Denitrifying bacteria and application thereof

Abstract

The invention relates to the field of microorganisms, and discloses a denitrifying bacterium and application thereof, the denitrifying bacterium is Pseudomonas peli, and the preservation number of the denitrifying bacterium is CGMCC (China General Microbiological Culture Collection Center) No. 30426. The denitrifying bacterium provided by the invention can effectively reduce nitrate nitrogen in a water body, and the strain has good ecological safety and is non-toxic to fishes.

Description

Technical Field

[0001] The invention relates to the technical field of microorganisms, and in particular to a denitrifying bacterium and application thereof. Background Art

[0002] my country is the world's largest aquaculture country. In 2022, my country's freshwater aquaculture output was 32.9 million tons, and the feed coefficient of aquatic animals was 1-2. More than 32.9 million tons of feed are used in various modes of freshwater aquaculture each year. Only 25% of the nutrients in the feed are converted into economic animals. Most of the nutrients in the feed exist in the water in organic or inorganic form. Various forms of pollutants including nitrogen and phosphorus generated by intensive aquaculture are discharged into the receiving water, causing the surrounding water environment to deteriorate. Among them, excessive nitrogen is one of the key indicators of pollution load.

[0003] With the increasing attention paid to the ecological environment and the green and sustainable development of aquaculture, freshwater aquaculture tailwater treatment facilities have been further strengthened. At present, most of the major aquaculture provinces and cities in the country have issued aquaculture tailwater discharge standards, requiring aquaculture tailwater to be treated and allowed to be discharged only when it meets the relevant standards. There are currently two main ways to treat aquaculture tailwater, one is centralized treatment and the other is in-situ treatment. Among these two methods, compared with chemical methods, the microbial denitrification process based on denitrification has attracted the attention of many researchers due to its low operating cost and no residue. Unlike domestic sewage biochemical pools, there are farmed animals in aquaculture ponds and aquaculture tailwater treatment facilities, and the dissolved oxygen is relatively high, so anaerobic denitrification is difficult to achieve in freshwater aquaculture. Due to the lack of denitrifying bacteria used in aquaculture and aquaculture tailwater treatment, the removal of nitrogen from aquaculture tailwater is still a problem. Summary of the invention

[0004] The purpose of the present invention is to overcome the problem that the prior art lacks denitrifying bacteria used in aquaculture and aquaculture tail water treatment, resulting in difficulty in denitrifying aquaculture tail water, and to provide a denitrifying bacterium and its application in freshwater aquaculture water.

[0005] The inventor of the present invention isolated a strain from the intestine of channel catfish, which was identified as Pseudomonas peli. The strain has a good function of removing nitrate and nitrite, and can be used for the removal of nitrate in aquaculture. Therefore, in order to achieve the above purpose, the first aspect of the present invention provides a denitrifying bacterium, which is Pseudomonas peli, and the deposit number is CGMCC No. 30426.

[0006] The second aspect of the present invention provides the use of the denitrifying bacteria described in the first aspect of the present invention in water treatment.

[0007] A third aspect of the present invention provides a method for treating a water body, characterized in that the method comprises contacting the denitrifying bacteria as described above with the water body to be treated.

[0008] Through the above technical solution, the present invention has at least the following beneficial effects:

[0009] (1) The denitrifying bacteria provided by the present invention are isolated from the intestine of channel catfish and are a strain of Pseudomonas sludge isolated from the intestine of farmed fish. They have good environmental ecological safety and are non-pathogenic to fish.

[0010] (2) The denitrifying bacteria provided by the present invention have good functions of removing nitrate nitrogen and nitrite nitrogen in freshwater aquaculture water, can be effectively used for the removal of nitrate nitrogen in aquaculture, and the treatment cycle is short.

