Thauera with denitrification capability as well as preparation method and application of thauera

Abstract

The invention discloses thauera with denitrification capacity and application of the thauera, the strain is preserved in the China Center for Type Culture Collection, the preservation number is CCTCC M 2024703, and the strain is named as TES. The strain can be subjected to enrichment culture under different environmental conditions, screening and domestication are performed, enrichment culture is diluted after 3-5 times of enrichment culture, after further culture is performed until macroscopic bacterial colonies are obtained, a pure strain is picked and subjected to plate streak separation, the pure strain is further prepared into a liquid microbial inoculum, the liquid microbial inoculum is inoculated into wastewater containing ammonia nitrogen, nitrate or nitrite, and the wastewater containing ammonia nitrogen, nitrate or nitrite is obtained. The strain grows and reproduces and plays a role in denitrification through culture, so that the content of ammonia nitrogen, nitrate or nitrite in wastewater is reduced, experimental conclusions prove that the denitrification treatment effect on treated wastewater in a high-aeration tank of a pharmaceutical wastewater and sewage treatment plant and landfill leachate of a refuse landfill is remarkable, and the nitrogen removal effect on the treated wastewater in the high-aeration tank of the pharmaceutical wastewater and sewage treatment plant and the landfill leachate of the refuse landfill is remarkably improved under the condition of normal temperature. The treatment effect is more obvious and efficient.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of environmental microbial application technology, and particularly relates to a bacterium with denitrification ability and biological activity, and also relates to a method for culturing the bacterium and application of the bacterium in nitrogen degradation in water bodies. BACKGROUND

[0002] Nitrogen in water bodies mainly exists in the form of organic nitrogen, ammonia nitrogen and nitrate nitrogen, and is one of the most common pollutants that cause water eutrophication, harm human health and destroy the ecological environment of water bodies. The discharge of agricultural nitrogen fertilizer, industrial wastewater and urban wastewater is the main source of nitrogen in water bodies. How to economically and efficiently remove nitrogen pollution in water bodies has become a research focus and hotspot in the field of water pollution control.

[0003] Biological denitrification is an environmentally friendly and economical method for wastewater denitrification, which uses specific microorganisms under suitable environmental conditions to convert nitrogen elements in wastewater into harmless substances. Thauera is a bacterium widely existing in natural environment, which has high denitrification ability. However, different Thauera strains have different denitrification performance and application effects

[0004] Traditional biological denitrification is divided into two relatively independent processes, namely nitrification process and denitrification process. Due to the different requirements of (aerobic) nitrification and (anaerobic) denitrification for dissolved oxygen, it is difficult to realize the synchronization and unity of the whole denitrification process, resulting in the disadvantages of long wastewater treatment process, huge investment in treatment facilities and equipment, high energy consumption and operation and maintenance cost, which greatly hinders the improvement of biological denitrification efficiency. The discovery of heterotrophic nitrification and aerobic denitrification changes the traditional cognition of biological denitrification theory and provides potential upward space for the new development of biological denitrification technology. The use of aerobic denitrification can realize the unity of nitrification process and denitrification process in time and space, and aerobic denitrification has the advantages of realizing simultaneous nitrification and denitrification, compensating for the alkalinity consumed by nitrification reaction and strong adaptability of bacterial flora. Aerobic denitrifying bacteria are widely distributed, and these bacteria are mainly isolated from activated sludge, soil and sediment, wastewater and wastewater treatment system, and lakes and other natural environments.

[0005] Therefore, in order to improve the efficiency and effect of wastewater denitrification treatment, it is necessary to screen and study Thauera with high denitrification ability, and to explore its possibility in practical application. The Thauera strain with denitrification ability obtained by screening can play an important role in wastewater treatment, improve the efficiency and effect of wastewater denitrification treatment, and make contributions to the development of environmental protection and wastewater treatment field. SUMMARY

[0006] The first object of the present application is to provide a Thauerabutanivorans with high nitrogen removal efficiency, which can remove more nitrogen compounds in a short time at room temperature and improve the treatment efficiency.

