A beautiful Halomonas strain with anaerobic half-denitrification function and its screening method and application

Abstract

The present invention discloses a beautiful Halomonas with anaerobic half-denitrification function and a screening method and application thereof, relating to the field of environmental protection microbial technology. The strain was deposited in the General Microbiological Center (CGMCC) of the China Microbiological Culture Collection Administration on April 25, 2024, with the deposit number CGMCC No. 30437. The strain BW‑01 was isolated from anaerobic heterotrophic denitrification sludge acclimated with seawater. It is a strain that uses organic matter for semi-heterotrophic denitrification under anaerobic conditions to reduce nitrate to nitrite. The denitrification efficiency is as high as 87% when the carbon source is peptone, the nitrogen source concentration is 100 mgN / L, C / N=3, and the salinity is 60 g / L. The strain reduces nitrate to nitrite, promotes the accumulation of nitrite, and is beneficial for the subsequent nitrite to participate in the autotrophic denitrification reaction with sulfide as the electron donor. It has the advantage of high coordination with desulfurization and denitrification, realizes synchronous desulfurization and denitrification, and can be used in environmental protection fields such as ship high-salt nitrogen-containing wastewater treatment.

Description

Technical Field

[0001] The invention relates to the technical field of microorganisms, in particular to a beautiful Halomonas strain with anaerobic half-denitrification function and a screening method and application thereof. Background Art

[0002] With the development of shipping economy, the pollutants discharged by ships during navigation are increasing day by day. X ) is one of the main pollutants in ship exhaust emissions. After washing, nitrate (NO3 - ) will lead to eutrophication of water bodies, reduce water quality, and excessive NO3 - It also affects the health of aquatic plants and animals. Ship sewage also contains NO3 - , and its discharge will also affect the marine water environment. The above ship wastewater also has the characteristics of high salinity, which greatly increases the difficulty of combined in-situ treatment of wastewater.

[0003] At present, the treatment methods for high-salinity and high-nitrogen wastewater from ships include physical methods such as membrane separation and ion exchange, chemical methods such as electrochemical reduction, and biological methods. Biological methods are widely used due to their advantages such as good economy, simple operation, and high treatment efficiency. The organic matter contained in ship wastewater is complex and difficult to degrade. It is not appropriate to use a single anaerobic process. Combining multiple processes can achieve the purpose of synergistic removal of multiple pollutants. Among them, anaerobic heterotrophic semi-denitrification can be used as a front-end treatment process to remove NO3 - Reduction to nitrite nitrogen (NO2 - ), promote NO2 - accumulation; subsequently, NO2 - Then it participates in the desulfurization and denitrification reaction and is finally converted into N2, achieving the requirement of complete removal of nitrogen-containing pollutants. However, high salt will have a strong inhibitory effect on microorganisms, resulting in low denitrification performance of high-nitrate nitrogen wastewater in a high-salt environment. If salt-tolerant bacteria that can treat high-salt nitrogen-containing wastewater are screened out and the optimal conditions for their degradation of pollutants are studied, it will play a vital role in treating high-salt nitrogen-containing wastewater from ships and maintaining the ecological balance of the ocean.

[0004] Therefore, providing a semi-anaerobic heterotrophic denitrifying bacteria that is tolerant to high salt and has high denitrification efficiency to participate in the combined process has important practical significance for the in-situ purification of high-salinity and nitrogen-containing wastewater from ships, and has broad application prospects. Summary of the invention

[0005] In view of the defects in the prior art, the present invention proposes a beautiful Halomonas with anaerobic semi-denitrification function. The taxonomic name of the beautiful Halomonas is Halomonas venusta BW-01, the preservation number is CGMCC No.30437, the preservation date is April 25, 2024, and the preservation address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

[0006] The biological characteristics of the Halomonas venusta BW-01 are:

