning on a sunny day, and the spraying was continued until the end of the breeding.

[0064] The influence of the composite microbial preparation on inorganic nitrogen in water in Example 3: 32.05%, 35.83% and 30.01% of ammonia nitrogen, nitrite and nitrate nitrogen in the water quality in the round pool decreased after 6 days of using the composite microbial preparation for the first time compared with not using the composite microbial preparation; the influence on the growth of tilapia: 9.85% and 24.62% of the average body length and average weight of tilapia in the round pool increased after 60 days of using the composite microbial preparation for the first time compared with not using the composite microbial preparation.

[0065] Example 4

[0066] A microbial preparation regulation method for inorganic nitrogen in water of a land-based round pool tilapia culture system, comprising the following steps:

[0067] Step S1, 3 loops of Candida utilis preserved on an inclined plane are taken with an inoculation loop one, and the inoculation loop one is placed in a yeast extract malt extract medium, and then the medium is placed in a shaking bed for culture for 12 hours, the bacterial liquid is turbid with obvious bacterial bodies, and an activated strain one is obtained, the rotation speed of the shaking bed is 150 rpm, and the temperature is 30 DEG C;

[0068] Step S2, the activated strain one is transferred to the yeast extract malt extract medium for expansion culture for 16 hours, and the bacterial liquid concentration is greater than 10 9 CFU / ml, 10 9 CFU / ml of Candida utilis bacterial liquid is obtained through dilution, the rotation speed of the shaking bed is 150 rpm, the pH is 7, the temperature is 30 DEG C, and the inoculation amount of the activated strain one is 2%, and Candida utilis bacterial liquid is obtained;

[0069] Step S3, 3 loops of Bacillus subtilis preserved on an inclined plane are taken with an inoculation loop two, and the inoculation loop is placed in an LB medium, and then the medium is placed in a shaking bed for culture for 10 hours, the bacterial liquid is turbid with obvious bacterial bodies, and an activated strain two is obtained, the rotation speed of the shaking bed is 150 rpm, and the temperature is 40 DEG C;

[0070] Step S4, the activated strain two is transferred to the LB medium for expansion culture for 12 hours, and the bacterial liquid concentration is greater than 10 9 CFU / ml, 10 9 CFU / ml of Bacillus subtilis bacterial liquid is obtained through dilution, the rotation speed of the shaking bed is 150 rpm, the pH is 7, the temperature is 40 DEG C, and the inoculation amount of the activated strain two is 2%;

[0071] Step S5, 3 loops of Rhodopseudomonas palustris preserved on an inclined plane are taken with an inoculation loop three, and the inoculation loop is placed in a light and bacterial culture medium for static culture for 60 hours, the bacterial liquid is obviously red, and an activated strain three is obtained, the light intensity is 4000 Lx, and the temperature is 30 DEG C;

[0072] The light and bacterial culture medium comprises 2.0 g / L of ammonium chloride, 3.0 g / L of sodium acetate, 0.5 g / L of magnesium sulfate, 0.1 g / L of calcium chloride, 0.5 g / L of potassium dihydrogen phosphate, 0.5 g / L of dipotassium hydrogen phosphate and 1.0 g / L of yeast extract.

[0073] In step S6, the activated strain three is transferred to the light and bacterial culture medium for expansion for 65 hours until the concentration of the Rhodopseudomonas palustris bacterial liquid is greater than 10 9 CFU / ml, 10 9 CFU / ml of the Rhodopseudomonas palustris bacterial liquid, the light intensity is 4000 Lx, the pH is 7, the temperature is 30 DEG C, and the inoculation amount of the activated strain three is 10%;

[0074] In step S7, the Rhodotorula mucilaginosa bacterial liquid, the Bacillus subtilis bacterial liquid and the Rhodopseudomonas palustris bacterial liquid are mixed at 40%, 40% and 20% of the total volume to obtain the compound microbial preparation.

[0075] In step S8, a plurality of small land-based circular pools with a volume of 1 m 3 are provided, and in late May, 50 healthy Oreochromis aureus fry with an average body length of 4-6 cm are raised in each circular pool. During the breeding period, the fry are fed with granular compound feed, three times a day for the first month, and the feeding amount is 2% of the body weight of the fish. Then, the fry are fed twice a day, and the feeding amount is increased to 4%. The dissolved oxygen concentration in the water is controlled to be greater than 4 mg / L by an oxygenator, and the water is changed by 10% every day during the breeding period.

