A method for achieving zero-carbon denitrification in water by coupling conductive bacteria with zero-valent iron

Abstract

This invention discloses a method for achieving zero-carbon denitrification of water by coupling conductive bacteria with zero-valent iron, belonging to the field of nitrogen-containing wastewater treatment technology. The invention involves placing polluted water containing nitrate nitrogen in a reactor, then adding conductive bacteria and zero-valent iron according to a specific ratio, followed by the addition of denitrifying microorganisms or anaerobic denitrifying sludge. Finally, the reactor is sealed after the oxygen is removed and subjected to constant temperature and vibration at 20-35°C for 2-20 days. No additional organic carbon source is artificially added to the reactor. This invention couples conductive bacteria with zero-valent iron, allowing the conductive bacteria to extract electrons from the zero-valent iron and directionally transfer them to the denitrifying bacteria, thereby improving the electron utilization rate of zero-valent iron and denitrification efficiency. Furthermore, compared to traditional methods, this invention does not require an additional organic electron donor, potentially reducing carbon emissions during the denitrification process.

Description

Technical Field

[0001] This invention belongs to the field of nitrogen-containing wastewater treatment technology, specifically relating to a method for achieving zero-carbon denitrification of water by coupling conductive bacteria with zero-valent iron. Background Technology

[0002] Nitrate is considered a typical pollutant in water bodies, primarily released into the environment through industrial activities and the large-scale use of chemical fertilizers in agricultural production. Although nitrate is an essential nutrient for organisms, excessive nitrate in water bodies can lead to eutrophication and harm human health. The problem of nitrate pollution in water bodies has attracted widespread attention from researchers and government departments and is a key research focus in the field of water treatment.

[0003] Anaerobic denitrification is a traditional microbial nitrate treatment technology that utilizes electrons generated during the respiration and metabolism of organic carbon by denitrifying bacteria to gradually reduce nitrates into nitrogen gas. It's easy to see that the treatment efficiency of heterotrophic anaerobic denitrification depends on the ratio of organic carbon to nitrate nitrogen. Theoretically, when the ratio of organic carbon (calculated as chemical oxygen demand (COD)) to nitrate nitrogen is 2.86, nitrate nitrogen can be completely converted into nitrogen gas. However, in practical scenarios, an organic carbon to nitrate nitrogen ratio of around 5 is needed to achieve complete removal of nitrate nitrogen from water. In water treatment processes, anaerobic digestion is often used to remove organic matter from wastewater, followed by anaerobic denitrification to remove nitrates. At this point, insufficient organic carbon is often encountered, requiring the addition of an extra carbon source to improve denitrification efficiency, thus increasing treatment costs. Therefore, it is necessary to find a highly efficient and green alternative electron donor to reduce the treatment costs in the denitrification process.

[0004] Zero-valent iron (ZVFe) and related iron-based materials are used by researchers to enhance the treatment of low C / N ratio wastewater due to their low cost and environmental friendliness. In this process, hydrogen generated from the anaerobic hydrogen evolution corrosion of ZVFe is typically used as an electron donor to improve the denitrification efficiency of low C / N ratio wastewater. However, due to the limited solubility of hydrogen, the electrons from iron-based materials cannot be efficiently utilized in the denitrification process. Furthermore, because the chemical and biological reduction processes of nitrates compete for electrons, some nitrate nitrogen is chemically reduced to ammonia nitrogen, thus affecting the denitrification efficiency of the water body. Therefore, finding a method to regulate the electron selectivity of ZVFe in the biological denitrification process (i.e., the proportion of electrons contributed by ZVFe to the reduction of nitrate nitrogen to nitrogen gas relative to the total electrons consumed in the reaction) has become a key issue in improving the electron utilization efficiency of ZVFe.

[0005] Studies have shown that conductive bacteria can participate in the microbial corrosion process of metals, obtaining electrons from metallic materials and redistributing them to electron acceptors in the environment through extracellular electron transfer. Therefore, using conductive bacteria to couple zero-valent iron (ZVFe) shows promise as an effective way to improve the electron selectivity of ZVFe and may even achieve denitrification of water bodies without the addition of organic carbon sources.