[0007] The present application also provides a preparation method of the Thauerabutanivorans, which comprises inoculating the Thauerabutanivorans TES strain into LB culture medium and culturing to the logarithmic growth phase, removing the supernatant of the obtained bacterial suspension by centrifugation, and washing with sterile water to obtain the Thauerabutanivorans TES strain mother liquor.

[0008] The third object of the present application is to provide an application of the Thauerabutanivorans TES in the nitrogen removal treatment of nitrogen-containing wastewater.

[0009] The extraction process of the Thauerabutanivorans TES with high nitrogen removal efficiency comprises the following steps:

[0010] A, take the perfume wastewater into a 250mL conical flask, and add liquid heterotrophic nitrification medium into the conical flask, and culture at a temperature of 30℃ and a shaking speed of 100r / min for 24-72 hours; monitor the change of ammonia nitrogen concentration during the culture process, and when no ammonia nitrogen is detected, take 1% of the culture to transfer to fresh liquid heterotrophic nitrification medium for the next round of enrichment culture.

[0011] The liquid heterotrophic nitrification medium contains the following components: ammonium chloride 0.1-6.0g / L, magnesium sulfate 0.18-0.42g / L, potassium hydrogen phosphate 0.08-0.8g / L, sodium bicarbonate 0.4-1.2g / L, anhydrous sodium acetate 2-10g / L, ferrous sulfate heptahydrate 0.02-0.08g / 10mL, sodium chloride 30-100g / L, trace elements 1mL, and distilled water 1000mL. Among them, the ferrous sulfate heptahydrate is added to the liquid heterotrophic nitrification medium in proportion after sterilization;

[0012] The liquid heterotrophic nitrification medium can be made into a solid heterotrophic nitrification medium by adding 1%-2% mass ratio of agar powder.

[0013] The trace elements comprise the following components: copper sulfate 0.025-0.13 g / L, zinc sulfate 0.15-0.85 g / L, cobalt chloride 0.25-0.869 g / L, manganese chloride 0.25-0.90 g / L, ethylenediaminetetraacetic acid 0.1-1.3 g / L, sodium molybdate 0.02-0.30 g / L, boric acid 0.01-0.25 g / L, calcium carbonate 0.02-0.50 g / L.

[0014] B, after 3-5 rounds of enrichment culture, the enrichment culture obtained in step A is diluted and plated on the same new heterotrophic nitrification solid medium as described in step A, and after the visible colonies are cultured, single colonies are picked for plate streaking separation, and after 3-5 streaking, the pure strain is separated.

[0015]

[0016] The colony morphology characteristics of the Tolumonas TES strain are that the colony of the Tolumonas TES strain on an agar plate is round, the colony diameter is large, the surface is smooth, the colony is cushion-shaped, is opaque, and the edge is complete.

[0017] The physiological and biochemical characteristics are that the Tolumonas TES strain belongs to gram-negative bacteria, can survive under aerobic and anaerobic conditions, and is positive in oxidation enzyme, contact enzyme, starch hydrolysis, citrate, nitrate reduction experiments, and is negative in acetic acid oxidation, pectin hydrolysis, gelatin liquefaction and lactose oxidation fermentation.

[0018] The application of a Tolumonas TES strain with denitrification ability in the denitrification treatment of nitrogen-containing wastewater is provided, and the steps are as follows:

[0019] A. The Tolumonas TES strain with high-efficiency denitrification is inoculated into the above-mentioned liquid heterotrophic nitrification culture medium to culture to the logarithmic growth phase, the obtained bacterial suspension is centrifuged to remove the supernatant, and then washed with sterile physiological saline to obtain a Tolumonas TES strain mother liquor.

[0020] B. The Tolumonas TES strain mother liquor prepared in step A is added to the liquid heterotrophic nitrification culture medium to perform fermentation culture, and a Tolumonas TES liquid bacterial agent is obtained.

[0021] C. The liquid bacterial agent obtained by fermentation in step B is added to various salinity, various C / N, various temperature and different types of high-salt nitrogen-containing wastewater to be treated, and high-efficiency denitrification can be achieved.

