Method for denitrification treatment of low carbon-nitrogen ratio water body

Abstract

The application provides a method for denitrification treatment of low carbon-nitrogen ratio water bodies, which utilizes a magnetite photoelectron coupling heterotrophic nitrification and aerobic denitrification bacteria system to remove nitrogen in low carbon-nitrogen ratio water bodies, and raw materials of the magnetite photoelectron coupling heterotrophic nitrification and aerobic denitrification bacteria system include magnetite and a Delftia sp. Y19 strain. 3‑ -N, NH 4+ -N, COD and NO 2‑ -N. The operation method is simple and easy to implement, and has a wide application prospect in the denitrification of low carbon-nitrogen ratio water bodies.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of industrial microorganisms, and in particular to a method for denitrification treatment of low carbon-nitrogen ratio water. BACKGROUND

[0002] Excessive nitrate in surface water and groundwater as drinking water sources can cause human methemoglobinemia, cancer and other diseases, and cause irreversible harm to human health. Therefore, removing nitrate and other nitrogen from low carbon-nitrogen ratio water is of great significance to protect the ecological environment and human health.

[0003] Low carbon-nitrogen ratio (C / N) sewage refers to sewage with a carbon-nitrogen ratio lower than that of normal water. This sewage can be considered as relatively "nutrient-poor" sewage because its carbon-nitrogen ratio is lower than that of normal water. The organic matter content in this sewage is relatively low, so it can be more easily decomposed by microorganisms, thereby reducing environmental pollution. However, there are still few studies on denitrification of low carbon-nitrogen ratio water. SUMMARY

[0004] The present application screens heterotrophic nitrifying and aerobic denitrifying bacteria, and constructs a magnetite photoelectron-promoted heterotrophic nitrifying and aerobic denitrifying bacteria denitrification system. The removal rates of NO3 - -N, NH4 + -N, the changes in the concentrations of COD and NO2 - -N are used as evaluation indexes to carry out dynamic experiments to study the denitrification efficiency of the system under light. Electrochemical technology is applied to analyze the photoelectrochemical response between magnetite and microorganisms, and to reveal the mechanism of magnetite photoelectron combined with HN-AD denitrification, so as to provide a theoretical basis and technical support for denitrification in river and lake water bodies.

[0005] The purpose of the present application is to provide a method for denitrification treatment of low carbon-nitrogen ratio water. The present application screens the heterotrophic nitrifying and aerobic denitrifying bacteria Delftia sp. Y19 strain, and constructs a magnetite photoelectron-promoted heterotrophic nitrifying and aerobic denitrifying bacteria denitrification system.

[0006] In order to achieve the above-mentioned purpose of the present application, the following technical solutions are adopted:

[0007] The present application provides a method for denitrification treatment of low carbon-nitrogen ratio water, which comprises the step of treating water with magnetite and Delftia sp. Y19 strain together, and the carbon-nitrogen ratio of the water is 10±1.

[0008] The Delftia sp. Y19 strain is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 26000.

[0009] In some embodiments, the magnetite is natural magnetite.

[0010] In some embodiments, the concentration of ammonia nitrogen (NH4 + -N) in the water body is 20±1.5 mg / L, the concentration of nitrate nitrogen (NO3 - -N) is 20±1.5 mg / L, and the concentration of COD is 400±10 mg / L.

[0011] In the present application, the Delftia sp. Y19 strain was deposited with the China General Microbiological Culture Collection Center on October 31, 2022, and has the classification name Delftia sp. with the preservation number CGMCC No. 26000 and the address of the preservation unit being No. 3, Beichen West Road, Chaoyang District, Beijing.

[0012] In the present application, the Delftia sp. Y19 strain is isolated and screened from the water body of a town river, and has the following biological characteristics:

[0013] Colony characteristics: intermediate convex, regular edge, smooth surface, beige, opaque;

[0014] Cell morphological characteristics: regular rod-shaped, cell size (1.1-1.9) μm x (0.5-0.7) μm, surface with wrinkles, no flagellum.

[0015] Biochemical characteristics:

[0016] The bacterium can grow in a mixed nitrogen source of ammonia salt and nitrate;

[0017] The bacterium can grow in yeast extract peptone;

[0018] The bacterium can grow under neutral or weakly alkaline conditions (pH 7-9);

[0019] The bacterium can grow in a dissolved oxygen concentration of 5.38-6.35 mg / L.

[0020] In one embodiment, the method for denitrification treatment of a low carbon-nitrogen ratio water body of the present application comprises the following steps:

[0021] S1, inoculating the Delftia sp. Y19 strain into LB liquid medium for activation;

[0022] S2, under light conditions, adding the bacterial solution obtained in S1 and magnetite into the water body to remove the nitrogen source.

[0023] In S1, preferably, the activation is carried out at 30℃, 120r / min oscillation culture for 12-36 hours, to OD 600 1.5.

[0024] In S1, preferably, the LB liquid medium is as follows: 10g / L of protein peptone, 5g / L of yeast powder, 10g / L of NaCl, and distilled water is added to 100mL, pH is 7.

[0025] In S2, preferably, the magnetite dosage is 2g / L, and the bacterial liquid dosage is 3%(v / v); at 30℃, 120r / min, oscillation culture for 2 days.

[0026] In S2, preferably, the light source of the light is a 300w xenon lamp.