[0007] (1) The Halomonas venusta BW-01 provided by the present invention can convert NO3 into - Directly reduced to NO2 through the denitrification process - , thereby achieving NO3 - The conversion of NO2 into a substrate for subsequent reactions does not involve NO2 - The conversion to N2 is a half-way denitrification process; the strain converts NO3 - Reduction to NO2 - , contributing to NO2 - The accumulation of NO2 - Participate in desulfurization and denitrification reactions as electron acceptors, using sulfide (S 2- ) is an electron donor and is eventually converted into N2 to achieve simultaneous desulfurization and denitrification. Therefore, this strain has the advantage of high compatibility with the desulfurization and denitrification process and can be applied in environmental protection fields such as high-salt nitrogen-containing wastewater treatment;

[0008] (2) The colonies are round, white, slightly raised in the middle, with intact, opaque edges;

[0009] (3) 16S rDNA gene sequencing:

[0010] The 16S rDNA gene sequence of Halomonas venusta BW-01 was amplified by PCR, sequenced and compared with BLAST. The 16S rDNA sequence is shown in SEQ ID NO: 1. The 16S rDNA gene sequence of Halomonas venusta BW-01 is highly similar to that of Halomonas venusta, with a similarity of 98.65%.

[0011] Another aspect of the present invention further provides a screening and culturing method for the Halomonas venusta BW-01, the screening method comprising the following steps:

[0012] From the anaerobic activated sludge with heterotrophic denitrification function, 10 ml of the sludge-water mixture is placed in 90 ml of liquid enrichment culture medium, and cultured in a constant temperature incubator at 100-150 rpm and 25-35°C for 48 hours; then the bacterial liquid is gradiently diluted and spread on the surface of the solid screening culture medium, and cultured at a constant temperature of 25-35°C for 48 hours. After the colonies grow on the solid culture medium, the colonies with clear plaques and larger diameters are selected for streaking separation, and the separation and purification are performed for 4-6 generations combined with microscopic observation until a purified single strain is obtained.

[0013] Furthermore, the culture medium for screening the Halomonas venusta BW-01 is prepared according to the following proportions:

[0014] The liquid enrichment medium: 0.9 g / L sodium nitrate, 1 g / L peptone, 1.5 g / L sodium bicarbonate, 1 L natural seawater, sterilized at 121° C. for 20 min;

[0015] The solid screening culture medium comprises: 0.9 g / L sodium nitrate, 1 g / L peptone, 1.5 g / L sodium bicarbonate, 12-14 g / L agar, 1 L natural seawater, and sterilized at 121° C. for 20 min.

[0016] In another aspect, the present invention provides the use of the beautiful Halomonas with anaerobic half-denitrification function in the treatment of high-salt nitrogen-containing wastewater from ships.

[0017] Furthermore, the high-salinity nitrogen-containing wastewater from ships is NO3 - Saline wastewater as a nitrogen source.

[0018] Furthermore, in the saline wastewater, peptone is the carbon source, the concentration of the nitrogen source is 100-250 mgN / L, C / N=2-3, and the salinity is 35-80 g / L.

[0019] The beneficial effects of the present invention are:

[0020] (1) The Halomonas venusta BW-01 of the present invention is isolated from anaerobic heterotrophic denitrification sludge domesticated with seawater, has a highly efficient salt-tolerant denitrification biological denitrification ability, and is a high-salt-tolerant biological denitrification strain of the genus Halomonas;

[0021] (2) The Halomonas venusta BW-01 provided by the present invention is different from the Halomonas venusta isolated in the past under aerobic conditions. The bacterium was isolated under anaerobic conditions and has the ability to withstand high-salinity denitrification under anaerobic conditions. Ship wastewater has a high salinity. Usually, denitrifying bacteria have low activity in high-salinity environments and the treatment effect is poor. However, this strain can maintain high activity in high-salinity environments and complete NO3 - This is of great significance for the anaerobic treatment of high-salinity nitrogen-containing wastewater from ships;

[0022] (3) The denitrification performed by Halomonas venusta BW-01 provided by the present invention is to convert NO3 - Reduction to NO2 - Half-way denitrification, easy to NO2 - The subsequent NO2 - Will participate in the desulfurization and denitrification reaction, and the relevant microorganisms will use NO2 - Sulfide (S 2- ) is an electron donor, and finally NO2 - Converted into N2. Therefore, the bacteria can promote the combined use of half-way denitrification and desulfurization and denitrification processes to achieve in-situ simultaneous removal of multiple pollutants from ships;