[0006] Based on the above reasons, this application is hereby submitted. Summary of the Invention

[0007] The technical problem solved by this invention is the low efficiency of anaerobic denitrification in wastewater with a low carbon-to-nitrogen ratio. It provides a highly efficient and inexpensive conductive bacteria coupling zero-valent iron coupling technology to reduce costs and carbon emissions, and to improve anaerobic denitrification efficiency and zero-valent iron electron utilization.

[0008] Based on the above reasons, and in view of the problems or defects existing in the prior art, the purpose of this invention is to provide a method for achieving zero carbon denitrification in water by coupling conductive bacteria with zero-valent iron, thereby solving or at least partially solving the aforementioned technical defects existing in the prior art.

[0009] To achieve the above-mentioned objectives of the present invention, the technical solution adopted by the present invention is as follows:

[0010] A method for achieving zero-carbon denitrification of water by coupling conductive bacteria with zero-valent iron, the method specifically includes the following steps:

[0011] The polluted water containing nitrate nitrogen is placed in a reactor. Then, conductive bacteria and zero-valent iron are added to the polluted water according to the specified ratio. Next, denitrification model microorganisms or anaerobic denitrification sludge are added. Finally, the oxygen in the reactor is removed and the reactor is sealed. The reactor is then subjected to constant temperature and vibration at 20-35℃ for 2-20 days. No additional organic carbon source is added to the reactor.

[0012] Furthermore, in the above-mentioned technical solution, the concentration of nitrate nitrogen in the polluted water body is between 28-70 ppm. Nitrate nitrogen refers to the nitrogen element contained in nitrates.

[0013] Furthermore, in a preferred embodiment of the present invention, the conductive bacteria described above are Shewanella oneidensis MR-1.

[0014] Furthermore, in the above technical solution, the *Shewanella onenei* MR-1 is added in the form of *Shewanella onenei* MR-1 bacterial suspension. In a preferred embodiment of the present invention, the *Shewanella onenei* MR-1 bacterial suspension is cultured using the following method:

[0015] Shewanella onychomycosis MR-1 was placed in LB medium and cultured at 30°C and 220 rpm for 12 h. It was then collected by centrifugation and the residual culture medium was washed with physiological saline. The LB medium was formulated as follows: 10 g / L tryptone, 5 g / L yeast extract, and 5 g / L sodium chloride.

[0016] Furthermore, in the above technical solution, the zero-valent iron includes commercial micron-sized zero-valent iron or ball-milled zero-valent iron.

[0017] Furthermore, in the above technical solution, the commercial micron-sized zero-valent iron is a 100-300 mesh reducing iron powder. In a preferred embodiment of the present invention, the size of the zero-valent iron is 100 mesh.

[0018] Furthermore, in the above technical solution, the average particle size of the ball-milled zero-valent iron is 0.5-2 micrometers, and the ball-milled zero-valent iron is specifically prepared by ball milling micrometer-sized zero-valent iron.

[0019] Preferably, in the above technical solution, the ball milling process is as follows: the ball milling speed is 200-800 rpm, and the ball milling time is 2-6 hours. In a preferred embodiment of the present invention, the ball milling speed is 500 rpm, and the ball milling time is 4 hours.

[0020] Furthermore, in the above technical solution, the ratio of zero-valent iron to polluted water is 0.5-2 g / L. In a preferred embodiment of the present invention, the ratio of zero-valent iron to polluted water is 1 g / L.

[0021] Furthermore, in a preferred embodiment of the present invention, the denitrification model microorganism is *Paracoccus denitrificans*. The *Paracoccus denitrificans* is added in the form of a bacterial suspension. In a preferred embodiment of the present invention, the *Paracoccus denitrificans* bacterial suspension is cultured using the following method:

[0022] Denitrifying paracocci were placed in LB culture medium and cultured at 30°C and 220 rpm for 24 h. The culture medium was then collected by centrifugation and the residual culture medium was washed away with physiological saline. The LB culture medium was formulated as follows: 10 g / L tryptone, 5 g / L yeast extract, and 5 g / L sodium chloride.

