A salt-tolerant and drug-resistant Pseudomonas stutzeri aerobic denitrifying bacterium RAS-L11, microbial agent and its application

Abstract

The present invention relates to the field of microbial technology, and in particular to a salt-resistant and drug-resistant Pseudomonas stutzeri aerobic denitrifying bacteria RAS-L11, a microbial agent and its application. The salt-resistant and drug-resistant Pseudomonas stutzeri aerobic denitrifying bacteria is named RAS-L11, classified as Pseudomonas stutzeri (Stutzerimonas stutzeri), and is preserved in the General Microbiological Center of the China Microbiological Culture Collection Administration Committee, with a preservation address of No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, a preservation date of May 20, 2024, and a preservation number of CGMCC No. 30692. The strain RAS-L11 has a wide range of antibiotic adaptability and salinity adaptability, can still maintain a high denitrification capacity in water bodies containing different types of antibiotics or having a certain salinity, and can simultaneously remove nitrate nitrogen, nitrite nitrogen and ammonia nitrogen in the water body.

Description

Technical Field

[0001] The invention relates to the technical field of microorganisms, and in particular to a salt-resistant and drug-resistant Pseudomonas stutzeri aerobic denitrifying bacteria RAS-L11, a microbial agent and applications thereof. Background Art

[0002] Nitrogen is an essential component for animal protein synthesis and plant growth, and is one of the most important nutrients for organisms. However, with the expansion of cities and the development of industry and agriculture in recent years, excessive inorganic nitrogen (mainly ammonia nitrogen, nitrate nitrogen and nitrite nitrogen) has been discharged into water bodies, becoming an important water pollutant. However, excessive nitrogen will not only destroy the aquatic ecosystem, leading to acidification and eutrophication of lakes, rivers, offshore oceans, etc., but also endanger the health of humans and organisms and even threaten their survival in severe cases. Traditional water denitrification technologies mainly include chemical denitrification, physical denitrification and biological denitrification, among which biological denitrification is widely used due to its low cost, mild reaction conditions, and degradation without secondary pollution. Traditional biological denitrification includes aerobic nitrification by autotrophic bacteria and anaerobic denitrification by heterotrophic bacteria, but because it consists of two parts of the process, it has the disadvantages of complex operation, high energy consumption and high operating costs. Moreover, traditional theory holds that denitrification can only be carried out under anaerobic or anoxic conditions. The discovery of aerobic denitrifying bacteria has broken through traditional understanding and can achieve denitrification under aerobic conditions, thereby achieving simultaneous nitrification and denitrification in the same reaction system, greatly saving operating time and costs.

[0003] A large number of studies have shown that most antibiotics have a broad antimicrobial spectrum and will kill certain species of microorganisms in the environment or inhibit the growth and reproduction of certain microorganisms. In sewage biological treatment systems, the presence and accumulation of antibiotics will have a significant negative effect on the aerobic denitrification of biological denitrification treatment technology, not only inhibiting the growth of aerobic denitrifying bacteria and reducing the activity of aerobic denitrifying enzymes, but also changing the structure of aerobic denitrifying microbial communities and reducing the abundance of aerobic denitrifying microbial communities, thereby affecting the aerobic denitrification rate. In addition, since pharmaceutical wastewater and domestic sewage contain some high-salinity wastewater and sewage, in a high-salinity environment, the increase in osmotic pressure will inhibit cell metabolism and enzyme activity, thereby affecting the efficiency of aerobic denitrification, and further affecting the sewage treatment effect. Summary of the invention

[0004] In view of this, the present invention provides a salt-tolerant and drug-tolerant Pseudomonas stutzeri aerobic denitrifying bacteria RAS-L11, a microbial agent and an application thereof. The aerobic denitrifying bacteria RAS-L11 can tolerate different antibiotics and higher salinity, and especially has a relatively high denitrification capacity in water bodies containing antibiotics or having a certain salinity.

[0005] To solve the above technical problems, the first aspect of the present invention provides a salt-resistant and drug-resistant Pseudomonas stutzeri aerobic denitrifying bacteria, the strain is named RAS-L11, classified as Pseudomonas stutzeri (Stutzerimonas stutzeri), and is deposited in the General Microbiological Center of the China Microbiological Culture Collection Administration, with a storage address of No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, a storage date of May 20, 2024, and a storage number of CGMCC No. 30692.