[0023] (4) The Halomonas venusta BW-01 provided by the present invention can utilize proteins, which are complex and difficult to degrade macromolecular organic matter. Under anaerobic conditions, the difficult-to-degrade organic matter is decomposed and converted into small molecular organic matter for use by other microorganisms. Therefore, the strain has the characteristic of working in coordination with other microorganisms, which is beneficial to the operation of the integrated wastewater treatment process of ships;

[0024] (5) The denitrification efficiency of Halomonas venusta BW-01 provided by the present invention is as high as 87%. High-salt wastewater is continuously produced during the voyage of ships, which occupies a large amount of limited ship space. The bacteria with high denitrification ability can promote the in-situ treatment of comprehensive wastewater, which can not only reduce the space pressure of ships during long-distance voyages, but also will not cause secondary pollution, and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0026] Figure 1 This is the colony morphology of strain BW-01 on the culture medium;

[0027] Figure 2 is the phylogenetic tree of strain BW-01;

[0028] Figure 3 The biological denitrification effect of strain BW-01 under different carbon source conditions; a is NO3 - The removal rate of NO2 - The production rate of bacteria, c is the OD of bacteria 600 value;

[0029] Figure 4 The biological denitrification effect of strain BW-01 under different nitrogen source concentrations; a is NO3 - The removal rate of NO2 - The production rate of bacteria, c is the OD of bacteria 600 value;

[0030] Figure 5 The biological denitrification effect of strain BW-01 under different carbon-nitrogen ratios C / N; a is NO3 - The removal rate of NO2 - The production rate of bacteria, c is the OD of bacteria 600 value;

[0031] Figure 6 The biological denitrification effect of strain BW-01 under different salinity conditions; a is NO3 - The removal rate of NO2 - The production rate of bacteria, c is the OD of bacteria 600 value;

[0032] Figure 7 This is the growth and degradation curve of strain BW-01 during anaerobic denitrification in high-salt nitrogen-containing wastewater from ships. DETAILED DESCRIPTION

[0033] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0034] The culture medium used in the examples of this application is as follows:

[0035] Liquid enrichment medium: sodium nitrate 0.9 g / L, peptone 1 g / L, sodium bicarbonate 1.5 g / L, 1 L natural seawater, sterilized at 121 °C for 20 min;

[0036] Liquid activation medium: The composition is the same as that of liquid enrichment medium;

[0037] Solid screening medium: sodium nitrate 0.9 g / L, peptone 1 g / L, sodium bicarbonate 1.5 g / L, agar 12-14 g / L, 1 L natural seawater, sterilized at 121 °C for 20 min;

[0038] Natural seawater denitrification medium: sodium nitrate 0.6-1.5g / L, peptone 0.6-5.4g / L, sodium bicarbonate 1.5g / L, 1L natural seawater, sterilized at 121℃ for 20min, where peptone can be replaced by sodium acetate, sodium propionate, sodium butyrate or glucose as a carbon source;

[0039] Artificial seawater denitrification medium: sodium nitrate 0.6 g / L, peptone 1.8 g / L, sodium bicarbonate 1.5 g / L, sodium chloride 35-120 g / L, magnesium chloride 2.26 g / L, magnesium sulfate 3.25 g / L, calcium chloride 1.15 g / L, potassium chloride 0.72 g / L, sodium bicarbonate 1.7 g / L, dissolved in ultrapure water, sterilized at 121°C for 20 min.

[0040] Embodiment 1:

[0041] (1) Enrichment culture:

[0042] Take 10 ml of the anaerobic activated sludge with heterotrophic denitrification function that has been acclimated with seawater. Pour 10 ml of the sludge-water mixture and 90 ml of the prepared enrichment culture medium into an anaerobic bottle (conical flask) on a sterile operating table. Cover with transparent paper and kraft paper in turn, seal with a rubber band, and place in a constant temperature incubator at 130 rpm and 30°C for 48 hours.