[0022] On the basis of the above technical solutions,

[0023] Preferably, the liquid heterotrophic nitrification culture medium contains the following components: ammonium chloride 0.38 g / L, magnesium sulfate 0.2 g / L, dipotassium hydrogen phosphate 0.1 g / L, sodium bicarbonate 1 g / L, anhydrous sodium acetate 3.4 g / L, ferrous sulfate heptahydrate 0.050 g / 25 mL, sodium chloride 2 g / L, trace elements 1 mL, distilled water 1000 mL, and pH 7-8.

[0024] The trace elements contain the following components: copper sulfate 0.12 g / L, zinc sulfate 0.80 g / L, cobalt chloride 0.80 g / L, manganese chloride 0.80 g / L, ethylenediaminetetraacetic acid 1.0 g / L, sodium molybdate 0.20 g / L, boric acid 0.20 g / L, and calcium carbonate 0.3 g / L.

[0025] Compared with the prior art, the present application has the following advantages and effects:

[0026] The Thauera TES bacteria with denitrification capacity provided by the application can realize efficient denitrification. The bacteria can grow to OD600 of 2.35 in 48 hours in wastewater with ammonia nitrogen concentration of 100 mg / L-1000 mg / L, and the ammonia nitrogen removal rate can reach 98.28%, and the total nitrogen removal rate can reach 90.83%. The strain can grow to OD600 of 1.65 in 36 hours under 0.2%-4% salinity, and the ammonia nitrogen removal rate can reach more than 90%, and the total nitrogen removal rate can reach more than 86%. Compared with traditional biological denitrification technology, the Thauera TES bacteria can remove more nitrogen compounds in a shorter time, and improve the processing efficiency. The Thauera TES bacteria denitrification technology reduces energy consumption and chemical consumption by optimizing reaction conditions and microbial population. This technology can operate at normal temperature and general pressure, reduce energy consumption, and reduce the amount of chemicals used, thereby reducing the operating cost. The Thauera TES bacteria denitrification is suitable for different types of wastewater treatment, including domestic sewage, industrial wastewater, etc., and can adapt to different water quality and treatment requirements, and has strong applicability. BRIEF DESCRIPTION OF DRAWINGS

[0027] Figure 1 It is a colony morphology diagram of Thauera TES bacteria on agar medium;

[0028] Figure 2 It is a denitrification efficiency diagram of Thauera TES bacteria under different salinity;

[0029] Figure 3 It is a denitrification efficiency diagram of Thauera TES bacteria under different temperature;

[0030] Figure 4 It is a denitrification efficiency diagram of Thauera TES bacteria under different pH;

[0031] Figure 5 It is a denitrification efficiency diagram of Thauera TES bacteria under different rotation;

[0032] Figure 6 It is a denitrification efficiency diagram of Thauera TES bacteria under different carbon source;

[0033] Figure 7 It is a denitrification efficiency diagram of Thauera TES bacteria under different nitrogen source;

[0034] Figure 8 It is a denitrification test effect diagram of Thauera TES bacteria in medical wastewater (30℃);

[0035] Figure 9 It is a denitrification test effect diagram of Thauera TES bacteria in landfill leachate (30℃).

[0036] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the following will briefly introduce the drawings needed to be used in the embodiments or prior art description. Obviously, the drawings in the following description are only some embodiments of the present application, and for those skilled in the art, other drawings can also be obtained without creative labor on the basis of these drawings. DETAILED DESCRIPTION

[0037] The technical solutions in the embodiments of the present application will be described clearly and completely in combination with the embodiments of the present application. Obviously, the described embodiments are only some embodiments of the present application, but not all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative labor are within the scope of protection of the present application.

[0038] Embodiment 1: Isolation of Thauera TES strain

[0039] The present application provides a denitrifying bacteria which denitrifies through heterotrophic nitrification-aerobic denitrification pathway. The strain is classified and named as Thauera butanivorans, and the strain number is TES; the strain was preserved in China Center for Type Culture Collection on April 18, 2024, and the preservation number is CCTCC M 2024703. Hereinafter, it is referred to as TES strain.

[0040] The strain is isolated from perfume wastewater by the inventors in 2023.