[0006] The aerobic denitrifying bacteria RAS-L11 provided by the present invention has strong adaptability to different types of antibiotics and salinity, can maintain a high denitrification capacity in water bodies containing different types of antibiotics or with a certain salinity, and can simultaneously remove nitrate nitrogen, nitrite nitrogen and ammonia nitrogen in the water body.

[0007] In combination with the first aspect, the colonies of the salt-resistant and drug-resistant Pseudomonas stutzeri aerobic denitrifying bacteria are milky white, slightly convex, with a moist and smooth surface, regular edges, uniform and opaque texture, and are easy to pick.

[0008] In combination with the first aspect, the salt-resistant and drug-resistant Pseudomonas stutzeri aerobic denitrifying bacteria have a long rod shape.

[0009] The second aspect of the present invention provides a use of the above-mentioned salt-resistant and drug-resistant Pseudomonas stutzeri aerobic denitrifying bacteria in water denitrification.

[0010] In combination with the second aspect, the denitrification includes removing nitrate nitrogen, ammonia nitrogen and / or nitrite nitrogen from the water body.

[0011] In combination with the second aspect, the salinity of the water body is 0% to 8%, preferably 3% to 5%.

[0012] In conjunction with the second aspect, the water body includes pharmaceutical wastewater, domestic sewage, aquaculture wastewater and industrial wastewater, but is not limited to the above water bodies, and is also applicable to other qualified water bodies. The strain is applied in the process of pharmaceutical wastewater treatment, domestic sewage treatment, aquaculture wastewater treatment and industrial wastewater treatment, which can significantly reduce the various antibiotics present in the water body and the influence of high salinity on the biological denitrification process, which is of great significance to ensure the normal operation of the treatment system.

[0013] In combination with the second aspect, the water body contains antibiotics, and the antibiotics include tetracycline antibiotics, quinolone antibiotics, macrolide antibiotics, nitroimidazole antibiotics and sulfonamide antibiotics.

[0014] In combination with the second aspect, the carbon-nitrogen ratio of the water body is not less than 4, preferably the carbon-nitrogen ratio is greater than or equal to 6, for example, the carbon-nitrogen ratio is 6, 8 or 10.

[0015] The third aspect of the present invention provides a microbial agent, the active ingredients of which include the above-mentioned salt-resistant and drug-resistant Pseudomonas stutzeri aerobic denitrifying bacteria RAS-L11. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 This is a photo of the enrichment of fresh activated sludge collected in LB medium;

[0017] Figure 2 The colony morphology characteristics of the obtained strain RAS-L11 are shown;

[0018] Figure 3 is the scanning electron micrograph of strain RAS-L11;

[0019] Figure 4 is the phylogenetic tree of strain RAS-L11;

[0020] Figure 5 is the cell growth curve of strain RAS-L11;

[0021] Figure 6 Aerobic denitrification characteristics of strain RAS-L11 at 25℃, C / N=6;

[0022] Figure 7 The short-range denitrification characteristics of strain RAS-L11 at 25℃, C / N=6;

[0023] Figure 8 The simultaneous nitrification and denitrification characteristics of strain RAS-L11 at 25℃ and C / N=6;

[0024] Fig. 9 is the growth curve of strain RAS-L11 in the presence of different concentrations of tetracycline;

[0025] Fig.10 The bar graph is a graph showing the concentration changes of nitrate nitrogen (in terms of nitrogen) of strain RAS-L11 at different time points in the presence of different concentrations of tetracycline;

[0026] Fig.11 It is a bar graph showing the concentration changes of total dissolved nitrogen (in terms of nitrogen) of strain RAS-L11 at different time points in the presence of different concentrations of tetracycline;

[0027] Fig.12 The growth curves of strain RAS-L11 in the presence of 10 mg / L ciprofloxacin, erythromycin, metronidazole and sulfamethoxazole, respectively;

[0028] Fig.13The line graphs are the concentration changes of nitrate nitrogen (in terms of nitrogen) of strain RAS-L11 at different time points in the presence of 10 mg / L ciprofloxacin, erythromycin, metronidazole and sulfamethoxazole;