[0043] (2) Separation and purification:

[0044] Take 1 ml of the supernatant of the enriched bacterial solution (shake well), add 9 ml of sterile water to dilute the bacterial solution to a concentration of 10 -1 , dilute step by step 10 -2 , 10 -3 , 10 -4 , 10 -5 , 10 -6 , 10 -7 and 10 -8In an anaerobic tube, use the plate spreading method to absorb 0.1 ml of the original solution and the diluted bacterial solution on the solid culture medium, and then place it in a 30°C constant temperature incubator for culture. After 48 hours of culture, select the colonies with good growth, clear plaques and large diameter in each dilution gradient for streaking separation. During the separation process, each streak selects colonies with different morphologies and good growth conditions, and streaks on the solid culture medium for culture. The entire operation process is carried out in a sterile operating table, and the culture of the bacterial solution and the plate colonies are carried out under constant temperature anaerobic conditions;

[0045] Separate and purify for 4-6 generations until the single colonies grown on the solid culture medium are consistent in shape, the strain purification is completed, and the pure culture is obtained. The single strain is frozen in glycerol at -20°C. Select different pure bacterial colonies for denitrification experiments and calculate the NO3 - Removal rate, screening out NO3 - The strain with the best removal effect is strain BW-01.

[0046] (3) Morphological identification

[0047] The strain BW-01 was inoculated onto solid culture medium and cultured at 30°C for 24 hours, and then its colony morphology was observed. Figure 1 It can be seen that the colony morphology of strain BW-01 on solid culture medium is round, white, with a slight bulge in the middle, and complete and opaque edges.

[0048] (4) Physiological and biochemical identification

[0049] Physiology and Biochemistry Project result Physiology and Biochemistry Project result Nitrate reduction + glucose + Hydrogen sulfide - sucrose - Starch hydrolysis - maltose + Gelatin liquefaction - Cellobiose + Indole production - Mannose + Methyl red - lactose - Arabinose - fructose +

[0050] Note: “+” means positive; “-” means negative.

[0051] (5) Molecular biological identification

[0052] The DNA of the selected strains was extracted using a bacterial genome extraction kit, and the strain BW-01 was amplified by PCR using universal primers 27F and 1492R of the 16S rDNA gene. The PCR reaction system consisted of 1.0 μL of genomic DNA (20 ng / μL), 10× Buffer (containing 2.5 mM Mg 2+) 5.0μL, Taq polymerase (5u / μL) 1.0μL, dNTP (10mM) 1.0μL, 27F primer (SEQ ID NO.2: 5'-AGAGTTTGATCCTGGCTCAG-3') (10μM) 1.5μL, 1492R primer (SEQ ID NO.3: 5'-GGCTACCTTGTTACGACTT-3') (10μM) 1.5μL, ddH2O 39.0μL, total volume 50.0μL. PCR amplification conditions were: 95℃ pre-denaturation for 5min; 95℃ denaturation for 30s; 58℃ annealing for 30s; 72℃ extension for 1min30s; 72℃ final extension for 7min; 35 cycles. After the reaction was completed, 3ul of PCR product was taken for 1% agarose gel electrophoresis to confirm the PCR amplification fragment, and the PCR product sequencing was completed by Qingdao Weilai Biotechnology Co., Ltd.

[0053] The 16S rDNA sequence of strain BW-01 is 1436 bp in length, and the nucleotide sequence is as follows:

[0054] SEQ ID NO.1:

[0055]

[0056] The sequence was submitted to the NCBI database for blast comparison and the phylogenetic tree was drawn using MEGA7.0. The phylogenetic tree of strain BW-01 is shown in Figure 2 As shown, the results showed that the strain BW-01 had a similarity of 98.65% with Halomonas venusta NBRC 102221 (GenBank accession number NR 114902.1). Based on the above identification results, the strain was identified as Halomonas venusta.

[0057] Example 2: Optimization of optimal growth and denitrification conditions for strain BW-01

[0058] (1) Effects of different carbon sources on the anaerobic denitrification effect of strain BW-01

[0059] Activation: Inoculate the strain BW-01 preserved in glycerol at -20°C into 50 mL of sterilized liquid activation medium, place it in a shaker at 30°C and 120 rpm for 24 h to allow the bacteria to grow to a certain abundance. After harvesting the bacteria by centrifugation, dilute it with sterile water to an OD600 value of about 0.1, and use the bacterial suspension for inoculation.