[0041] A preparation method of Thauera TES strain, the steps of which are:

[0042] I. Isolation of the strain

[0043] (1) Preparation of culture medium:

[0044] The liquid heterotrophic nitrification culture medium contains the following components (g / L) in mass / volume ratio: ammonium chloride 0.38 g / L, magnesium sulfate 0.2 g / L, potassium phosphate dibasic 0.1 g / L, sodium bicarbonate 1 g / L, anhydrous sodium acetate 3.4 g / L, ferrous sulfate heptahydrate 0.050 g / 25 mL, sodium chloride 2 g / L, trace elements 1 mL, distilled water 1000 mL, pH 7-8. Among them, the ferrous sulfate heptahydrate is added to the liquid heterotrophic nitrification culture medium in proportion after sterilization.

[0045] The solid culture medium is prepared by adding 1-2% of agar powder in mass percentage on the basis of the aforementioned liquid heterotrophic nitrification culture medium.

[0046] The trace elements contain the following components: copper sulfate 0.12 g / L, zinc sulfate 0.80 g / L, cobalt chloride 0.80 g / L, manganese chloride 0.80 g / L, ethylenediaminetetraacetic acid 1.0 g / L, sodium molybdate 0.20 g / L, boric acid 0.20 g / L, calcium carbonate 0.3 g / L.

[0047] (2) Isolation method:

[0048] The flavor wastewater is taken into a 250 mL conical flask, and the heterotrophic nitrification medium is added to the conical flask. The screening and domestication are carried out at a temperature of 30°C, a salinity of 0.2% (calculated by NaCl), and 100 r / min for 24-72 hours, and the concentration of ammonia nitrogen is monitored. When the concentration of ammonia nitrogen cannot be monitored, the sample is inoculated into the heterotrophic nitrification medium for enrichment culture. After 3-5 times of enrichment culture, the enrichment culture is diluted and coated on a solid culture medium. After the culture reaches the visible colony, a single colony is picked for plate streaking separation. After 3-5 times of streaking, the Thauera TES pure strain is separated.

[0049] II. Strain identification:

[0050] 1. Bacterial morphological observation:

[0051] The Thauera TES strain is cultured at a temperature of 30°C, a salinity of 0.2% (calculated by NaCl), and 100 r / min to the logarithmic phase. After the bacterial solution is diluted, it is uniformly coated on a solid plate and cultured in a biochemical incubator at 30°C. After a single colony forms on the plate, observation is carried out, and gram staining microscopic examination is performed.

[0052] 2. Colony morphological characteristics:

[0053] The colony of the Thauera TES strain on the agar plate is milky white, opaque, with a protruding periphery and a sunken middle, a smooth surface, and a neat edge. The results are shown in Table 1. Figure 1

[0054] 3. Strain molecular identification:

[0055] The 16S rRNA is amplified by using the bacterial genome as a DNA template and 27F (5'-AGAGTTTGATCCTGGCTCAG-3') and 1492R (5'-GGTTACCTTGTTACGACTT-3') as the upper and lower primers. The PCR reaction conditions are as follows: 94°C pre-denaturation for 4 min; 94°C denaturation for 25 s, 58°C annealing for 30 s, 72°C extension for 30 s, a total of 25 cycles; and 72°C final extension for 5 min. After amplification, the PCR product is subjected to 1% agarose gel electrophoresis to detect the integrity of the PCR product. Finally, the PCR product is sent to the sequencing department of GenScript Biotech (Shanghai) Co., Ltd. for sequencing.​

[0056] The obtained 16S rDNA gene of the bacterial strain has an effective length of about 1443 bp nucleotide sequence, and the 16S rDNA sequence is shown as SEQ ID NO: 1. The sequence is input into GenBank. The Blast software is used for comparison analysis with the database sequence, and the result shows that the similarity with the 16S rDNA sequence of Thauera butanivorans is high, which is 99.2%. Based on the phylogenetic analysis result of the 16S rDNA gene and the physiological and biochemical characteristics, it is identified as a new bacterial strain of Thauera butanivorans, named as TES; and the bacterial strain is preserved in China Center for Type Culture Collection, and the preservation number is CCTCC M2024703.