[0029] Fig.14 : (a) is the removal rate of nitrate nitrogen by strain RAS-L11 in normal denitrification medium at different times and the growth curve of the strain, (b) is the removal rate of nitrate nitrogen by strain RAS-L11 in denitrification medium with 3% salinity at different times and the growth curve of the strain, (c) is the removal rate of nitrate nitrogen by strain RAS-L11 in denitrification medium with 5% salinity at different times and the growth curve of the strain, (d) is the removal rate of nitrate nitrogen by strain RAS-L11 in denitrification medium with 8% salinity at different times and the growth curve of the strain. DETAILED DESCRIPTION

[0030] In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with specific embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0031] In view of the fact that when the current biodegradation method is used to treat water bodies containing antibiotics or having a certain salinity, the microorganisms used are easily affected by the antibiotics contained in the water body or the high salinity, thereby resulting in a reduced aerobic denitrification efficiency of the microorganisms and a poor treatment effect, the present invention provides a Pseudomonas stutzeri aerobic denitrifying bacterium RAS-L11, which has a wide adaptability to antibiotics and salinity and can still maintain a high denitrification capacity in water bodies containing antibiotics or having a certain salinity.

[0032] Example 1

[0033] The screening and enrichment process of the aerobic denitrifying bacteria Pseudomonas stutzeri RAS-L11 provided by the present invention is as follows:

[0034] Fresh activated sludge samples were collected from a pharmaceutical wastewater treatment plant in Shijiazhuang City. 10 mL of fresh activated sludge was placed in a 250 mL conical flask containing 100 mL of LB culture medium (such as Figure 1As shown), enrichment culture was carried out in a constant temperature shaking box at 25°C at a speed of 90rpm, and the culture medium was replaced every 2 days for a total of 4 days. After the culture was completed, 5% of the suspension (7.5mL) was transferred to a 250mL conical flask containing 150mL of denitrification culture medium, and was still placed in a constant temperature shaking box at 25°C and a speed of 90rpm for culture. The volume ratio of the denitrification culture medium to the pharmaceutical wastewater (the total volume was calculated as 10) was prepared in accordance with 10 / 0, 9 / 1, 8 / 2, 7 / 3, 6 / 4, 5 / 5, 4 / 6, 3 / 7, and 2 / 8, respectively. All culture media were continuously digested at 121°C for 30min, and each concentration of culture medium was cultured for 3d, repeated twice, and acclimated for 54 days.

[0035] Take 1 mL of the suspension after acclimation and add it to 9 mL of buffer. -1 , 10 -2 , 10 -3 , 10 -4 dilute in multiples, then from 10 -4 Take 0.2 mL of the mixed solution of the concentration and transfer it to the solid denitrification medium. Use a sterile coating rod to evenly spread it on the plate, seal it and invert it to culture at 25°C for 2 days. When colonies are visible to the naked eye, streak them. After a single colony is isolated, streak it several times to obtain a purified single bacterium. The purified single bacterium is cultured at 25°C and then placed at 4°C for screening.

[0036] The isolated single colonies were inoculated into a conical flask containing 100 mL of LB medium for enrichment culture for 2 days. After the culture was completed, the bacterial liquid was poured into a sterilized 50 mL centrifuge tube, centrifuged at 6000 r / min, the supernatant was taken out, the obtained bacteria were mixed with the buffer solution and shaken well, 10 mL of the mixed solution was inoculated into a 250 mL conical flask containing 150 mL of denitrification medium, and single bacteria with a nitrate nitrogen removal rate of more than 90% were screened out.

[0037] 10 mL of bacterial suspension with a nitrate-nitrogen removal rate of more than 90% was taken and inoculated into a conical flask containing fresh denitrification culture medium. The culture was transferred three times and the acclimatization time was 9 days. Single bacteria with a nitrate-nitrogen removal rate of more than 90% were screened out, including strain RAS-L11.

[0038] In the above screening and separation process, the components of the culture medium used include:

[0039] LB medium: tryptone 10 g / L, yeast extract powder 5 g / L, NaCl 10 g / L, sterilize at 121°C for 30 min.