[0060] Grouping: Use 10 anaerobic bottles and divide them into 5 groups, with 2 anaerobic bottles in each group. Add 50mL of natural seawater denitrification medium into each anaerobic bottle. Among them, the carbon sources in each group of anaerobic bottles are 1g / L sodium acetate, 1g / L sodium propionate, 1g / L sodium butyrate, 1g / L peptone, and 1g / L glucose, and the other ingredients are consistent in each group (sodium nitrate 0.9g / L, sodium bicarbonate 1.5g / L, 1L natural seawater (salinity 35g / L)); after sterilization and cooling to room temperature, inoculate the bacterial solution into 10 anaerobic bottles at a 2% (v / v) inoculation rate. Then culture at a constant temperature of 30°C and 120rpm for 72h, and take water samples every 12 hours to measure NO2 - 、NO3 - Concentration and solution OD 600 Absorbance. Note: All operations were performed on a sterile workbench. Figure 3 As shown by Figure 3 (c) It can be seen that strain BW-01 only grows well when peptone is used as the carbon source. 600 The value increased rapidly within 24 hours, approached 0.5 and reached a stable state after 60 hours, and the abundance of the strain no longer increased. Figure 3 (a) and Figure 3 (b) It can be seen that the strain began to rapidly degrade NO3 after inoculation. - , conduct NO3 - To NO2 -After 60h of cultivation, NO3 - The removal rate reached 83%, NO2 - The production rate reached 95.64%, and in the denitrification medium of other carbon sources, strain BW-01 not only utilized NO3 - The effect is poor and growth is stagnant, therefore, peptone is the most suitable carbon source.

[0061] (2) Effect of nitrogen source concentration on anaerobic denitrification of strain BW-01

[0062] The activation step is the same as the activation step in (1);

[0063] Grouping: 8 anaerobic bottles were used and divided into 4 groups, with 2 anaerobic bottles in each group. 50 mL of natural seawater denitrification medium was added to each anaerobic bottle. The nitrogen source concentrations in each group of anaerobic bottles were 100 mg N / L, 150 mg N / L, 200 mg N / L, and 250 mg N / L, respectively. The carbon source was peptone, and the other components were kept consistent in each group (sodium nitrate 0.9 g / L, sodium bicarbonate 1.5 g / L, 1 L natural seawater (salinity 35 g / L)); after sterilization and cooling to room temperature, the bacterial solution was inoculated into 8 anaerobic bottles at a 2% (v / v) inoculation rate. Then, the culture was carried out at a constant temperature of 30°C and 120 rpm for 72 hours, and water samples were taken every 12 hours to measure NO2 - 、NO3 - Concentration and solution OD 600 Absorbance. Note: All operations were performed on a sterile workbench. Figure 4 As shown by Figure 4 (a) It can be seen that the denitrification performance of the strain is the best under the condition of 100 mgN / L, NO3 - The removal rate reached 84.36% in 48 hours, which was earlier than the time when the denitrification efficiency reached more than 80% under other conditions, indicating that the nitrogen source concentration had a great influence on the denitrification performance of strain BW-01. Figure 4 (c)OD 600 Value and NO2 - From the perspective of the production rate, the bacterial growth of the strain at 100 mg N / L in each time period was better than that under other conditions. Figure 4 (b) It can be seen that, except for the NO2 - The generation rate was significantly lower than that of the other three groups. - The generation rate can reach more than 90%, achieving NO3 - To NO2 - Therefore, for strain BW-01, the optimal nitrogen source concentration is 100 mgN / L.