[0027] The magnetite can produce e - and h + under light. - e + has reducing property, and h - has oxidizing property. + e - can be effectively separated, e 3- can take NO 3- as substrate, and reduce NO - to NO2 + , NH4 + and N2, and the organic acid produced in the system through nitrification can be oxidized by h - , so as to remove e 2+ (Formula (1)-Formula (7)). The part of Fe 2+ dissolved in the wastewater participates in the extracellular biological denitrification. The system of the application using the magnetite / Y19 coupling system includes the biological denitrification and the redox of the oxidized iron mineral, and the electronic energy of the magnetite mineral can affect the growth and metabolism of Y19 through the extracellular electron transfer, which enriches the energy acquisition mode of Y19. The part of Fe - dissolved in the wastewater participates in the extracellular biological denitrification, and the reaction formula is as follows:

[0028] NO3 - +2H + →NO2 - +H2O (1)

[0029] NO3 - +8e - +10H + →NH4 + +3H2O (2)

[0030] 2NO3 - +10e- +12H + →N2+6H2O (3)

[0031] NO2 - +6e - +8H + →NH4 + +2H2O (4)

[0032] 2NO2 - +6e - +8H + →N2+4H2O (5)

[0033] HCOO - +h + →H + +·CO2 - (6)

[0034] 2NO3 - +12H + +10·CO2 - →N2+6H2O+10CO2 (7)

[0035] Fe 2+ +2NO3 - +24H2O→10Fe(OH)3+N2+18H + (8)

[0036] The reaction process does not need to add an organic carbon source, and avoids the secondary pollution problem that may be caused by adding a carbon source in low C / N wastewater.

[0037] The magnetite / Y19 coupling system can effectively remove NO3 - -N, NH4 + -N, COD and NO2 - -N, and the magnetite photocatalysis generates electrons as extracellular electrons, promotes the growth and metabolism of Y19, and thus improves the denitrification performance of Y19. - -N, NH4 + -N and COD are respectively 100%, 36% and 100%, and the COD can be completely removed around the fourth day, and there is no accumulation of NO2 - -N in the system. The operation method is simple and easy to implement, and has a wide application prospect in the denitrification of low carbon-nitrogen ratio water bodies.

[0038] The magnetite / Y19 coupling system of the application is compared with Y19 alone in terms of denitrification performance in low carbon-nitrogen ratio water, and the ability of Y19 to utilize the photo-generated electrons of magnetite for denitrification is superior to the ability of Y19 alone to denitrify. BRIEF DESCRIPTION OF DRAWINGS

[0039] Figure 1 It is the screening result of heterotrophic nitrification and aerobic denitrification bacteria.

[0040] Figure 2 It is the denitrification ability of different strains.

[0041] Figure 3 It is the denitrification ability of different strains under low C / N conditions.

[0042] Figure 4 It is the colony map after the bacterial purification of Delftia sp. Y19 strain is streaked on a plate.

[0043] Figure 5 It is the electron microscope scanning map of Delftia sp. Y19 strain.

[0044] Figure 6 It is the growth curve of Delftia sp. Y19 strain in LB liquid medium.

[0045] Figure 7 It is the phylogenetic tree map of Delftia sp. Y19 strain.

[0046] Figure 8 It is the growth curve map of Delftia sp. Y19 under illumination.

[0047] Figure 9 It is the utilization ability map of Delftia sp. Y19 to photo-generated electrons of magnetite.

[0048] Figure 10 It is the influence map of magnetite dosage on the denitrification of the coupling system.

[0049] Figure 11 It is the influence map of Delftia sp. Y19 dosage on the denitrification of magnetite / Y19.

[0050] Figure 12 It is the denitrification performance map of different systems.

[0051] Figure 13 It is the electrochemical test map of the coupling system under illumination.

[0052] Figure 14The test device diagram for coupling system to repair low C / N sewage.

[0053] Figure 15 The diagram for simulating the change of three nitrogen concentrations in wastewater with time. DETAILED DESCRIPTION

[0054] The technical solutions of the present application will be clearly and completely described below in combination with the embodiments. Obviously, the described embodiments are only a part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments in the present application, all the other embodiments obtained by those skilled in the art without creative work belong to the protection scope of the present application.

[0055] The present application will be further described below by way of examples. Unless otherwise specified, the materials in the examples are prepared according to the existing methods, or directly purchased from the market.

[0056] The determination methods of ammonia nitrogen, nitrite nitrogen, nitrate nitrogen and COD involved in the following examples are as follows respectively:

[0057] The ammonia nitrogen is determined by the method of Determination of Ammonia Nitrogen in Water by Nessler's Reagent Spectrophotometry (HJ535-2009);

[0058] The nitrite nitrogen (NO2 - -N) is determined by the method of Determination of Nitrite Nitrogen in Water by Diazotization Coupling Spectrophotometry (GB7493-87);

[0059] The nitrate nitrogen (NO3 - -N) is determined by the method of Determination of Nitrate Nitrogen in Water by Ultraviolet Spectrophotometry (HZ-HJ-SZ-0138), which quantitatively determines the nitrate ion by its absorption at 220 nm wavelength;

[0060] The COD is determined by the method of Rapid Digestion Spectrophotometry (GB11914-89).