[0040] Denitrification medium: CH 3 COONa 4.69g / L, KNO3 0.722g / L, Vickers salt solution (including K 2 HPO 4 ·3H 2 O 0.75 g / L, NaH 2 PO 4 ·2H 2 O 0.25g / L, NaCl 0.12g / L, MnSO 4 ·H 2 O 0.01g / L, MgSO 4 7H 2 O 0.05g / L, FeSO 4 7H 2 O 0.01g / L), and sterilized by high pressure at 121°C for 30min. This denitrification medium is also applicable to the following examples.

[0041] Solid denitrification medium: CH 3 COONa 4.69g / L, KNO 3 0.722g / L, K 2 HPO 4 ·3H 2 O0.75g / L, NaH 2 PO 4 ·2H 2 O 0.25g / L, NaCl 0.12g / L, MnSO 4 ·H 2 O 0.01g / L, MgSO 4 7H 2 O 0.05g / L, FeSO 4 7H 2 O 0.01g / L, agar 20g / L, autoclave at 121℃ for 30min.

[0042] Buffer: NaCl 8g / L, KCl 0.2g / L, Na 2 HPO 4 12H 2 O 3.63g / L, KH 2 PO 4 0.24g / L, adjust pH to 7.4.

[0043] Example 2

[0044] Observe the morphology of strain RAS-L11 isolated and screened in Example 1, and record its morphological characteristics: the colony diameter is about 1-1.5 mm (such as Figure 2The colonies are milky white, slightly convex, moist and smooth, with regular edges, uniform texture, opaque, and easy to pick. The bacteria are long rod-shaped (as shown in the figure) under a scanning electron microscope. Figure 3 shown).

[0045] Further molecular identification of strain RAS-L11:

[0046] The genomic DNA of the strain RAS-L11 was extracted, and the PCR product was amplified using the universal primer 27F / 1492R for bacterial 16SrRNA gene amplification, and sent to Shanghai Meiji Biopharmaceutical Technology Co., Ltd. for sequence sequencing (the sequence is shown in SEQ ID NO.1), and compared in the NCBI database to analyze and identify its species relationship. It was found that the strain RAS-L11 was closest to Pseudomonas stutzeri and had the highest similarity (99.93%). Based on the above analysis, the strain was preliminarily identified as Pseudomonas stutzeri, and the taxonomic name was: Pseudomonas stutzeri RAS-L11, and its phylogenetic tree results were as follows: Figure 4 shown.

[0047] Pseudomonas stutzeri RAS-L11 was deposited in the General Microbiology Center of China Culture Collection Administration on May 20, 2024, with the deposit number CGMCC No. 30692, and the deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

[0048] Example 3

[0049] Growth curve of Pseudomonas stutzeri RAS-L11

[0050] The activated seed liquid of Pseudomonas stutzeri RAS-L11 was inoculated into 150 mL of fresh denitrification medium at a rate of 2%, and cultured at a constant temperature of 25 ° C and a rotation speed of 90 rpm. Samples were taken at 0 h, 6 h, 12 h, 15 h, 18 h, 21 h, 24 h, 27 h, 30 h, 33 h, 36 h, 42 h, 48 h, 57 h, 60 h, 66 h, 72 h, 81 h, 84 h, 90 h, 96 h, 108 h, 120 h, and 144 h to determine the OD of the strain. 600 With the culture time t as the horizontal axis, OD 600 The value is used to draw a growth curve on the vertical axis, such as Figure 5 shown.

[0051] Among them, the composition formula of fresh denitrification medium is: CH 3 COONa 2.05g / L, KNO3 0.722g / L, Vickers salt solution (including K 2 HPO 4 ·3H 2 O 0.75 g / L, NaH 2 PO 4 ·2H 2 O0.25g / L, NaCl 0.12g / L, MnSO 4 ·H 2 O0.01g / L, MgSO 4 7H 2 O 0.05g / L, FeSO 4 7H 2 O 0.01g / L), sterilized by high pressure at 121℃ for 30min.

[0052] It can be seen that the lag period is from 0 to 48 hours, which is the stage of bacteria adapting to the new environment. In this stage, the cell volume increases, the metabolism is active, the cells proliferate, and there are sufficient reserves of enzymes, energy and intermediate metabolites; the logarithmic growth period is from 48 to 108 hours, and the bacteria proliferate rapidly at a geometric rate; the stable period is from 108 to 144 hours, and the bacterial reproduction trend gradually decreases due to the consumption of nutrients in the culture medium and the accumulation of toxic metabolites; after 144 hours is the decline period, and with the development of the stable period, the bacterial reproduction rate is lower than the rate of cell autolysis and death.