[0057] Example 2: Bacterial strain test

[0058] (1) Denitrification effect of Thauera TES bacterial strain under different salinity:

[0059] At a temperature of 30℃, the TES bacterial suspension is inoculated into a 250 mL triangular flask containing 100 mL of the above-mentioned heterotrophic nitrification medium at a 1‰ inoculation ratio, and is cultured under different salinity (calculated by NaCl) values (0%, 0.2%, 1%, 2%, 4%, 6%). During the culture process, the shaking speed is set to 100 r / min. In addition, the culture solution is sampled regularly during the culture process, and the ammonia nitrogen concentration is detected.

[0060] Under the salinity of 0.2%, the Thauera TES bacterial strain can remove 99.87% of the ammonia nitrogen and 64.94% of the total nitrogen; under the salinity of 1% and 2%, the ammonia nitrogen can also be removed by more than 99%, and the total nitrogen can also be removed by more than 63.54%; under the salinity of 4% and 6%, the removal rate of ammonia nitrogen is only about 30%, and the removal rate of total nitrogen is about 28%. The experimental results are shown in Table 1. Figure 2

[0061] (2) Denitrification effect of Thauera TES bacterial strain under different temperatures:

[0062] Under the salinity of 0.2% (calculated by NaCl), the TES bacterial suspension of Thauera is inoculated into a 250 mL triangular flask containing 100 mL of the above-mentioned heterotrophic nitrification medium at a 1‰ inoculation ratio, and is cultured under different temperature values (10℃, 20℃, 30℃, 40℃). During the culture process, the shaking speed is set to 100 r / min. In addition, the culture solution is sampled regularly during the culture process, and the ammonia nitrogen concentration is detected.

[0063] ​The Thauera TES strain can remove 85.54% of ammonia nitrogen in a 20℃ environment, and the removal rate of ammonia nitrogen can reach more than 99% at 30℃ and 40℃. The strain cannot grow at 10℃. The experimental results are shown in Figure 3 .

[0064] (3) Denitrification effect of Thauera TES strain under different pH values:

[0065] At a temperature of 30℃, a salinity of 0.2% (calculated by NaCl), and a 1‰ inoculation ratio, Thauera TES strain bacterial suspension was inoculated into a 250mL triangular flask containing 100mL of the above-mentioned heterotrophic nitrification medium, and cultured under different pH values (4, 6.5, 7.5, 8.5, 10). During the culture process, the shaking speed was set to 100r / min. In addition, samples were taken regularly during the culture process to detect the ammonia nitrogen concentration of the culture solution.

[0066] The removal rate of ammonia nitrogen was more than 99.00% under pH values of 7.5, 8.5 and 10, and the removal rate of nitrogen could reach about 82.05% under a pH value of 6.5. Thauera TES strain did not grow in an acidic environment with a pH value of 4. The experimental results are shown in Figure 4 .

[0067] Denitrification effect of Thauera TES strain under different rotation speeds:

[0068] At a temperature of 30℃, a salinity of 0.2% (calculated by NaCl), and a 1‰ inoculation ratio, Thauera TES strain bacterial suspension was inoculated into a 250mL triangular flask containing 100mL of the above-mentioned heterotrophic nitrification medium, and cultured under different rotation speed values (0r / min, 50r / min, 75r / min, 100r / min, 150r / min). In addition, samples were taken regularly during the culture process to detect the ammonia nitrogen concentration of the culture solution.

[0069] The removal rate of ammonia nitrogen could reach 51.65% at a rotation speed of 0r / min, and the removal rate of ammonia nitrogen by Thauera TES strain increased greatly with the increase of rotation speed. When the rotation speed was 100r / min, the removal rate could reach 99%, and when the rotation speed increased to 150r / min, the removal rate of ammonia nitrogen reached 100%. The experimental results are shown in Figure 5 .

[0070] Denitrification effect of Thauera TES strain under different carbon sources:

[0071] Inoculate the Thauera Strain into the 250 mL flask containing 100 mL of the above-mentioned heterotrophic nitrification medium at a 1‰ inoculation ratio at a temperature of 30℃ and a salinity of 0.2% (calculated as NaCl), and cultivate under different carbon sources (anhydrous sodium acetate, sodium citrate, sodium succinate, sucrose, glucose), with the rotation speed of the shaker set to 100 r / min. In addition, periodically take samples during the cultivation process to detect the ammonia nitrogen concentration of the culture solution.