[0061] The magnetite is obtained from Zhengzhou City, Henan Province, and is commercially available. The magnetite is washed with tap water for multiple times, and then washed with deionized water to remove the surface impurities. After drying in a 60℃ oven, the magnetite is ground to 200 mesh and sealed for storage.

[0062] The medium formula used in the following examples is as follows:

[0063] In a 1L container, the components in Table 1 are added, and sterilized water is added to make up to 1000mL.

[0064] Table 1 Medium formula

[0065]

[0066]

[0067] Note: Vials salt solution: 6.5 g / L KH2PO4·3H2O, 2.5 g / L MgSO4·7H2O, 2.5 g / L NaCl, 0.05 g / L FeSO4·7H2O, 0.04 g / L MnSO4·7H2O; Bromothymol blue solution: 1 g bromothymol blue solution in 100 mL ethanol; Trace element solution: 4 g / L CuSO4·5H2O, 0.7 g / L FeSO4·7H2O, 7 g / L FeCl3·7H2O, 0.2 g / L CoCl3·6H2O, 3.4 g / L NaMoO4·2H2O, 2 g / L CaCl2·2H2O.

[0068] All the culture media were sterilized by high-pressure steam sterilization method (121℃, 20 min) during the operation process, and were disinfected; all the operations were carried out beside the alcohol lamp of the super-clean workbench.

[0069] The simulated wastewater in the examples was prepared by using pure water, NH4Cl, KNO3 and C6H 12 O6, etc., simulating low C / N wastewater, and mainly containing 20 mg / L NH4 + -N, 20 mg / L NO3 - -N and 400 mg / L COD.

[0070] The trace element mixture mainly contains 4 g / L CuSO4·5H2O, 0.7 g / L FeSO4·7H2O, 7 g / L FeCl3·7H2O, 0.2 g / L CoCl3·6H2O, 3.4 g / L NaMoO4·2H2O, 2 g / L CaCl2·2H2O.

[0071] Example 1 Screening and separation and identification of heterotrophic nitrification-aerobic denitrification strain

[0072] Enrichment of each strain in river water: river water was taken from Yuekui River (E: 105°; N: 30°) in Anyue County, Ziyang City, Sichuan Province, 5 mL of fresh river water was mixed in 100 mL of sterilized LB liquid medium, and was placed in a constant temperature shaker at 30℃ and 120 r / min for 48 h of enrichment, 5 mL of bacterial suspension was inoculated in fresh LB liquid medium, and the enrichment was repeated three times to obtain a bacterial suspension with rich colonies.

[0073] Screening of heterotrophic nitrifying strain: 5 mL of the enriched bacterial suspension was added to 100 mL of heterotrophic nitrifying medium, mixed uniformly, and placed in a 30°C, 120 r / min constant temperature shaker for 48 h of shaking culture. Whether the culture medium was turbid was observed. If it was turbid, 5 mL of the cultured heterotrophic nitrifying medium was inoculated into 100 mL of fresh heterotrophic nitrifying medium, and placed in a 30°C, 120 r / min constant temperature shaker for 48 h of shaking culture. This was repeated three times. If the culture medium was not turbid, the above enrichment and screening steps were repeated again.

[0074] Isolation of heterotrophic nitrifying strain: 1 mL of the bacterial suspension was gradient diluted (diluted by 10, 10 2 , 10 3 , 10 4 , 10 5 , 10 6 , 10 7 times successively) with sterile water. The heterotrophic nitrifying solid medium was sterilized at 121°C for 20 min, and then poured into sterile plates while hot. After solidification, 100 μL of the diluted bacterial suspension was taken with a pipette gun and uniformly spread on the heterotrophic nitrifying solid medium with a spreader. Each plate was sealed with a sealing film and placed in a 30°C constant temperature incubator for 48 h of culture.

[0075] Purification of heterotrophic nitrifying strain: different colonies with better growth and faster growth rate were selected according to the colony morphology. The selected single colonies were streaked on LB solid medium with a loop, and the plate was sealed with a sealing film and cultured at 30°C until complete colonies were visible to the naked eye. The purified strain was obtained. The purified strain was inoculated into 100 mL of LB liquid medium, and placed in a 30°C, 120 r / min constant temperature shaker for 24 h of culture to obtain a bacterial suspension.

[0076] Screening of heterotrophic nitrifying and aerobic denitrifying strain: 5 mL of the bacterial suspension obtained in the above step was inoculated into 100 mL of aerobic denitrifying liquid medium, and placed in a 30°C, 120 r / min constant temperature shaker for 48 h of culture. 5 mL of the cultured aerobic denitrifying medium was inoculated into 100 mL of fresh aerobic denitrifying liquid medium, and placed in a 30°C, 120 r / min constant temperature shaker for 48 h of shaking culture. This was repeated three times.

[0077] Identification of heterotrophic nitrifying and aerobic denitrifying strain: the bacterial suspension was taken with a loop and streaked on a bromothymol blue medium. The plate was sealed with a sealing film after streaking, and cultured at 30°C until complete colonies were visible to the naked eye. When the bromothymol blue medium turned blue, it indicated that the strain had denitrification ability. Seven strains Y4, Y5, Y6, Y8, Y9, Y19 and Y20 that made the bromothymol blue medium turn blue were screened out, as shown in FIG. 1.Figure 1 As shown.