[0011] Biological Deposit

[0012] The Pseudomonas peli of the present invention is numbered H46-1 and was deposited in the General Microbiology Center of China Microorganism Culture Collection Committee (address: No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, Postal Code: 100101) on April 24, 2024 (the abbreviation of the depository unit is CGMCC), and the deposit number is CGMCC No.30426. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 It is the hemolytic activity of Staphylococcus aureus (left) and Pseudomonas sludge of the present invention (right);

[0014] Figure 2 The growth of the Pseudomonas sludge of the present invention in a nitrate nitrogen medium;

[0015] Figure 3 This is a diagram showing the effect of the sludge pseudomonas bacteria of the present invention in removing nitrate nitrogen;

[0016] Figure 4 The growth of the Pseudomonas sludge of the present invention in a nitrite nitrogen culture medium;

[0017] Figure 5 This is a diagram showing the effect of the sludge Pseudomonas bacteria of the present invention in removing nitrite nitrogen;

[0018] Figure 6 This is a diagram showing the effect of the present invention's Pseudomonas sludge bacteria on the removal of nitrite nitrogen in a circulating water system (C / N=15);

[0019] Figure 7 This is a diagram showing the effect of the present invention's Pseudomonas sludge bacteria on the removal of nitrate nitrogen in a circulating water system (C / N=15);

[0020] Figure 8 This is a diagram showing the effect of the present invention's Pseudomonas sludge bacteria on the removal of nitrite nitrogen in a circulating water system (C / N=10);

[0021] Fig. 9 This is a diagram showing the effect of the Pseudomonas sludge bacteria of the present invention on the removal of nitrate nitrogen in a circulating water system (C / N=10). DETAILED DESCRIPTION

[0022] The endpoints and any values ​​of the ranges disclosed in this article are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of each range, the endpoint values ​​of each range and the individual point values, and the individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed in this article.

[0023] During the research, the inventors of the present invention isolated a denitrifying bacterium from the intestine of channel catfish, which was identified as Pseudomonas peli by 16s RNA gene sequencing. The deposit number of the denitrifying bacterium is CGMCC No. 30426. It is numbered H46-1 in the present invention.

[0024] The 16S rRNA gene sequence of the denitrifying bacteria provided by the present invention is shown in SEQ ID NO: 1.

[0025] SEQ ID NO:1

[0026]

[0027] The morphological characteristics of the denitrifying bacteria of the present invention are as follows: after being cultured on a denitrifying agar plate at 28° C. for 48 hours, the colonies are round, with neat edges, a diameter of about 1 mm, and slightly raised surfaces. Under a microscope, the cells are rod-shaped, moving, no spores are observed, and Gram staining is negative. The denitrifying agar plate is composed of: 5 g / L sodium citrate, 1 g / L K2HPO4, 1 g / L KH2PO4, 0.2 g / L MgSO4·7H2O, 1 g / L NaNO3, 2 mL / L trace element solution, and 0.05 g / L bromophenol blue. Trace element solution: 0.2 g / L CaCl2, 1 g / L FeSO4·7H2O, 0.35 g / L CoCl2·6H2O, 0.4 g / L CuSO4·5H2O, 0.8 g / L MnCl2·7H2O, (NH4)6Mo7O 24 ·4H2O 0.65g / L, ZnSO4 0.4g / L.

[0028] The denitrifying bacteria of the present invention were subjected to API 20NE detection, and the physiological and biochemical characteristics thereof are shown in Table 1 below.

[0029] Table 1 Physiological and biochemical characteristics of Pseudomonas sludge

[0030]

[0031] The second aspect of the present invention provides the use of the denitrifying bacteria described in the first aspect of the present invention in water treatment.

[0032] In the present invention, the water body may be a common water body that needs to be treated with nitrate nitrogen, preferably a freshwater aquaculture water body, such as a freshwater fish aquaculture water body.

[0033] In the present invention, the method can treat water with a C / N ratio of 5-20, preferably 10-15.

[0034] In the present invention, the method can treat water bodies with a nitrite nitrogen content of 0-20 mg / L and a nitrate nitrogen content of 0-50 mg / L. Preferably, the nitrite nitrogen content in the water body is 0.2-5 mg / L, and the nitrate nitrogen content is 5-30 mg / L.

[0035] The third aspect of the present invention provides a water treatment method, the treatment method comprising contacting the denitrifying bacteria as described above with the water to be treated. The water may be as described above.

[0036] In the present invention, the contacting method between the denitrifying bacteria and the water body includes directly adding the denitrifying bacteria and / or loading the denitrifying bacteria onto a carrier and then adding the denitrifying bacteria to the water body to be treated.

[0037] In the present invention, in order to achieve a better treatment effect of nitrate nitrogen, the concentration of denitrifying bacteria in the water body is 1×10 2 -1×10 8 CFU / mL, preferably 1×10 3 -1×10 6 CFU / mL.