[0064] (3) Effects of different carbon-nitrogen ratios (C / N) on the anaerobic denitrification effect of strain BW-01

[0065] The activation step is the same as the activation step in (1);

[0066] Grouping: 12 anaerobic bottles were used and divided into 6 groups, with 2 anaerobic bottles in each group. 50 mL of natural seawater denitrification medium was added to each anaerobic bottle. Among them, in order to explore the effect of different carbon-nitrogen ratios C / N on the anaerobic denitrification of the strain, it was set as follows: the culture medium in the 12 anaerobic bottles used peptone as the carbon source, the nitrogen source concentration was 100 mgN / L, and the other components were kept consistent in each group (sodium nitrate 0.9 g / L, sodium bicarbonate 1.5 g / L, 1L natural seawater (salinity 35 g / L)). The C / N value was controlled by adjusting the content of peptone. C / N was set to 1, 2, 3, 5, 7, and 9, respectively. After the denitrification medium was sterilized and cooled to room temperature, the bacterial solution was inoculated into 12 anaerobic bottles at a 2% (v / v) inoculation rate. Then it was cultured at a constant temperature of 30°C and 120 rpm for 72 hours, and water samples were taken every 12 hours to measure NO2 - 、NO3 - Concentration and solution OD 600 Absorbance; Note: All operations are performed on a sterile workbench;

[0067] The results are as follows Figure 5 As shown by Figure 5 (a) It can be seen that when the carbon-nitrogen ratio C / N is 3, the denitrification efficiency of the strain is the highest, reaching 87.61% in 48 hours. When the carbon-nitrogen ratio C / N is 1, 2, and 5, the denitrification capacity of the strain is not much different, with a denitrification efficiency of about 80% in 48 hours. When the carbon-nitrogen ratio C / N is 7 or 9, the denitrification capacity of the strain decreases significantly. Figure 5 As shown in (b), when C / N is 1-5, NO2 - The generation rate is above 90%, which is a relatively high level. When C / N is 7 or 9, NO2 - The production rate is less than 90%, because when the carbon source is increased to a higher concentration, the strain begins to produce NO2 - to N2; Figure 5 (c) OD shown 600 From the value, the strain grows best when the C / N ratio is 3; so for strain BW-01, the optimal carbon-nitrogen ratio C / N is 3.

[0068] (4) Effect of salinity on anaerobic denitrification of strain BW-01

[0069] The activation step is the same as the activation step in (1);

[0070] Grouping: 10 anaerobic bottles were divided into 5 groups, with 2 anaerobic bottles in each group. 50 mL of artificial seawater denitrification medium was added to each anaerobic bottle. In order to explore the effect of salinity on anaerobic denitrification of strains, the following settings were used: the culture medium in the 10 anaerobic bottles used peptone as the carbon source, the nitrogen source concentration was 100 mgN / L, the carbon-nitrogen ratio C / N was 3, and the other components were consistent in each group (sodium nitrate 0.6 g / L, sodium bicarbonate 1.5 g / L, magnesium chloride 2.26 g / L, magnesium sulfate 3.25 g / L, calcium chloride 1.15 g / L, potassium chloride 0.72 g / L, sodium bicarbonate 1.7 g / L), and the sodium chloride concentration was 35 g / L, 60 g / L, 80 g / L, 100 g / L, and 120 g / L, respectively. At this time, the salinity of the five culture media was guaranteed to be 3.5%, 6%, 8%, 10%, and 12%, respectively. After the denitrification medium was sterilized and cooled to room temperature, the bacterial solution was inoculated into 10 anaerobic bottles at a 2% (v / v) inoculation rate. Then, the culture was carried out at a constant temperature of 30°C and 120 rpm for 72 hours, and water samples were taken every 12 hours to measure NO2 - 、NO3 - Concentration and solution OD 600 Absorbance. Note: All operations were performed on a sterile workbench. Figure 6 As shown by Figure 6 (a) It can be seen that when the sodium chloride concentration is in the range of 35-80g / L, the NO3 - The removal rate can reach more than 78%, and the denitrification effect of the strain is best when the salinity is 60g / L. - The removal rate of NO3 can reach 87.05% in 48h, and then the removal rate tends to be stable. The denitrification performance of the strains at other salinities is relatively low, especially when the salinity reaches above 100g / L. - The removal rate was less than 10% in the first 24 hours, which was because the salinity exceeded the optimal tolerance range of the strain. Figure 6 (b) NO2 - From the perspective of generation rate, the NO2 - The production rates were all high, indicating that the strains still mainly produced NO3 - To NO2 - The transformation from Figure 6 (c)OD 600 The values ​​show that the strain grows best at a salinity of 60 g / L, so for strain BW-01, the optimal salinity is 60 g / L.