[0023] Furthermore, in a preferred embodiment of the present invention, the concentration ratio of Oneydashewanella MR-1 to Paracoccus denitrifyingis in the polluted water is 2:1.

[0024] Furthermore, in the above technical solution, when using anaerobic denitrification sludge for treatment, the inoculum amount (wet weight) of the anaerobic denitrification sludge is 1-10 kg / m³. 3In a preferred embodiment of the present invention, the inoculum amount (wet weight) of the anaerobic denitrification sludge is 5 kg / m³. 3 The Oneida Shewanella MR-1OD 600 =0.08.

[0025] The technical principles involved in this invention are as follows:

[0026] In heterotrophic denitrification water treatment, the nitrogen removal efficiency is mainly limited by the availability of organic carbon, essentially due to limited electron donors. This often necessitates the addition of organic carbon sources to enhance denitrification, increasing treatment costs and carbon emissions. Therefore, it is necessary to find inexpensive and readily available inorganic electron donors to save costs and reduce carbon emissions. Zero-valent iron (ZVFe) is an inexpensive and readily available inorganic electron donor with excellent application prospects in water treatment. This invention utilizes conductive bacteria to enhance the electron utilization efficiency of ZVFe and enable its electrons to be efficiently applied in the denitrification process. Specifically, cytochrome c in the outer membrane of conductive bacteria achieves transmembrane electron transport through changes in its redox state. When bacteria come into contact with ZVFe, cytochrome c is reduced, and the acquired electrons are then transferred into the cell for redistribution through a series of conductive proteins. When conductive bacteria come into contact with other environmental functional microorganisms (such as denitrifying bacteria), a nanoporous conductive structure can be formed between them. The excess electrons of the conductive bacteria can be transferred to the environmental functional microorganisms through this conductive structure to assist them in performing their respective environmental functions. Therefore, using conductive bacteria to couple zero-valent iron can enhance the electron utilization efficiency of zero-valent iron and improve the electron selectivity of zero-valent iron during denitrification.

[0027] The experimental results of this invention show that, when the inoculum size of denitrifying bacteria and the amount of zero-valent iron added are constant, the denitrification efficiency of the system is positively correlated with the inoculum size of conductive bacteria. This is because conductive bacteria regulate the chemical and biological processes involved in nitrate production within the denitrification system. Specifically, when the inoculum size of conductive bacteria is insufficient, some zero-valent iron will react with nitrate to form ammonia nitrogen, which is detrimental to the complete denitrification of the water.

[0028] Compared with the prior art, the present invention has the following beneficial effects:

[0029] (1) The zero-valent iron used in this invention has advantages such as low cost and environmental friendliness, and is usually used as an electron donor in environmental pollution remediation processes. This invention applies it as an electron donor to the anaerobic denitrification process to replace traditional electron donors (methanol, sodium acetate, etc.) in order to achieve efficient denitrification of wastewater with low carbon-to-nitrogen ratio and reduce costs.

[0030] (2) In traditional methods, hydrogen produced by the anaerobic corrosion of zero-valent iron is usually used to provide electrons for microbial metabolism. However, due to the low solubility of hydrogen, microorganisms cannot efficiently utilize the hydrogen produced by the corrosion of zero-valent iron. Conductive bacteria can obtain electrons from zero-valent iron by inducing metal corrosion. Furthermore, conductive bacteria can transfer electrons produced by their own metabolic processes to other electron acceptors in the environment through extracellular electron transport, thereby regulating the electron flow in the environment.

[0031] (3) In traditional iron-based materials for enhancing water denitrification technology, iron-based materials are often prepared by sintering or adding modifiers. In the method of this invention, zero-valent iron does not need to be processed in a complicated way, and the material is simple and easy to obtain.