[0038] In the present invention, the contacting conditions may include: a temperature of 25-30° C. and a time of 1-30 days.

[0039] The present invention will be described in detail below by way of examples. It should be understood that the following examples are only used to further explain and illustrate the content of the present invention, and are not intended to limit the present invention.

[0040] Unless otherwise specified, the reagents and materials used in the following examples were purchased from regular chemical reagent suppliers and were of analytical grade.

[0041] The formula of the Pseudomonas sludge culture medium used in the following examples is K2HPO4 1g / L, KH2PO4 1g / L, MgSO4·7H2O 0.2g / L, trace element solution 2mL / L, NaNO3 0.085g / L, trisodium citrate dihydrate 0.856g / L. The trace element solution includes CaCl2 0.2g / L, FeSO4·7H2O 1g / L, CoCl2·6H2O 0.35g / L, CuSO4·5H2O0.4g / L, MnCl2·7H2O 0.8g / L, (NH4)6Mo7O 24 ·4H2O 0.65g / L, ZnSO4 0.4g / L, the solution was sterilized by filtration through a 0.22μm filter membrane.

[0042] The nitrite nitrogen concentration in the following examples was determined using the "Water quality - Determination of nitrite nitrogen - Spectrophotometric method GB 7493-87". The nitrate nitrogen concentration was determined using the "Water quality - Determination of nitrate nitrogen - Ultraviolet spectrophotometric method HJ / T 346-2007".

[0043] Example 1

[0044] This example is used to illustrate the safety of the denitrifying bacteria provided by the present invention.

[0045] The density of Pseudomonas sludge was set to 0 and 1×10 5 , 1×10 6 and 1×10 7CFU / mL, and three parallel experiments were set up in each group. Each experimental group had 15 zebrafish. After anesthesia, the scales of about 20% of the body surface area were scraped off the tail and placed in the water containing bacteria for immersion infection test. The observation lasted for 14 days, and the mortality rate was counted every day and the symptoms of infection were observed. In the mud, the density of Pseudomonas species was 1×10 5 , 1×10 6 and 1×10 7 In the experimental groups with three CFU / mL concentrations and the control group without bacteria, there was no death of zebrafish within 14 days and no obvious abnormal symptoms were observed, indicating that Pseudomonas sludge is not pathogenic to zebrafish.

[0046] Example 2

[0047] This example is used to illustrate the safety of the denitrifying bacteria provided by the present invention.

[0048] Pseudomonas sludge was activated and cultured for 24 hours, 2 μL of the bacterial solution was spotted on a blood agar plate, and cultured at 28°C for 48 hours to observe whether a hemolytic ring was produced. Staphylococcus aureus was used as a positive control.

[0049] like Figure 1 As shown, on the blood agar plate, the positive control Staphylococcus aureus produced an obvious hemolytic ring, while Pseudomonas sludge had no hemolytic ring.

[0050] Example 3

[0051] This example is used to illustrate the denitrification performance of the denitrifying bacteria provided by the present invention on nitrate nitrogen.

[0052] Nitrate nitrogen removal test culture medium: sodium citrate dihydrate 5g / L, K2HPO4 1g / L, KH2PO4 1g / L, MgSO4·7H2O 0.2g / L, trace element solution 2mL / L, NaNO3 1g / L.

[0053] Prepare the nitrate denitrification capacity test medium, and make 3 parallels for each group. Take the bacterial solution in the logarithmic growth period and inoculate it into the test medium, so that the initial OD600 is 0.1-0.2, and shake and culture at 28℃ and 180r / min. Take samples every 12h to measure the OD600 and nitrate nitrogen concentration until 72h.

[0054] like Figure 2 As shown in Figure 2, Pseudomonas sludge grew well in the nitrate nitrogen denitrification test medium, and the bacterial density reached the maximum after 24 hours, at which time the nitrate nitrogen removal rate was greater than 95.3% ( Figure 3 ), within 36-72h, the level of nitrate nitrogen was lower than 10mg / L.

[0055] Example 4

[0056] This example is used to illustrate the denitrification performance of the denitrifying bacteria provided by the present invention on nitrite nitrogen.

[0057] Nitrite nitrogen removal test culture medium: sodium citrate dihydrate 5g / L, K2HPO41 g / L, KH2PO4 1g / L, MgSO4·7H2O 0.2g / L, trace element solution 2mL / L, KNO2 0.5g / L.