[0078] Take 2 mL of bacterial suspension from each of the above 7 strains and culture them until the logarithmic OD phase. 600 A bacterial suspension of strain 1.5 was transferred to 100 mL of a secondary screening liquid medium containing sufficient carbon source and incubated at 30 °C and 120 rpm for 72 h. The final NH4 content in the medium was then measured. + -N, NO2 - -N, NO3 - -N, TN (total nitrogen), and COD concentrations, such as Figure 2 As shown. After 48 hours of cultivation, these 7 bacterial strains showed resistance to NH4+ in the culture medium. + -N, NO3 - The removal effects on -N, TN and COD are all good, and the removal effect on NH4 is also good. + The removal rate of -N was generally above 90%, with strains Y8 and Y20 showing particularly good removal rates of NH4+. + The removal rates of -N were 98.7% and 98.4%, respectively; Y8 and Y19 removed NO3. - The removal rates of nitrogen (N) were 89.6% and 85.3%, respectively; in terms of COD removal, Y5 and Y8 achieved COD removal rates of 78.3% and 83.4%, respectively. Based on the screening of microorganisms' ability to remove nitrogen and carbon in water bodies with sufficient organic matter, Y5, Y8, Y19, and Y20 were identified as strains with relatively good capabilities.

[0079] The selected strains Y5, Y8, Y19, and Y20, which exhibited the best denitrification capabilities, were cultured in LB medium until the logarithmic growth phase. 2 mL of the bacterial suspension was then added to 100 mL of simulated wastewater with a low C / N ratio. After reacting for 72 h, NH4 content was measured. + -N, NO2 - -N, NO3 - -N, TN, and COD concentrations. Strains exhibiting good denitrification efficiency under low C / N conditions are screened; these strains are the target strains. For example... Figure 3 As shown, strain Y5 is effective against NO3. - -N, NH4 + The removal rates of NO2- and COD were 71.6%, 62.9%, and 70.4%, respectively, while after 48 hours of cultivation, NO2- - The concentration of -N was 18 mg / L, indicating that the system contained NH4+. + -N, NO3 - -N was mostly converted into NO2 by strain Y5 under low C / N conditions. - -N, no nitrogen was removed from the wastewater. Y19 exhibited better denitrification ability under low C / N conditions, particularly for NH4+. + -N, NO3 -- The removal rates of N, TN and COD were 51%, 94%, 56% and 71%, respectively. There was a small amount of NO2 - - N accumulation at the end of the experiment, which might be related to the denitrification pathway of the heterotrophic nitrification-aerobic denitrification (HN-AD) strain, NH4 + - N, NO3 - - N were first oxidized and reduced to NO2 - - N. After analysis, Y19 can be used as the experimental strain for follow-up research.

[0080] Characteristics of the screened strain Y19 on solid medium: intermediate convex, edges neat, smooth surface, beige, opaque, as shown in Figure 1; its morphological characteristics are regular rod-shaped, size (1.1-1.9) μm x (0.5-0.7) μm, as shown in Figure 2. Figure 4 Figure 5 Characteristics of the screened strain Y19 on solid medium: intermediate convex, edges neat, smooth surface, beige, opaque, as shown in Figure 1; its morphological characteristics are regular rod-shaped, size (1.1-1.9) μm x (0.5-0.7) μm, as shown in Figure 2.

[0081] Detection of the nitrogen removal ability of the heterotrophic nitrification-aerobic denitrification strain: take 1 inoculation ring of the target strain in 100 mL of LB liquid medium and incubate in a constant temperature shaker at 30°C and 120 r / min for 24 h to obtain a bacterial suspension. Take 2 mL of the bacterial suspension into 100 mL of the rescreening liquid medium and incubate in a constant temperature shaker at 30°C and 120 r / min for 72 h. Measure the final NH4 + , NO2 - , NO3 - and COD concentrations in the medium.

[0082] Growth curve determination of the heterotrophic nitrification-aerobic denitrification strain: take 1 inoculation ring of the target strain into 100 mL of LB liquid medium and incubate in a constant temperature shaker at 30°C and 120 r / min for 36 h. Take samples at 1, 2, 4, 6, 8, 12, 24, 32 and 36 h. Measure the absorbance at a wavelength of 600 nm using a UV spectrophotometer. Draw the growth curve according to the absorbance, as shown in Figure 3. Figure 6 As can be seen from Figure 3, the Delftia sp. Y19 strain grows faster and the strain metabolizes faster. The strain reaches the fastest growth period at 12 h, OD 600 = 1.5, and reaches the stationary phase after 24 h.

[0083] Example 2 PCR amplification and sequence determination of the 16s rDNA gene of the Delftia sp. Y19 strain

[0084] ① DNA extraction of the strain: the strain obtained in the above step is subjected to bacterial genomic DNA extraction kit (Tsingke) according to the instructions;

[0085] ​② PCR analysis of the strain: The full-length 16S rRNA gene sequence of the strain's genomic DNA obtained in step ① was sequenced and amplified by PCR using universal primers.

[0086] The universal primers used are: 27F: 5'-GAGAGTTTGATCCTGGCTCAG-3' (SEQ ID NO.2); 1492R: 5'-TACGGCTACCTTGTTACGAC-3' (SEQ ID NO.3);

[0087] Amplification conditions: 1 μl DNA template, 45 μl 1×TSE101 Gold Mix, 2 μl each of upstream and downstream universal primers;

[0088] After mixing, place it in a PCR instrument for amplification. The PCR program is as follows: 98℃ pre-denaturation for 3 min; 98℃ denaturation for 10 s, 55℃ annealing for 15 s, 72℃ extension for 5 min, 39 cycles.