[0053] Example 4

[0054] This example studies the aerobic denitrification characteristics of Pseudomonas stutzeri RAS-L11. The experimental temperature is 25°C, the medium C / N=6, and the steps are as follows:

[0055] The activated seed solution of Pseudomonas stutzeri RAS-L11 was inoculated into a fresh denitrification medium (the formula was the same as the denitrification medium in Example 3, wherein the mass concentration of nitrogen atoms was rounded off to 100 mg / L) at a rate of 5%, and cultured at 25°C and 90 rpm for 48 h. Samples were taken at 12 h, 15 h, 18 h, 24 h, 36 h, and 48 h to measure the OD 600 , nitrate nitrogen, ammonia nitrogen, nitrite nitrogen and total dissolved nitrogen concentrations. The results are as follows Figure 6 shown.

[0056] It can be seen that the removal rate of nitrate nitrogen by strain RAS-L11 in the denitrification medium with a nitrate nitrogen concentration (in terms of nitrogen) of 106.77 mg / L can reach 98% at 24 hours, and the concentration of nitrate nitrogen (in terms of nitrogen) at this time is 1.95 mg / L; moreover, the accumulation of nitrite nitrogen (in terms of nitrogen) reaches the highest (33.69 mg / L) at 15 hours, and then the concentration (in terms of nitrogen) is only 0.88 mg / L at 24 hours; the removal rate of dissolved total nitrogen also reaches 99% at 24 hours, indicating that the strain can quickly remove nitrate nitrogen and nitrite nitrogen accumulated during the removal process of nitrate nitrogen. It can be seen that strain RAS-L11 has good aerobic denitrification performance under the conditions of 25°C and C / N=6.

[0057] Example 5

[0058] This example studies the short-range denitrification characteristics of Pseudomonas stutzeri RAS-L11. The experimental temperature is 25°C, the medium C / N=6, and the steps are as follows:

[0059] The activated seed solution of Pseudomonas stutzeri RAS-L11 was inoculated into fresh short-range denitrification medium at a rate of 5%, and cultured at 25°C and 90 rpm for 72 h. Samples were taken at 12 h, 24 h, 36 h, 48 h, 60 h, and 72 h to measure OD. 600 , nitrite nitrogen and dissolved total nitrogen concentrations, the results are as follows Figure 7 As shown. The short-range denitrification culture medium formula used is: CH 3 COONa2.05g / L, NaNO 2 0.4929 g / L, Vickers salt solution (including K 2 HPO 4 ·3H 2 O 0.75 g / L, NaH 2 PO 4 ·2H 2 O 0.25g / L, NaCl 0.12g / L, MnSO 4 ·H 2 O 0.01g / L, MgSO 4 7H 2 O0.05g / L, FeSO 4 7H 2 O 0.01g / L), sterilized by high pressure at 121℃ for 30min.

[0060] It can be seen that the removal rate of nitrite nitrogen by strain RAS-L11 in the short-range denitrification medium with a nitrite nitrogen concentration (in terms of nitrogen) of 90.99 mg / L can reach 100% in 24 hours, and the nitrite nitrogen concentration at this time is 0.14 mg / L; the removal rate of total dissolved nitrogen reaches 99% in 24 hours, and the concentration is reduced to 0.33 mg / L. It can be seen that strain RAS-L11 has good short-range aerobic denitrification performance under the conditions of 25°C and C / N=6.

[0061] Example 6

[0062] This example studies the simultaneous nitrification and denitrification characteristics of Pseudomonas stutzeri RAS-L11. The experimental temperature is 25°C, the medium C / N=6, and the steps are as follows:

[0063] The activated seed solution of Pseudomonas stutzeri RAS-L11 was inoculated into fresh synchronous nitrification and denitrification medium at a rate of 5%, and cultured at 25°C and 90 rpm for 72 h. Samples were taken at 12 h, 24 h, 36 h, 48 h, 60 h, and 72 h to measure OD. 600 , nitrate nitrogen, ammonia nitrogen, nitrite nitrogen and total dissolved nitrogen concentrations. The results are as follows Figure 8 As shown. The formula of the synchronous nitrification and denitrification culture medium used is: CH 3 COONa 2.05g / L, (NH 4 ) 2 SO 4 0.236g / L, KNO 3 0.361g / L, Vickers salt solution (including K 2 HPO 4 ·3H 2 O 0.75 g / L, NaH 2 PO 4 ·2H 2 O 0.25g / L, NaCl 0.12g / L, MnSO 4 ·H 2 O 0.01g / L, MgSO 4 7H 2 O 0.05g / L, FeSO 4 7H 2 O 0.01g / L), sterilized by high pressure at 121℃ for 30min.