[0076] In step S9, 90 ml of the compound microbial preparation in step S7 is sprayed into the circular pool every 7 days after 20 days of breeding, and the spraying time is preferably selected in the morning on a sunny day until the end of breeding.

[0077] Influence of the compound microbial preparation in Example 4 on inorganic nitrogen in water: After 6 days of using the compound microbial preparation for the first time, the ammonia nitrogen, nitrite and nitrate nitrogen in the water in the circular pool decreased by 20.88%, 21.39% and 25.43% respectively compared with the case where the compound microbial preparation is not used. Influence on the growth of the tilapia: After 60 days of using the compound microbial preparation for the first time, the average body length and average body weight of the tilapia in the circular pool increased by 8.1% and 16.9% respectively compared with the case where the compound microbial preparation is not used.

[0078] Comparative Example 1

[0079] A method for regulating inorganic nitrogen in water in a land-based circular pool tilapia breeding system, comprising the following steps:

[0080] Step S1, take 3 rings of Candida utilis preserved in the slant with inoculation ring one, and place the inoculation ring one in the yeast extract malt extract medium, then place the medium in the shaker for culture for 12h to obtain activated strain one, the rotation speed of the shaker is 150rpm, and the temperature is 30℃;

[0081] Step S2, transfer the activated strain one to the yeast extract malt extract medium for expansion culture for 16h until the concentration of the bacterial liquid is >10 9 CFU / ml, obtain 10 9 CFU / ml of Candida utilis bacterial liquid by dilution, the rotation speed of the shaker is 150rpm, the pH is 7, the temperature is 30℃, and the inoculation amount of the activated strain one is 2%;

[0082] Step S3, take 3 rings of Bacillus subtilis preserved in the slant with inoculation ring two, and place the inoculation ring in the LB medium, then place the medium in the shaker for culture for 10h until the bacterial liquid is turbid with obvious bacterial bodies to obtain activated strain two, the rotation speed of the shaker is 150rpm, and the temperature is 40℃;

[0083] Step S4, transfer the activated strain two to the LB medium for expansion culture for 12h until the concentration of the bacterial liquid is >10 9 CFU / ml, obtain 10 9 CFU / ml of Bacillus subtilis bacterial liquid by dilution, the rotation speed of the shaker is 150rpm, the pH is 7, the temperature is 40℃, and the inoculation amount of the activated strain two is 2%;

[0084] Step S5, take 3 rings of Rhodopseudomonas palustris preserved in the slant with inoculation ring three, and place the inoculation ring in the light and bacterial medium for static culture for 60h until the bacterial liquid is obviously red to obtain activated strain three, the light intensity is 4000Lx, and the temperature is 30℃;

[0085] The light and bacterial medium comprises ammonium chloride 2.0g / L, sodium acetate 3.0g / L, magnesium sulfate 0.5g / L, calcium chloride 0.1g / L, potassium dihydrogen phosphate 0.5g / L, dipotassium hydrogen phosphate 0.5g / L, and yeast extract 1.0g / L.

[0086] Step S6, transfer the activated strain three to the light and bacterial medium for expansion culture for 65h until the concentration of the bacterial liquid is >10 9 CFU / ml, obtain 10 9 CFU / ml of Rhodopseudomonas palustris bacterial liquid by dilution, the light intensity is 4000Lx, the pH is 7, the temperature is 30℃, and the inoculation amount of the activated strain three is 10%;

[0087] Step S7, the Candida utilis bacterial liquid, Bacillus subtilis bacterial liquid, Rhodopseudomonas palustris bacterial liquid are mixed according to 45%, 10%, 45% of the total volume, to obtain a composite microbial preparation;

[0088] Step S8, a plurality of small land-based circular pools with a volume of 1m 3 are provided, and in late May, 50 healthy Oreochromis aureus fry with an average body length of 4-6cm are raised in each circular pool, and during the breeding period, the fry are fed with granular compound feed, 3 times a day for the first month, and the feeding amount is 2% of the body weight of the fish; then 2 times a day, and the feeding amount is increased to 4%, and the dissolved oxygen concentration in the water is controlled to be greater than 4mg / L by an oxygenator, and the water needs to be replaced by 10% every day during the breeding period;

[0089] Step S9, after 20 days of breeding, the composite microbial preparation in step S7 is sprayed into the circular pool at 110ml every 7 days, and the spraying time is preferably selected in the morning on a sunny day, until the end of the breeding.