[0058] Prepare the denitrification test medium for nitrite, and make 3 parallels for each group. Take the bacterial solution in the logarithmic growth phase and inoculate it into the test medium, so that the initial OD600 is 0.1-0.2, and shake and culture at 28℃ and 180r / min. Take samples every 12h to measure the OD600 and nitrite nitrogen concentration until 72h.

[0059] like Figure 4 As shown in Figure 2, Pseudomonas sludge grew well in the nitrite denitrification ability test medium, and the bacterial density reached the maximum after 24 hours, at which time the nitrite removal rate reached 95.28% ( Figure 5 ), the concentration of nitrite nitrogen ranged from 3.7 to 7.7 mg / L within 36 to 72 hours.

[0060] Example 5

[0061] This example is used to illustrate the performance of the denitrifying bacteria provided by the present invention in removing nitrite nitrogen and nitrate nitrogen in a circulating water system.

[0062] The experimental circulating water system consists of aquaculture facilities, a circulating pump and a filtering device. A 60L plastic basin is used as aquaculture facilities, each basin contains 30L of tap water, and a filtering device is set up above the plastic basin. The filtering device consists of a filter box and filter materials, and the filter materials are volcanic rocks, gravel and filter cotton.

[0063] The entire circulating water system was treated with trichlorfon disinfection and aerated for two days. The water temperature was 26±2℃. Fifteen zebrafish were placed in each system, with a body weight of 0.25±0.1g / fish. They were fed once a day, with a feeding amount of about 3% of their body weight. The circulating water was closed for 30 minutes during feeding, and feeding was continued for 4 days. Day 0 was the day after 4 days of feeding.

[0064] Soak the filter cotton in 1.8L fresh sludge Pseudomonas culture medium, inoculate 200mL sludge Pseudomonas, continue aeration, and culture at 28℃ for 12h. At this time, the bacterial density is 2×10 8 CFU / mL. Take out the filter cotton containing bacteria and put it into the filtration device on Day 0. Turn on the circulation pump. The bacterial density of the aquaculture water is about 1×10 6 CFU / mL.

[0065] Six independent circulating water systems were used, and three parallel experiments were set up for the control group without bacteria and the treatment group with bacteria. Sodium nitrite was added to the control group and the treatment group until the nitrite nitrogen was 4 mg / L, and sodium nitrate was added to the aquaculture facility water body until the nitrate nitrogen was 41 mg / L, and C / N was set to 15. Filter cotton containing Pseudomonas sludge and sodium citrate dihydrate with a final concentration of 2.756 g / L were added. Nitrite nitrogen and nitrate nitrogen indicators were measured on Day 0, 1, 3, 5, 7, 9, and 11, respectively.

[0066] like Figure 6 As shown, in the circulating water system, the nitrite nitrogen level in the control group was in the range of 3.62-4.10 mg / L within 11 days. The nitrite nitrogen level in the control group on Day 0 was 3.96±0.09 mg / L, and that in the treatment group was 4.04±0.10 mg / L. After 1 day, the nitrite nitrogen in the treatment group increased significantly, reaching 20.07±1.39 mg / L, which was significantly higher than the control group (p<0.05). On the 3rd day, the nitrite nitrogen in the treatment group decreased to 3.97±0.28 mg / L. After the 5th day, the nitrite nitrogen concentration in the treatment group was already below the detection limit (0.003 mg / L), and the nitrite nitrogen on the 5th, 7th, 9th and 11th days was below the detection limit.

[0067] like Figure 7 As shown, in the circulating water system, the nitrate nitrogen level in the control group was in the range of 35.11-42.84 mg / L within 11 days. The nitrate nitrogen level in the control group on Day 0 was 40.11±1.72 mg / L, and that in the treatment group was 40.00±2.36 mg / L. After 1 day, the nitrate nitrogen level in the treatment group was significantly lower than that in the control group (p<0.05), reaching 12.61±2.49 mg / L. After 3 days, the nitrate nitrogen concentration in the treatment group dropped to 3.66±0.92 mg / L, which was significantly different from the control group (p<0.05). On days 5, 7, 9, and 11, the nitrate nitrogen concentration in the treatment group dropped below 3 mg / L.