[0032] (4) This invention couples conductive bacteria with zero-valent iron, allowing the conductive bacteria to extract electrons from the zero-valent iron and directionally transfer them to the denitrifying bacteria, thereby improving the electron utilization rate of zero-valent iron and the denitrification efficiency. Furthermore, compared to traditional methods, this invention does not require additional organic electron donors, potentially reducing carbon emissions during the denitrification process. Attached Figure Description

[0033] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0034] Figure 1 The image shows the Raman spectrum of cytochrome c on the surface of the conductive bacteria used in Example 1.

[0035] Figure 2 The graph shows a comparison of the denitrification effects of three different treatment methods on water bodies: Example 2 (N2 group), Example 3 (N3 group), and Comparative Example 1 (N1 group).

[0036] Figure 3 This is a diagram showing the denitrification effect in water when the inoculation ratio of *Shewanella onychomycosis* to *Paragonimus denitrifyingus* was 2:1 in Example 3.

[0037] Figure 4 This is a diagram showing the denitrification effect in water when the inoculation ratio of *Shewanella onychomycosis* to *Paragonimus denitrifyingus* was 1:1 in Example 4.

[0038] Figure 5 The graph shows a comparison of the zero-carbon denitrification effect of the denitrifying sludge in different systems for Example 5, Comparative Example 2, and Comparative Example 3. Detailed Implementation

[0039] The present invention will be further described in detail below through implementation examples. These implementation examples are carried out based on the technology of the present invention. Detailed implementation methods and specific operating procedures are provided to illustrate the inventiveness of the present invention, but the scope of protection of the present invention is not limited to the following implementation examples.

[0040] Based on the information contained in this application, various modifications to the precise description of the invention can be readily made by those skilled in the art. It should be understood that the scope of the invention is not limited to the defined processes, properties, or components, as these embodiments and other descriptions are merely illustrative of specific aspects of the invention.

[0041] To better understand the invention and not to limit its scope, all figures indicating amounts, percentages, and other numerical values ​​used in this application should, in all cases, be understood to be modified by the word "approximately." Therefore, unless otherwise stated, the numerical parameters listed in the specification are approximate values ​​and may vary depending on the desired properties being sought. Each numerical parameter should at least be considered as obtained based on reported significant figures and through conventional rounding methods.

[0042] The equipment and raw materials used in this invention are all commercially available or commonly used in the field. Unless otherwise specified, the methods in the following embodiments are conventional methods in the field.

[0043] The *Shewanella oneidensis* MR-1 (ATCC700550) involved in the following examples was obtained from the China General Microbiological Culture Collection Center.

[0044] The Paracoccus denitrificans (ATCC 17741) involved in the following examples was obtained from Ningbo Mingzhou Biotechnology Co., Ltd.

[0045] The anaerobic denitrification sludge used in the following examples came from the biological treatment tank of the Baita Wastewater Treatment Plant in Ganzhou City, Jiangxi Province. It was acclimatized to a laboratory sequencing batch reactor (SBR) before use. The reactor had an effective volume of 10.24 L and a water exchange rate of 50%. The reactor operated intermittently with a cycle of 6 hours, including four processes: influent (3 min), reaction (345 min), sedimentation (20 min), and effluent (2 min). Mechanical agitation was used during the reaction to promote sludge granulation. 16S rRNA sequencing revealed that the dominant denitrifying microorganisms belonged to the genera Thaurera, Ohtaekwangia, and Nitrosomonas.

[0046] Example 1

[0047] Characterization of cytochrome c on the surface of conductive bacteria after contact with zero-valent iron