[0089] After PCR, agarose gel electrophoresis (2 μl sample + 6 μl bromophenol blue) was performed at 300V for 12 minutes to detect whether the PCR amplification was successful.

[0090] When electrophoresis yields a 1500bp fragment, PCR amplification is considered successful; conversely, when electrophoresis does not yield the relevant fragment, PCR amplification is considered a failure.

[0091] ③ DNA sequencing of the strain: The strain was sequenced using universal primers (27F / 1492R) at Qingke Biotechnology Co., Ltd., and its nucleotide sequence is shown in SEQ ID NO.1. The sequence length is 1537 bp.

[0092] The selected strains were sequenced using 16S rRNA and compared with data in NCBI. The results showed that the isolated bacteria belonged to *Delftia sp.*. Figure 7 As shown. This bacterium was deposited at the China General Microbiological Culture Collection Center (CGMCC) on October 31, 2022, with accession number CGMCC No. 26000.

[0093] Example 3: Growth curve of strain Y19 under light irradiation

[0094] Take 5 mL of cultured medium until the logarithmic phase (12 h) OD 600 Centrifuge the bacterial suspension at 8000 rpm for 5 min to a concentration of 1.5, then dilute with PBS buffer to an OD value of 1.5. 600The bacterial suspension of 0.5 was added to a quartz single-chamber reactor containing 100 mL of good-nourishing denitrification liquid medium; 30 mg of magnetite ground to 200 mesh was mixed with 100 μL of 5% nafion reagent, and then smeared on a 20 mm x 20 mm x 1.1 mm FTO electrode using a glass rod. The FTO electrode was cleaned with anhydrous ethanol and deionized water twice to remove surface impurities, and then dried at 60°C for standby. The conductive surface of the FTO electrode was determined using a multimeter, and the magnetite was evenly smeared on the conductive surface of the FTO electrode and placed in the above-mentioned quartz single-chamber reactor. The reactor was placed in a constant-temperature shaker at 30°C and 120 r / min, and irradiated with a 300-watt xenon lamp for 96 h. Two control groups were set up: one was the above-mentioned reactor (bacterial suspension + magnetite) wrapped with tin foil paper, and the other was a quartz single-chamber reactor prepared by adding only the bacterial suspension (the process was the same as above) and wrapped with tin foil paper. The growth curves were determined under the same conditions.

[0095] The results are shown in Figure 8 The growth of bacteria in the light-magnetite-Y19 system was significantly better than that in the dark-magnetite-Y19 system. This indicates that the photo-generated electrons excited by the magnetite under light can effectively stimulate and promote the growth of microorganisms. Before 24 h, the growth of bacteria under dark conditions was better than that under light, indicating that Delftia sp. Y19 preferentially utilized the organic carbon source in the system for growth and metabolism in the early stage of the experiment. Under dark conditions, due to the adaptability of the strain and the influence of simulated natural light on Delftia sp. Y19, the growth of Delftia sp. Y19 in the first 12 h under dark conditions was better than that under light-Delftia sp. Y19-magnetite. Since Delftia sp. Y19 gradually adapted to the environment, and the magnetite had the ability to absorb ultraviolet light, it could reduce the cell damage of microorganisms caused by simulated natural light, so the bacteria could grow normally under simulated natural light. As the experiment proceeded, the microorganisms may gradually adhere to the surface of the magnetite, affecting the conversion of light energy by the magnetite, and the e - The production amount was small, and due to the adhesion of the strain, the content of microorganisms in the solution was low, so the growth curve of the strain began to decline after 72 h.

[0096] Example 4 Utilization ability of Delftia sp. Y19 for magnetite photo-generated electrons

[0097] The determination of cyclic voltammetry curves (CV), linear sweep voltammetry tests (LSV), and current-time curves (I-t) was performed using an electrochemical workstation (Shanghai Chenhua) (as shown in Figure 9 ).

[0098] The working electrode is a magnetite-coated or uncoated FTO electrode, the reference electrode is an Ag / AgCl electrode, and the counter electrode is a platinum wire electrode.

[0099] Electrolyte: Add 5 mL of bacterial suspension and 0.1 mol / L KH2PO4 to 100 mL of LB medium solution. The 5 mL bacterial suspension is prepared by centrifuging 5 mL of bacterial suspension cultured to the logarithmic growth phase (12 h) at 8000 r / min for 5 min. Dilute the lowermost bacterial suspension with PBS buffer to OD200. 600 A bacterial suspension of 0.5 g (5 mL).

[0100] The device was tested after 2 hours of light-protected reaction. Illuminated samples were tested under illumination (xenon lamp irradiation, 300W), while dark samples were tested after being wrapped in tin foil.

[0101] CV curve experimental parameters: scan rate set to 100mV / s, number of scan segments 6, sampling interval 50mV, sensitivity 10. - 4 A / V; LSV curve experimental parameters: initial potential 1.5V, endpoint potential -1V, scan rate 15mV / s, sampling interval 0.001V, interval time 2s, sensitivity is 10. -4 A / V; It curve experimental parameters: initial potential 0.3V, sampling interval 0.1s, running time 600s (alternating between 50s in darkness and 50s in light), settling time 0s, sensitivity 10 -6 A / V.