[0068] Example 6

[0069] This example is used to illustrate the performance of the denitrifying bacteria provided by the present invention in removing nitrite nitrogen and nitrate nitrogen in a circulating water system.

[0070] The experimental circulating water system consists of aquaculture facilities, a circulating pump and a filtering device. A 60L plastic basin is used as aquaculture facilities, each basin contains 30L of tap water, and a filtering device is set up above the plastic basin. The filtering device consists of a filter box and filter materials, and the filter materials are volcanic rocks, gravel and filter cotton.

[0071] The entire circulating water system was treated with trichlorfon disinfection and aerated for two days. The water temperature was 26±2℃. Fifteen zebrafish were placed in each basin, with a body weight of 0.25±0.1g / fish. They were fed once a day, with a feeding amount of about 3% of their body weight. The circulating water was closed for 30 minutes during feeding, and feeding was continued for 4 days. After 4 days of feeding, it was Day 0.

[0072] Soak the filter cotton in 1.8L fresh sludge Pseudomonas culture medium, inoculate 200mL sludge Pseudomonas, continue aeration, and culture at 28℃ for 12h. At this time, the bacterial density is 2×10 8 CFU / mL. Take out the filter cotton containing bacteria and put it into the filtration device on Day 0. Turn on the circulation pump. The bacterial density of the aquaculture water is about 1×10 6 CFU / mL.

[0073] Six independent circulating water systems were used, and three parallel experiments were set up for the control group without bacteria and the treatment group with bacteria. Sodium nitrite was added to the control group and the treatment group until the nitrite nitrogen was 4 mg / L, and sodium nitrate was added to the nitrate nitrogen in the aquaculture facility water body to 41 mg / L, and C / N was set to 10. Filter cotton containing Pseudomonas sludge and sodium citrate dihydrate with a final concentration of 1.837 g / L were added. Nitrite nitrogen and nitrate nitrogen indicators were measured on Day 0, 1, 3, 5, 7, 9, and 11, respectively.

[0074] like Figure 8 As shown, in the circulating water system, the nitrite nitrogen level in the control group was in the range of 3.62-4.10 mg / L within 11 days. The nitrite nitrogen level in the control group on Day 0 was 3.96±0.09 mg / L, and that in the treatment group was 3.95±0.12 mg / L. After 1 day, the nitrite nitrogen in the treatment group increased significantly, reaching 19.34±1.27 mg / L, which was significantly higher than that in the control group (p<0.05). On the 3rd day, the treatment group decreased to 9.57±1.25 mg / L. On the 5th day, the nitrite nitrogen concentration in the treatment group was 4.87±1.41 mg / L, which was similar to that in the control group. On the 7th day, it was 1.05±0.17 mg / L, on the 9th day, it was 0.76±0.23 mg / L, and on the 11th day, it was 0.76±0.14 mg / L.

[0075] like Fig. 9 As shown, in the circulating water system, the nitrate nitrogen level in the control group was in the range of 35.11-42.84 mg / L within 11 days. The nitrate nitrogen level in the control group on Day 0 was 40.11±1.72 mg / L, and that in the treatment group was 40.91±3.43 mg / L. After 1 day, the nitrate nitrogen level in the treatment group was significantly lower than that in the control group (p<0.05), reaching 21.42±7.98 mg / L. After 3 days, the nitrate nitrogen concentration in the treatment group dropped to 3.43±1.47 mg / L, which was significantly different from that in the control group (p<0.05). On the 5th, 7th, 9th, and 11th days, the nitrate nitrogen concentration in the treatment group dropped to below 3 mg / L.

[0076] Example 7

[0077] This example is used to illustrate the performance of the denitrifying bacteria provided by the present invention in removing nitrite nitrogen and nitrate nitrogen in a pond.

[0078] The strain was tested on nitrite and nitrate nitrogen in ponds at a farm in Beijing. The test was conducted in early September. The details are as follows:

[0079] The nitrite nitrogen in pond 2# was higher at 0.56mg / L, and the nitrate nitrogen was 12.63mg / L. The nitrite nitrogen in pond 5# was 0.38mg / L, and the nitrate nitrogen was 10.82mg / L. Pond 2# was used as the treatment group, and the whole pond was sprayed with the culture solution of Pseudomonas sludge. Pond 5# was used as the control group, and no treatment was done. The area of ​​pond 2# was 4.6 mu, and the area of ​​pond 5# was 7 mu. Ponds 2# and 5# mainly raised channel catfish, silver carp and carp, with similar stocking density, specifications and time.