[0072] The removal rate of ammonia nitrogen can reach more than 99% when using anhydrous sodium acetate, sodium citrate, and sodium succinate as the carbon source, and the removal rate of total nitrogen can reach more than 50%. The experimental results are shown in Table 1. Figure 6

[0073] Denitrification effect of Thauera Strain under different nitrogen sources:

[0074] Based on the heterotrophic nitrification medium without the ammonium chloride component, different nitrogen sources (NH4+-N, NO3--N, NO2--N) are supplemented to test the denitrification effect of the Thauera Strain on different nitrogen sources. Inoculate the Thauera Strain into the 250 mL flask containing 100 mL of the above-mentioned heterotrophic nitrification medium at a 1‰ inoculation ratio at a temperature of 30℃ and a salinity of 0.2% (calculated as NaCl), and cultivate under different nitrogen sources, with the rotation speed of the shaker set to 100 r / min. In addition, periodically take samples during the cultivation process to detect the ammonia nitrogen, nitrite nitrogen, nitrate nitrogen, and total nitrogen concentrations of the culture solution. The removal rate of ammonia nitrogen can reach 100% when using ammonia nitrogen as the sole nitrogen source; the Thauera Strain can remove 80.84% of NO3--N when using nitrate nitrogen as the sole nitrogen source; and the Thauera Strain can remove 90.63% of NO2--N when using nitrite nitrogen as the sole nitrogen source. The experimental results are shown in Table 2. Figure 7

[0075] Example 3: Application of Thauera Strain in Pharmaceutical Wastewater

[0076] A preparation method of a Thauera TES bacterial agent and its application in pharmaceutical wastewater, the steps of which are as follows:

[0077] A. Inoculate the Thauera Strain into the liquid heterotrophic nitrification medium and cultivate to the logarithmic growth phase. Centrifuge the obtained bacterial suspension to remove the supernatant, and then wash with sterile physiological saline to obtain the Thauera Strain mother liquor.

[0078] B. Prepare the liquid heterotrophic nitrification medium, which needs to be sterilized in a high-pressure sterilization pot at 103.4 kpa and 121℃ for 20 min, and then cooled to room temperature (20-25℃).

[0079] ​​C. To the liquid heterotrophic nitrification medium prepared in step B after cooling, the Tolumes TES strain mother liquor obtained in step A is added, and culture is carried out in a shaking incubator, and when the culture OD600 reaches 1.2, the Tolumes TES liquid inoculum is obtained.

[0080] D. The TES liquid inoculum obtained in step C is added to the corresponding medical wastewater.

[0081] 5L of wastewater is collected from the high-exposure tank of the wastewater treatment system of the Hubei Xiangyang Huazhong Pharmaceutical Industrial Park, and the initial water quality parameters of the wastewater are: ammonia nitrogen 480-510 mg / L, total nitrogen 532 mg / L, COD 1000 mg / L, and salinity 0.6%. The wastewater is divided into three 1L plastic cups, wherein group A is added with 5% bacterial liquid, group B is added with 10% bacterial liquid, and group C is not added with bacterial liquid, and is placed at room temperature, while being aerated with a microporous sand core air head at a ventilation rate of 180 mL / min; during the test, the ammonia nitrogen change of the system is regularly detected to analyze the denitrification ability of the strain.

[0082] The denitrification test effect of the Tolumes TES inoculum on medical wastewater is shown in Figure 8 After 7 days, the ammonia nitrogen removal rate of group A reaches 58.94%, the ammonia nitrogen removal rate of group B is 85.49%, and the ammonia nitrogen removal rate of group C is 26.48%.

[0083] Example 4: Application of Tolumes TES inoculum in landfill leachate wastewater

[0084] 5L of landfill leachate after preliminary treatment is taken from a landfill in Wuhan, Hubei, and the average initial water quality parameters are: ammonia nitrogen 2800-3200 mg / L.