[0064] It can be seen that in the simultaneous nitrification and denitrification medium with ammonia nitrogen concentration of 49.55 mg / L and nitrate nitrogen concentration of 49.62 mg / L, the nitrate nitrogen concentration of strain RAS-L11 was reduced to 0.56 mg / L in 24 hours, and the removal rate could reach 99%; the ammonia nitrogen concentration was reduced to 0.52 mg / L in 48 hours, and the removal rate could reach 99%; the dissolved total nitrogen concentration was reduced to 0.72 mg / L in 48 hours, and the removal rate could reach 99%. It can be seen that strain RAS-L11 has good simultaneous nitrification and denitrification performance under the conditions of 25℃ and C / N=6.

[0065] Example 7

[0066] This example studies the tetracycline-resistant characteristics of Pseudomonas stutzeri RAS-L11:

[0067] The activated seed solution of Pseudomonas stutzeri RAS-L11 was inoculated into fresh denitrification medium containing 50 μg / L, 250 μg / L and 500 μg / L tetracycline (the formula is the same as the denitrification medium in Example 3) at a 5% inoculum amount, and cultured at 25°C and 90 rpm for 72 h. Samples were taken at 12 h, 18 h, 24 h, 36 h, 42 h, 48 h, 60 h and 72 h to measure OD 600 The concentrations of nitrate nitrogen and total dissolved nitrogen were as follows: Figures 9-11 shown.

[0068] It can be seen that compared with the control group, tetracycline antibiotics have an inhibitory effect on the growth rate of the strain ( Fig. 9 ), the higher the tetracycline concentration, the more obvious the inhibitory effect; and the greater the concentration of tetracycline, the greater its impact on the aerobic denitrification rate of the strain ( Fig.10 ), but it will not affect the final removal efficiency. At 72h, the removal rate of nitrate nitrogen and dissolved total nitrogen (in terms of nitrogen) in the culture medium containing different concentrations of tetracycline by strain RAS-L11 reached more than 99%. This shows that Pseudomonas stutzeri RAS-L11 can tolerate the toxicity of tetracycline antibiotics and perform aerobic denitrification, that is, the strain has a wide range of adaptability to tetracycline antibiotics.

[0069] Example 8

[0070] This example studies the resistance of Pseudomonas stutzeri RAS-L11 to quinolones, macrolides, nitroimidazoles and sulfonamide antibiotics:

[0071] The activated seed solution of Pseudomonas stutzeri RAS-L11 was inoculated into a fresh denitrification medium containing 10 mg / L of ciprofloxacin (quinolone), erythromycin (macrolide), metronidazole (nitroimidazole) and sulfamethoxazole (sulfonamide) at a concentration of 10 mg / L (the formula is the same as the denitrification medium in Example 3) at 25°C and 90 rpm for 96 h, and samples were taken at 24 h, 48 h, 72 h and 96 h to measure OD 600 and nitrate nitrogen concentrations, the results are as follows Fig.12 , Fig.13 shown.

[0072] It can be seen that different types of antibiotics have a certain inhibitory effect on the growth of strains ( Fig.12 ), among which ciprofloxacin had the greatest inhibitory effect on the growth of the strain in this experiment, followed by erythromycin, then sulfamethoxazole, and metronidazole had the least effect in this experiment; Fig.13 At 96h, the nitrate nitrogen concentration in the culture medium containing different types of antibiotics decreased from 103.92mg / L to 0mg / L, 1.03mg / L, 3.04mg / L and 0.03mg / L, respectively, and the removal rate was more than 97%. It can be seen that Pseudomonas stutzeri RAS-L11 can tolerate the toxicity of different types of antibiotics to carry out aerobic denitrification, that is, the strain has a wide range of antibiotic adaptability.