[0080] After culturing in the sludge Pseudomonas medium for 24 h under laboratory conditions, the concentration of the strain was 2×10 8 CFU / mL, 10L of bacterial solution was added to the sludge Pseudomonas culture medium and 5kg of brown sugar in a 100L clean bucket in the farm for further culture. After 12h of culture, the plate count bacterial density was 1×10 8 CFU / mL, 100L of the culture solution was poured into the treatment pond, and the final concentration of Pseudomonas in the pond water was about 1×10 3 CFU / mL.

[0081] The control group ponds and the treatment group ponds adopted the same management measures such as feeding and oxygenation, and the pond water temperature, dissolved oxygen, nitrite nitrogen and nitrate levels were monitored at 0d, 1d, 2d, 4d, 8d, 10d and 12d respectively.

[0082] Pseudomonas sludge could significantly reduce the nitrite level in pond 2#. After 8 days, the nitrite level was reduced to 0.24 mg / L, a 57.14% decrease from day 0. The nitrite levels on days 10 and 12 were 0.3 and 0.31 mg / L respectively. The nitrite level in the control pond was slightly increased, as shown in Table 2.

[0083] Table 2 Changes in nitrite concentration in the aquaculture pond after using Pseudomonas sludge in 12 days

[0084]

[0085] Pseudomonas sludge could significantly reduce the nitrate level in the two ponds. After 8 days, the nitrate nitrogen in the treatment group 2 was reduced to 6.27 mg / L, a 50.35% decrease from the 0th day. The nitrate nitrogen on the 10th and 12th days was 6.57 and 6.42 mg / L respectively. The nitrate nitrogen level in the control group pond 5# was similar to that on the 0th day, with no change.

[0086] Table 3 Changes in nitrate concentration in aquaculture ponds after using Pseudomonas sludge at 12 days

[0087]

[0088] The preferred embodiments of the present invention are described in detail above, but the present invention is not limited thereto. Within the technical concept of the present invention, the technical solution of the present invention can be subjected to a variety of simple modifications, including the combination of various technical features in any other suitable manner, and these simple modifications and combinations should also be regarded as the contents disclosed by the present invention and belong to the protection scope of the present invention.

Claims

1. A denitrifying bacterium, characterized in that: The denitrifying bacteria is Pseudomonas peli, and its preservation number is CGMCC No.30426.

2. The denitrifying bacteria according to claim 1, wherein The sequence of 16S rRNA of Pseudomonas sludge is shown as SEQ ID NO:

1.

3. Use of the denitrifying bacteria according to claim 1 or 2 in water treatment.

4. The use according to claim 3, wherein: The water body is a freshwater aquaculture water body.

5. The use according to claim 3, wherein: The C / N ratio of the water body is 5-20, preferably 10-15.

6. The use according to claim 3, wherein: The nitrite nitrogen content in the water body is 0-20 mg / L, and the nitrate nitrogen content in the water body is 0-50 mg / L. Preferably, the nitrite nitrogen content in the water body is 0.2-5 mg / L, and the nitrate nitrogen content in the water body is 5-30 mg / L.

7. A method for treating a water body, characterized in that: The treatment method comprises: contacting the denitrifying bacteria described in claim 1 or 2 with the water body to be treated.

8. The method according to claim 7, wherein: The contact method is to directly add the denitrifying bacteria and / or load the denitrifying bacteria onto a carrier and then add the denitrifying bacteria to the water body to be treated.

9. The method according to claim 7, wherein: The dosage of the denitrifying bacteria is 1×10 2 -1×10 8 CFU / mL water, preferably 1×10 3 -1×10 6 CFU / mL water.

10. The method according to claim 7, wherein: The water body is a freshwater aquaculture water body; and / or, the C / N ratio of the water body is 5-20, preferably 10-15; And / or, the nitrite nitrogen content in the water body is 0-20 mg / L, and the nitrate nitrogen content in the water body is 0-50 mg / L. Preferably, the nitrite nitrogen content in the water body is 0.2-5 mg / L, and the nitrate nitrogen content in the water body is 5-30 mg / L.

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