[0085] The landfill leachate is diluted to about 1000 mg / L, and the wastewater is divided into three 3L triangular flasks, three of which are added with 5% Tolumes TES liquid inoculum, and the other three are used as a control group (CK) and placed at 30°C for treatment; the shaking speed is 100 r / min; during the test, the ammonia nitrogen change of the system is regularly detected to analyze the aerobic denitrification ability of the strain.

[0086] The liquid bacterial liquid is inoculated into the landfill leachate and placed at 30°C for 5 days, as shown in Figure 9 The initial ammonia nitrogen of the inoculated group (TES) and the control group (CK) is 1112.5 mg / L. The ammonia nitrogen removal rate of the inoculated group (TES) reaches 45.61%, and the ammonia nitrogen removal rate of the control group (CK) is 9.8%.

[0087] Therefore, it is shown that the strain of the present application has good adaptation to the whole temperature zone and has good aerobic nitrification and denitrification ability.

[0088] The above merely provides the preferred embodiment of the present application, and is not used to limit the present application. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A Thauera bacteria, classified as Thauera butanivorans, deposited in China Center for Type Culture Collection, with the accession number CCTCC M 2024703, and named as TES.

2. The Taura's bacillus according to claim 1, characterized by, The Thauera bacteria comprise a 16s rDNA sequence as shown in SEQ ID NO.

1.

3. The bacterium of claim 1, wherein the bacterium is a Clostridium sp. that is, The Thauera bacteria have a denitrification ability, and can reduce ammonia nitrogen, nitrate and nitrite of a certain concentration to nitrogen in a short time.

4. The bacterium according to claim 1, wherein The Thauera bacteria strain has the following biological characteristics: The colony surface is milky white, opaque, with a smooth surface and a neat edge, and the gram staining reaction is negative.

5. An inoculant characterized in that, The bacterial agent comprises the Thauera bacteria according to any one of claims 1-4, and the OD600 value of the bacterial agent is greater than 1.

2.

6. The method of claim 5, wherein the bacterial agent is prepared by the steps of: The method comprises the following steps: A. The Thauera bacteria strain is inoculated into a liquid heterotrophic nitrification culture medium to the logarithmic growth phase, the obtained bacterial suspension is centrifuged to remove the supernatant, and then washed with sterile normal saline to obtain a Thauera bacteria strain mother liquor; B. The liquid heterotrophic nitrification culture medium is sterilized in a high-pressure sterilization pot at 103.4 kPa and 121℃ for 20 min before use, and then cooled to room temperature; C. The Thauera bacteria strain mother liquor is added to the cooled liquid heterotrophic nitrification culture medium prepared in step B, and cultured in a shaking incubator, and the Thauera bacteria liquid bacterial agent is obtained when the OD600 value of the culture reaches 1.

2.

7. The method of claim 6, wherein the bacterial agent is prepared by, The liquid heterotrophic nitrification culture medium comprises the following components in a mass / volume ratio: 0.1-6.0 g / L of ammonium chloride, 0.18-0.42 g / L of magnesium sulfate, 0.08-0.8 g / L of dipotassium hydrogen phosphate, 0.4-1.2 g / L of sodium bicarbonate, 2-10 g / L of anhydrous sodium acetate, 0.02-0.08 g / 10 mL of ferrous sulfate heptahydrate, 30-100 g / L of sodium chloride, 1 mL of trace elements, 1000 mL of distilled water, and pH 7-8, wherein the ferrous sulfate heptahydrate is added to the liquid heterotrophic nitrification culture medium after sterilization in a proportion. The trace elements comprise the following components in a mass / volume ratio: 0.025-0.13 g / L of copper sulfate, 0.15-0.85 g / L of zinc sulfate, 0.25-0.869 g / L of cobalt chloride, 0.25-0.90 g / L of manganese chloride, 0.1-1.3 g / L of ethylenediaminetetraacetic acid, 0.02-0.30 g / L of sodium molybdate, 0.01-0.25 g / L of boric acid, and 0.02-0.50 g / L of calcium carbonate. 8.The Thauera bacteria according to any one of claims 1-4 are applied to wastewater treatment.

9. Use according to claim 8, characterized in that, The wastewater comprises medical wastewater and landfill leachate.

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