[0071] Therefore, the best denitrification conditions for strain BW-01 are: peptone as carbon source, 100 mg N / L nitrogen source concentration, C / N ratio of 3, salinity of 60 g / L, NO3- The removal rate can reach 87.05% in 48 hours.

[0072] Example 3: Evaluation of the nitrogen removal effect of strain BW-01 on ship high-salinity nitrogen-containing wastewater

[0073] The composition of the ship high-salt nitrogen-containing wastewater used in this embodiment is: 1 g / L peptone, 0.9 g / L sodium nitrate (150 mg N / L), 1 L natural seawater, and the salinity is 35 g / L.

[0074] The strain stored in glycerol at -20°C was inoculated into 50 mL of sterilized liquid activation medium and cultured in a shaker at 30°C and 120 rpm for 24 h to allow the bacteria to grow to a certain abundance. The bacteria were harvested by centrifugation and diluted with sterile water to OD 600 The value is about 0.1, and the bacterial suspension is used for inoculation; 2% (v / v) bacterial liquid inoculation is inoculated into 50mL of ship high-salinity nitrogen-containing wastewater, and cultured at 30℃ and 120rpm for 96h. Water samples are taken every 12h to measure NO2 - 、NO3 - Concentration and solution OD 600 Absorbance. The results are as follows Figure 7 The results showed that strain BW-01 began to grow rapidly after inoculation and reached the maximum OD at 60h. 600 , was 0.405, and then the strain growth entered a stable period. The strain BW-01 decomposed NO3 while growing. - , maximum NO3 - The removal rate reached 81.84%, NO3 - During the degradation process, obvious NO2 - Accumulation, NO2 - The maximum production rate reached 99.91%, indicating that the main activity of the bacteria was NO3 - To NO2 - denitrification mode; after 60h, as the growth of the strain entered a stable period, NO3 - The removal rate gradually decreased, which shows that strain BW-01 has a good biological denitrification effect in high-salt nitrogen-containing wastewater from ships.

[0075] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A beautiful Halomonas strain having anaerobic half-denitrification function, characterized in that: The taxonomic name of the beautiful Halomonas venusta is (Halomonas venusta) BW-01, which is deposited in the General Microbiological Center of the China Microbiological Culture Collection Administration CGMCC. The deposit date is April 25, 2024, the deposit number is CGMCC No.30437, and the base length of 16S rDNA is 1436bp.

2. The beautiful Halomonas with anaerobic half-denitrification function according to claim 1, characterized in that The 16S rDNA sequence of the strain BW-01 is shown in SEQ ID NO.

1.

3. The beautiful Halomonas with anaerobic half-denitrification function according to claim 1, characterized in that The strain BW-01 can grow on a liquid culture medium with peptone as a carbon source and nitrate nitrogen as a nitrogen source. The colony of the strain BW-01 is round, white, has a bulge in the middle, and has a complete and opaque edge.

4. The beautiful Halomonas with anaerobic half-denitrification function according to claim 1, characterized in that The strain BW-01 can utilize peptone organic matter to perform denitrification and denitrification, and the denitrification and denitrification process is a half-denitrification process that converts nitrate nitrogen into nitrite nitrogen.

5. The use of the beautiful Halomonas with anaerobic half-denitrification function according to claim 1 in the treatment of high-salt nitrogen-containing wastewater from ships, characterized in that: The high-salt nitrogen-containing wastewater is saline wastewater with nitrate nitrogen as the nitrogen source.

6. The use of the beautiful Halomonas with anaerobic half-denitrification function in the treatment of high-salt nitrogen-containing wastewater from ships according to claim 5, characterized in that: In the saline wastewater, peptone is a carbon source, the concentration of the nitrogen source is 100-250 mgN / L, C / N=2-3, and the salinity is 35-80 g / L.

Images