[0102] Depend on Figure 9 It can be seen that when treating pollutants, the e in the system - and h + Effective transfer and separation can be achieved; the magnetite / Y19 coupling system uses redox reactions to transfer NH4+. + -N and NO3 - -N removal: Delftiasp. Y19 can transfer extracellular electrons between magnetite electrodes, exhibiting strong redox capabilities. HN-AD facilitates heterotrophic nitrification and aerobic denitrification, photogenerated electron reduction of magnetite, and Fe... 2+ The involvement of biological denitrification and magnetite adsorption is the key to NH4 removal. + -N and NO3 - The main pathway of -N.

[0103] Example 5: Effect of magnetite dosage under illumination on denitrification of the magnetite / Y19 coupled system

[0104] Magnetite was added to different conical flasks at 1 g / L, 2 g / L and 5 g / L, respectively, with a simulated wastewater volume of 300 mL, a trace element mixture dosage of 1 mL / L, a Delftia sp. Y19 dosage of 1%, and a pH control of 7.0. NO3 - -N and NH4 + The initial concentration of -N was 20 mg / L, and the COD concentration was 400 mg / L. Each experimental group was placed in a device at 120 r / min and 30°C, with a light source of 300 w xenon lamp, and the reaction was carried out for 72 h. Sampling was performed every 24 h, and the sample was filtered with a 0.45 μm filter head before determination of NO3 - -N, NO2 - -N, NH4 + The concentration of -N and COD.

[0105] When the magnetite dosage was 2 g / L, the removal rates of NO3 - -N, NH4 + The removal rates of -N, NH4 - -N were 54.2%, 88.4% and 91.1%, respectively. When the magnetite dosage was 1 g / L and 5 g / L, the removal rates of NO3 + -N were 49.1%, 44.3%, the removal rates of NH4 - -N were 83.7%, 69.5%, and the removal rates of COD were 85.9%, 76.7% (as shown in Table 2). Therefore, when the magnetite dosage was 2 g / L, the denitrification effect of the magnetite / Y19 coupling system under simulated sunlight was best. Figure 10

[0106] Example 6 Influence of Y19 dosage on denitrification of magnetite / Y19 coupling system under light

[0107] Delftia sp. Y19 was added to conical flasks containing 300 mL of simulated wastewater at dosages of 1%, 3%, 5% and 7%, with a trace element mixture dosage of 1 mL / L, a magnetite dosage of 2 g / L, and a pH control of 7.0. NO3 - -N and NH4 + The initial concentration of -N was 20 mg / L, and the COD concentration was 400 mg / L. Each experimental group was placed in a device at 120 r / min and 30°C, with a light source of 300 w xenon lamp, and the reaction was carried out for 72 h. Sampling was performed every 24 h, and the sample was filtered with a 0.45 μm filter head before determination of NO3 - -N, NO2 - -N, NH4 + The concentration of -N and COD.

[0108] NO3 - -N, NH4+ The removal rates of NO3- and COD increased with increasing dosage of Delftia sp. Y19. When the dosage was 7%, the removal rates of NO3- and COD- were significantly higher. - The removal of -N reached its maximum at 24 hours, with a removal rate of 96.8%. COD was also rapidly removed within 24 hours. This resulted in a lack of organic carbon source for microorganisms, and due to excessive microbial addition, a biofilm quickly formed on the magnetite surface, leading to a decrease in the e-carbon content of the magnetite. - The amount produced is reduced, the conversion of nitrogen is incomplete, and NO2 production is low at the end of the experiment. - -N accumulated significantly, reaching a concentration of 15.7 mg / L. At a dosage of 1%, NO3... - -N, NH4 + The removal rates of NO2- and -N were 64.1% and 66.2%, respectively. In the later stages of the experiment... - -N accumulation was severe at 9.0 mg / L. The dosage was 3%, NO3... - The removal rate of -N was 77.3%, and NH4+ was... + The removal rate of -N was 93.2%, and the removal rate of COD was 96% (from...). Figure 11 (As shown), NO2 - The cumulative concentration of -N was 3.4 mg / L. When the dosage was increased further, the removal rate did not increase. Therefore, a dosage of 3% Delftia sp. Y19 is more conducive to denitrification in the magnetite / Y19 coupled system.

[0109] Example 7: Simulation of nitrogen removal performance of magnetite / Y19 coupled system under sunlight