[0073] Example 9

[0074] This example studies the salt tolerance of Pseudomonas stutzeri RAS-L11:

[0075] The activated seed solution of Pseudomonas stutzeri RAS-L11 was inoculated into fresh denitrification medium with NaCl mass fractions of 0%, 3%, 5% and 8% respectively (the formula was the same as the denitrification medium in Example 3, respectively recorded as 0% salinity, 3% salinity, 5% salinity and 8% salinity), and cultured at 25°C and 90rpm for 120h (312h for 8% salinity), and samples were taken at intervals of 24h (except for 8% salinity) to determine the concentrations of nitrate nitrogen, ammonia nitrogen and nitrite nitrogen (all in terms of nitrogen). The results are shown in Table 1 (the results were rounded to two decimal places), and the removal rate of nitrate nitrogen (in terms of nitrogen) in denitrification medium of different salinities at different times and the growth of the strain were plotted, as shown in Table 1. Fig.14 shown.

[0076] It can be seen that under the influence of 3% salinity, the nitrate nitrogen removal rate of Pseudomonas stutzeri RAS-L11 can reach 93% at 48h, and the concentration (in terms of nitrogen) decreases from the initial 101.58mg / L to 7.14mg / L; under the influence of 5% salinity, the nitrate nitrogen removal rate of the strain reaches 99% at 96h, and the concentration (in terms of nitrogen) decreases from 101.58mg / L to 0.19mg / L; 8% salinity has a greater impact on the aerobic denitrification process of the strain, but the nitrate nitrogen removal rate can reach more than 20% at 72h, and the nitrate nitrogen removal rate reaches 85% at 312h, and the concentration (in terms of nitrogen) decreases from the initial 101.58mg / L to 15.17mg / L. It can be seen that the strain RAS-L11 can adapt to higher salinities than other salt-tolerant bacteria and has a wider range of salinity adaptability.

[0077] Table 1

[0078]

[0079] What is described above is only a preferred specific implementation manner of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can make equivalent replacements or changes according to the technical scheme and inventive concept of the present invention within the technical scope disclosed by the present invention, which should be covered by the protection scope of the present invention.

Claims

1. A salt-tolerant and drug-tolerant Pseudomonas stutzeri aerobic denitrifying bacterium, characterized in that: The strain was named RAS-L11 and classified as Pseudomonas stutzeri ( Stutzerimonas stutzeri ), deposited in the General Microbiology Center of China Microorganism Culture Collection Administration, the deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, the deposit date is May 20, 2024, and the deposit number is CGMCC No.30692.

2. Use of the salt-tolerant and drug-tolerant Pseudomonas stutzeri aerobic denitrifying bacteria according to claim 1 in denitrification of water bodies.

3. The use of the salt-tolerant and drug-tolerant Pseudomonas stutzeri aerobic denitrifying bacteria in water denitrification as claimed in claim 2, characterized in that: The denitrification includes removing nitrate nitrogen, ammonia nitrogen and / or nitrite nitrogen from the water body.

4. The use of salt-tolerant and drug-tolerant Pseudomonas stutzeri aerobic denitrifying bacteria in water denitrification as claimed in claim 2, characterized in that: The salinity of the water body is 0% to 8%.

5. The use of the salt-tolerant and drug-tolerant Pseudomonas stutzeri aerobic denitrifying bacteria in water denitrification as claimed in claim 2, characterized in that: The water bodies include pharmaceutical wastewater, domestic sewage, aquaculture wastewater and industrial wastewater.

6. The use of salt-tolerant and drug-tolerant Pseudomonas stutzeri aerobic denitrifying bacteria in water denitrification as claimed in claim 2, characterized in that: The water body contains at least one of tetracycline, ciprofloxacin, erythromycin, metronidazole and sulfamethoxazole.

7. The use of salt-tolerant and drug-tolerant Pseudomonas stutzeri aerobic denitrifying bacteria in water denitrification as claimed in claim 2, characterized in that: The carbon-nitrogen ratio of the water body is not less than 4.

8. A microbial agent, characterized in that: It includes the salt-resistant and drug-resistant Pseudomonas stutzeri aerobic denitrifying bacteria RAS-L11 described in claim 1.

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