[0090] Influence of the composite microbial preparation in Comparative Example 1 on inorganic nitrogen in water: after 6 days of using the composite microbial preparation for the first time, the ammonia nitrogen and nitrate nitrogen in the water in the circular pool decreased by 11.05% and 14.21% respectively compared with not using the composite microbial preparation, and the nitrite nitrogen did not decrease significantly; influence on the growth of tilapia: after 60 days of using the composite microbial preparation for the first time, the average body length of tilapia in the circular pool increased by 4.9% compared with not using the composite microbial preparation, and the body weight did not change significantly.

[0091] Comparative Example 2

[0092] A method for regulating inorganic nitrogen in water in a land-based circular pool tilapia breeding system, comprising the following steps:

[0093] Step S1, 3 loops of Candida utilis preserved on an inclined plane are taken with an inoculation loop one, and the inoculation loop one is placed in a yeast extract malt extract medium, and then the medium is placed in a shaker for culture for 12h to obtain an activated strain one, the rotation speed of the shaker is 150rpm, and the temperature is 30℃;

[0094] Step S2, the activated strain one is transferred to the yeast extract malt extract medium for expansion culture for 16h, to a bacterial liquid concentration of >10 9 CFU / ml, 10 9 CFU / ml of Candida utilis bacterial liquid is obtained by dilution, the rotation speed of the shaker is 150rpm, the pH is 7, the temperature is 30℃, and the inoculation amount of the activated strain one is 2%, to obtain a Candida utilis bacterial liquid;

[0095] Step S3, take the Bacillus subtilis 3 ring preserved in the slope with the inoculation ring, and place the inoculation ring in the LB culture medium, then place the culture medium in the shaker for culture for 10h, the bacterial liquid is turbid with obvious bacteria, and the activated strain two is obtained, the rotation speed of the shaker is 150rpm, and the temperature is 40℃;

[0096] Step S4, transfer the activated strain two to the LB culture medium for expansion culture for 12h, to the bacterial liquid concentration >10 9 CFU / ml, obtain 10 9 CFU / ml of Bacillus subtilis liquid by dilution, the rotation speed of the shaker is 150rpm, the pH is 7, the temperature is 40℃, and the inoculation amount of the activated strain two is 2%;

[0097] Step S5, take the Rhodopseudomonas palustris 3 ring preserved in the slope with the inoculation ring three, and place the inoculation ring in the light and bacterial culture medium for static culture for 60h, the bacterial liquid is obviously red, and the activated strain three is obtained, the light intensity is 4000Lx, and the temperature is 30℃;

[0098] The light and bacterial culture medium comprises ammonium chloride 2.0g / L, sodium acetate 3.0g / L, magnesium sulfate 0.5g / L, calcium chloride 0.1g / L, potassium dihydrogen phosphate 0.5g / L, dipotassium hydrogen phosphate 0.5g / L, and yeast extract 1.0g / L.

[0099] Step S6, transfer the activated strain three to the light and bacterial culture medium for expansion culture for 65h, to the bacterial liquid concentration >10 9 CFU / ml, obtain 10 9 CFU / ml of Rhodopseudomonas palustris liquid by dilution, the light intensity is 4000Lx, the pH is 7, the temperature is 30℃, and the inoculation amount of the activated strain three is 10%;

[0100] Step S7, mix the Candida utilis liquid, Bacillus subtilis liquid and Rhodopseudomonas palustris liquid according to 45%, 45% and 10% of the total volume to obtain the compound microbial preparation;

[0101] Step S8, provide a plurality of small land-based circular pools with a volume of 1m 3 Each circular pool is stocked with 50 healthy Oreochromis aureus fry with an average body length of 4-6cm in late May, and the fry is fed with granular compound feed during the breeding period, 3 times a day for the first month, and the feeding amount is 2% of the fish weight; then 2 times a day, and the feeding amount is increased to 4%, and the dissolved oxygen concentration in the water is controlled to be >4mg / L by the oxygenator, and the water needs to be replaced by 10% every day during the breeding period;

[0102] Step S9, 20 days after the start of breeding, the compound microbial preparation 70ml in step S7 is sprayed into the round pool every 7 days, and the spraying time is chosen as much as possible in the morning on sunny days until the end of breeding.

[0103] The influence of the compound microbial preparation in Comparative Example 2 on inorganic nitrogen in water: 6 days after the first use of the compound microbial preparation, the nitrate nitrogen in the water in the round pool decreased by 7.83% compared with the case where the compound microbial preparation was not used, and the ammonia nitrogen and nitrite nitrogen did not decrease significantly; the influence on the growth of tilapia: 60 days after the first use of the compound microbial preparation, the average body length of the tilapia in the round pool increased by 4.1% compared with the case where the compound microbial preparation was not used, and the body weight did not change significantly.