[0110] Three coupled systems were designed: light-magnetite / Y19, light-Delftia sp. Y19, and darkness-magnetite / Y19. The magnetite dosage was 2 g / L, the Delftia sp. Y19 dosage was 3% (v / v), the pH was controlled at 7.0, and NO3 was controlled at... - -N and NH4 + The initial concentrations of NO3- were all 20 mg / L, and the COD concentration was 400 mg / L. Each experimental group was placed in an apparatus at 120 rpm and 30°C, with a 300 W xenon lamp as the light source. The dark group was wrapped in yellow paper. The reaction proceeded for 72 hours. Samples were taken every 24 hours, filtered through a 0.45 μm filter, and NO3 was measured. - -N, NO2 - -N, NH4 + The concentrations of -N and COD were recorded, and the pH changes for each group were also recorded. Figure 12 It can be seen that the ability of Delftia sp. Y19 to denitrify using photogenerated electrons from magnetite under simulated sunlight is significantly better than other systems, especially for NO3.- - N, NH4 + The removal rates of -N were 71.3% and 80.1%, respectively, and the removal rate of COD was 97.9%. At the end of the experiment, there was NO2 - -N, and the accumulated -N was 6.4 mg / L. Under the condition of low C / N, due to the insufficient organic carbon source and the damage of light to Delftia sp. Y19 cells, the Delftia sp. Y19 system alone had poor removal ability for NO3 - - N, NH4 + - N, and the removal rates were about 15.2% and 20.3%. Under dark conditions, the magnetite-Y19 system had poor removal ability for NO3 - - N, NH4 + - N, and the removal rates were about 15.2% and 20.3%. Under dark conditions, the magnetite-Y19 system had poor removal ability for NO3

[0111] In order to verify the electrochemical characterization of the coupling system under light, cyclic voltammetry and linear sweep voltammetry tests were carried out, and the results are shown in Figures 8 to 10. Figure 13 From the CV curves, it can be seen that light improves the reduction peak intensity of the magnetite / Y19 coupling system, but does not shift the reduction potential. Light can produce extracellular electrons in Delftia sp. Y19 and magnetite, and will not change the properties of the mineral itself. The DPV curves verify the redox reactions in the coupling system under light, and under the condition that the organic carbon source in the system is basically removed, the microorganism continues to carry out redox reactions using the photo-generated electrons produced by the magnetite, and the extracellular electrons produced by the microorganism are transferred to the surface of the magnetite electrode, indicating the electron transfer between the microorganism and the semiconductor. From the LSV curves, it can be seen that the photo-generated charges in the coupling system can be effectively separated. Under light, both the magnetite and Delftia sp. Y19 have obvious photoresponse, and the photocurrent curve of the magnetite / Y19 coupling system stably rises, indicating that the electron transfer process between the coupling systems is relatively active, and the extracellular electrons produced by Delftia sp. Y19 can be accepted by the magnetite electrode. These results show that under the condition of insufficient organic carbon source, Delftia sp. Y19 can use the photoelectrons produced by the magnetite to carry out growth and metabolism activities, and continue to carry out denitrification, and the denitrification efficiency of the magnetite / Y19 coupling system is higher under light.

[0112] Example 8 Application of Delftia sp. Y19 Coupling Magnetite to Repair Nitrogen-Containing Wastewater

[0113] The magnetite / Y19 coupling system device column is an organic glass cylinder, as shown in Figure 11. Figure 14As shown, the column is 35 cm high, 5 cm in inner diameter, and has a volume of 0.69 L. To simulate a real aquatic environment, a 0.1 m thick layer of bottom sediment was placed at the bottom of the device. The wastewater volume was 350 mL. 10 mL of activated Delftia sp. Y19 was added to the microbiome. In the magnetite / Y19 coupling system, the amount of magnetite added was 1 g, and the amount of activated Delftia sp. Y19 added was 10 mL. The experimental temperature was maintained between 15 and 25 °C for 10 days. Water samples were taken daily for NH4+ analysis. + -N, NO3 - -N, NO2 - The concentration of -N was determined, and pH was measured using a portable pH meter at each sampling.

[0114] For NO3 - The change of -N is as follows Figure 15 As shown in (a), in the two systems on day 10, NO3 - -N removal rate was 100%. In the first five days, the microbial system alone achieved 100% removal of NO3. - The removal rate of NO3- was 87.9%, while the magnetite / Y19 coupling system showed a better removal rate of NO3-. - The removal rate of -N was 80.8%, indicating that Delftia sp. Y19 has good environmental adaptability and can quickly mobilize indigenous microorganisms to participate in denitrification; for NH4... + The change of -N is as follows Figure 15 As shown in (b), the removal rate was approximately 36% on day 5, while in wastewater treated only with Delftia sp. Y19, the NH4+ removal rate was significantly lower. + The NO3- concentration continued to rise, reaching approximately 32.8 mg / L on the second day. This indicates that NO3- in the Delftia sp. Y19 system was increasing. - The decrease in -N concentration only reduces NO3- concentration. - -N was mostly converted into NH4. + -N, NO3 was not implemented. - Removal of -N; for NO2 - Changes in -N, NO2 in individual strain groups - -N accumulated to its maximum level on day 5, reaching approximately 10.8 mg / L. This indicates that NO3- levels were significantly lower in the system containing only Delftia sp. Y19. - -N was converted to NH4 + -N and NO2 - -N, in the first five days, basically no nitrogen removal was achieved. From the sixth day onwards, the microorganisms began to utilize NO2. --N, it is possible that with the reaction proceeding, the denitrifying bacteria in the system gradually dominated, began to denitrify with NO2 - -N as the main substrate, and finally NO2 - -N was completely removed. The NO2 - -N accumulation trend was similar to the group only adding microorganisms, but the maximum accumulation was 5.8 mg / L, and it is possible that part of the photo-generated electrons produced by magnetite was used for NO2 - -N reduction, so the NO2 - -N concentration was not as high as that of the single microorganism group, and after the sixth day, with the denitrifying bacteria dominating, NO2 - -N was gradually removed.