[0104] Finally, it should be noted that the above specific embodiments are only used to illustrate the technical solutions of the present application and not to limit it. Although the present application has been described in detail with reference to the examples, it should be understood by those skilled in the art that the technical solutions of the present application can be modified or replaced equivalently without departing from the spirit and scope of the technical solutions of the present application, and they should be covered in the scope of the claims of the present application.

Claims

1. A method for regulating the inorganic nitrogen in water in a land-based circular pond tilapia farming system using a microbial formulation, characterized by: The method comprises the following steps: Step S1, taking 2-3 circles of Candida utilis preserved in a slant with a inoculation loop one, and placing the inoculation loop one in a yeast extract malt wort medium, and then placing the medium in a shaker for culture for at least 12 hours to obtain an activated strain one; Step S2, the activated strain is transferred to a yeast extract wort medium for expansion for at least 12 h, until the bacterial liquid concentration is >10 9 CFU / ml, Candida utilis bacterial liquid is obtained; Step S3, taking 2-3 circles of Bacillus subtilis preserved in a slant with a inoculation loop two, and placing the inoculation loop in an LB medium, and then placing the medium in a shaker for culture for at least 10 hours to obtain an activated strain two; Step S4, the activated strain is subcultured in LB medium for at least 10 h to a bacterial liquid concentration > 10 9 CFU / ml, obtaining a Bacillus subtilis bacterial liquid; Step S5, taking 2-3 circles of Rhodopseudomonas palustris preserved in a slant with a inoculation loop three, and placing the inoculation loop in a light and bacteria medium for static culture for at least 60 hours to obtain an activated strain three; Step S6, the activated strain is subcultured in light and bacterial culture medium for at least 60 h to obtain a Rhodopseudomonas palustris bacterial liquid with a concentration of >10 9 CFU / ml, obtaining a Rhodopseudomonas palustris bacterial liquid; Step S7, mixing Candida utilis liquid, Bacillus subtilis liquid and Rhodopseudomonas palustris liquid according to 40-45%, 40-45% and 10-20% of the total volume to obtain a compound microbial preparation; Step S8, provide a plurality of small land-based circular pools with a volume of 1 m 3 In late May, 40-50 healthy Oreochromis aureus fry with an average body length of 4-6 cm were raised in each circular pool. During the breeding period, pellet feed was used for feeding, and the dissolved oxygen concentration in the water was controlled to be >4 mg / L by an oxygenator. Step S9, after 20-30 days of breeding, spraying 90-110 ml of the compound microbial preparation in step S7 into the round pool every 7-10 days until the end of breeding; The feeding in step S8 is specifically as follows: feeding 3-4 times a day with a feeding amount of 1%-3% of the weight of the fish in the first 1-2 months; and then feeding 2-3 times a day with a feeding amount of 4%-6%.

2. The method of claim 1, wherein the method is characterized by: The rotation speed of the shaker in step S1 is 130-180 rpm, and the temperature is 28-32 DEG C; the rotation speed of the shaker in step S2 is 130-180 rpm, the pH is 7-8, and the temperature is 28-32 DEG C; and the inoculation amount of the activated strain one is 2%-3%.

3. The method of claim 1, wherein the method is characterized by: The rotation speed of the shaker in step S3 is 130-180 rpm, and the temperature is 30-40 DEG C; the rotation speed of the shaker in step S4 is 130-180 rpm, the pH is 6-8, and the temperature is 30-40 DEG C; and the inoculation amount of the activated strain two is 2%-3%.

4. The method for the microbial manipulation of inorganic nitrogen in the water of a land-based circular tank tilapia farming system according to claim 1, characterized in that: The light and bacteria medium in step S5 and step S6 comprises 2.0 g / L of ammonium chloride, 3.0 g / L of sodium acetate, 0.5 g / L of magnesium sulfate, 0.1 g / L of calcium chloride, 0.5 g / L of potassium dihydrogen phosphate, 0.5 g / L of dipotassium hydrogen phosphate and 1.0 g / L of yeast extract.

5. The method of claim 1, wherein the method is characterized by: The light intensity of the light and bacteria medium in step S5 is 3500-5500 Lx, and the temperature is 30-35 DEG C; the light intensity in step S6 is 3500-5500 Lx, the pH is 6-8, and the temperature is 30-35 DEG C; and the inoculation amount of the activated strain three is 10-20%.

6. The method of claim 1, wherein the method is characterized by: In step S8, 10-15% of water needs to be changed every day during breeding.