[0115] From the above examples, the optimal dosages of magnetite and microorganisms were 2 g / L and 3%, respectively. The results of the 10-day denitrification performance verification experiment of the magnetite / Y19 coupling system under simulated sunlight showed that the coupling system had good denitrification effect, and the removal rates of NO3 - -N, NH4 + -N were 100%, 36%, respectively, and the removal rate of COD was 100%, which had strong feasibility in application.

[0116] SEQ ID NO. 1

[0117] GTCGAACGGTAACAGGTCTTCGGACGCTGACGAGTGGCGAACGGGTGAGTAATAC

[0118] ATCGGAACGTGCCCAGTCGTGGGGGATAACTACTCGAAAGAGTAGCTAATACCGCA

[0119] TACGATCTGAGGATGAAAGCGGGGGACCTTCGGGCCTCGCGCGATTGGAGCGGCC

[0120] GATGGCAGATTAGGTAGTTGGTGGGATAAAAGCTTACCAAGCCGACGATCTGTAGC

[0121] TGGTCTGAGAGGACGACCAGCCACACTGGGACTGAGACACGGCCCAGACTCCTA

[0122] CGGGAGGCAGCAGTGGGGAATTTTGGACAATGGGCGAAAGCCTGATCCAGCAATG

[0123] GCGTGCAGGATGAAGGCCTTCGGGTTGTAAACTGCTTTTGTACGGAACGAAAA

[0124] AGCTCCTTCTAATACAGGGGGCCCATGACGGTACCGTAAGAATAAGCACCGGCTAA

[0125] CTACGTGCCAGCAGCCGCGGTAATACGTAGGGTGCGAGCGTTAATCGGAATTACTG

[0126] GGCGTAAAGCGTGCGCAGGCGGTTATGTAAGACAGATGTGAAATCCCCGGGCTCA

[0127] ACCTGGGAACTGCATTTGTGACTGCATGGCTAGAGTACGGTAGAGGGGGATGGAA

[0128] TTCCGCGTGTAGCAGTGAAATGCGTAGATATGCGGAGGAACACCGATGGCGAAGG

[0129] CAATCCCCTGGACCTGTACTGACGCTCATGCACGAAAGCGTGGGGAGCAAACAGG

[0130] ATTAGATACCCTGGTAGTCCACGCCCTAAACGATGTCAACTGGTTGTTGGGAATTA

[0131] GTTTTCTCAGTAACGAAGCTAACGCGTGAAGTTGACCGCCTGGGGAGTACGGCCG

[0132] CAAGGTTGAAACTCAAAGGAATTGACGGGGACCCGCACAAGCGGTGGATGATGT

[0133] GGTTTAATTCGATGCAACGCGAAAAACCTTACCCACCTTTGACATGGCAGGAAGTT

[0134] TCCAGAGATGGATTCGTGCTCGAAAGAGAACCTGCACACAGGTGCTGCATGGCTG

[0135] TCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTT

[0136] GTCATTAGTTGCTACATTCAGTTGAGCACTCTAATGAGACTGCCGGTGACAAACCG

[0137] GAGGAAGGTGGGGATGACGTCAAGTCCTCATGGCCCTTATAGGTGGGGCTACACA

[0138] CGTCATACAATGGCTGGTACAGAGGGTTGCCAACCCGCGAGGGGGAGCTAATCCC

[0139] ATAAAACCAGTCGTAGTCCGGATCGCAGTCTGCAACTCGACTGCGTGAAGTCGGA

[0140] ATCGCTAGTAATCGCGGATCAGCATGCCGCGGTGAATACGTTCCCGGGTCTTGTAC

[0141] ACACCGCCCGTCACACCATGGGAGCGGGTCTCGCCAGAAGTAGGTAGCCTAACCG

[0142] CAAGGAG

Claims

1. A method for denitrification treatment of a low carbon-nitrogen ratio water body, characterized in that, The method comprises the step of treating the water body with magnetite and Delftia sp. Y19 strain together, and the carbon-nitrogen ratio of the water body is 10±1. The Delftia sp. Y19 strain is preserved in the China General Microbiological Culture Collection Center, and the preservation number is CGMCC No. 26000. The method comprises the following steps: S1, inoculating the Delftia sp. Y19 strain into LB liquid medium for activation; S2, under the light condition, adding the bacterial solution obtained in S1 and magnetite into the water body to remove the nitrogen source; In S2, the adding amount of magnetite is 2 g / L, and the adding amount of bacterial solution is 3% (v / v); under the condition of 30℃, the rotation speed is 120 r / min, and the oscillation culture is carried out for 2 days; In S2, the light source of the light is a 300 W xenon lamp.

2. The method of claim 1, wherein, The magnetite is natural magnetite.

3. The method of claim 1, wherein, The concentration of ammonia nitrogen (NH4 + -N) in the water body is 20±1.5 mg / L, the concentration of nitrate nitrogen (NO3 - -N) is 20±1.5 mg / L, and the concentration of COD is 400±10 mg / L.

4. The method of claim 1, wherein, In S1, the activation is at 30 °C, 120 r / min oscillation culture for 12-36 hours, to OD 600 1.

5.

5. The method of claim 1, wherein, In S1, the LB liquid medium is as follows: peptone 10 g / L, yeast powder 5 g / L, NaCl 10 g / L, and distilled water is constant volume to 100 mL, and the pH is 7.

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