[0048] Conductive bacteria were cultured in combination with zero-valent iron: *Shewanella onedida* MR-1 was cultured in standard LB medium (LB medium formula: 10 g / L tryptone, 5 g / L yeast extract, 5 g / L sodium chloride) at 30°C and 200 rpm for 12 hours. After enrichment of the bacteria at 7000 rpm, 100 mL of physiological saline containing 1 g / L ball-milled zero-valent iron was added, and its OD was increased. 600 =0.1. After deoxygenation with argon gas, the sample was sealed and cultured at 30℃ and 30 rpm, with samples taken at intervals. The samples were then analyzed using a Raman spectrometer. The Raman spectrometer used a laser wavelength of 532 nm and a laser power of 40 mW. The change of the Raman spectrum of cytochrome c on its surface over time is shown below. Figure 1 As shown in the figure, it can be observed that with the extension of culture time, the characteristic peak of cytochrome c in the sample (750 cm⁻¹) becomes increasingly smaller. -1 1129cm -1 and 1584cm -1 All showed significant enhancement. Among them, 750cm -1 The characteristic peak represents the strength of the electrostatic attraction between pyrrole nitrogen atoms in cytochrome c. When cytochrome c is reduced, the ferric ion at its center is reduced to ferrous ion, and the electrostatic attraction between the pyrrole nitrogen atoms coordinated to it becomes stronger, corresponding to an enhanced Raman signal. This indicates that during the mixed culture of conductive bacteria and zero-valent iron, the cytochrome c on the surface of the conductive bacteria is reduced. (1584 cm⁻¹) -1 The intensity of the characteristic peak represents the relative content of reduced cytochrome c on the bacterial surface. It can be observed that the relative content of reduced cytochrome c on the conductive bacteria surface increases significantly with prolonged co-cultivation time of conductive bacteria and zero-valent iron. Raman spectroscopy results indicate that zero-valent iron can provide electrons to reduce cytochrome c.

[0049] Example 2

[0050] This embodiment presents a method for achieving zero-carbon denitrification of water by coupling conductive bacteria with zero-valent iron. The method specifically includes the following steps:

[0051] For the remediation of nitrate nitrogen-contaminated water, this embodiment uses a 100 mL aqueous solution of 28 ppm nitrate nitrogen as simulated wastewater. 0.1 g of 100-mesh commercial micron-sized zero-valent iron is added, along with conductive bacteria *S. oneylon* MR-1 and denitrifying *Paracoccus* bacterial solutions. After purging oxygen from the reaction flask with argon gas, the flask is sealed and placed in a constant-temperature shaker at 30 rpm at 30°C. Samples are taken periodically, and the concentrations of nitrate, nitrite, and ammonium are determined using ion chromatography and converted to nitrate nitrogen, nitrite nitrogen, and ammonia nitrogen concentrations. Wherein:

[0052] The nitrate nitrogen aqueous solution consists of 250 mg / L NaHCO3, 25 mg / L NaH2PO4, and 2 mmol / L KNO3;

[0053] The concentration ratio (inoculation ratio) of *Shewanella oneneidae* MR-1 to *Paragonimella denitrificans* in the polluted water was 2:1; specifically, the OD of *Shewanella oneneidae* MR-1 was... 600 (MR-1) = 0.16; the OD of the denitrifying paracoccus 600 (Pd) = 0.08;

[0054] The Onenaeida Shewanella MR-1 bacterial suspension was cultured using the following method:

[0055] Shewanella onychomycosis MR-1 was placed in LB medium and cultured at 30°C and 220 rpm for 12 h. It was then collected by centrifugation and the residual culture medium was washed with physiological saline. The LB medium was formulated as follows: 10 g / L tryptone, 5 g / L yeast extract, and 5 g / L sodium chloride.

[0056] The denitrifying Paracoccus bacterial culture was prepared by the following method:

[0057] Denitrifying paracocci were placed in LB culture medium and cultured at 30°C and 220 rpm for 24 h. The culture medium was then collected by centrifugation and the residual culture medium was washed away with physiological saline. The LB culture medium was formulated as follows: 10 g / L tryptone, 5 g / L yeast extract, and 5 g / L sodium chloride.

[0058] Example 3

[0059] This embodiment describes a method for achieving zero-carbon denitrification of water by coupling conductive bacteria with zero-valent iron. The method is basically the same as that in Embodiment 2, except that the zero-valent iron used in this embodiment is ball-milled zero-valent iron. The specific preparation method is as follows: 100-mesh commercial micron zero-valent iron is ball-milled at 500 rpm for 4 hours, and the average particle size of the obtained ball-milled zero-valent iron is 1.31 μm.

[0060] Comparative Example 1

[0061] This comparative example describes a method for achieving zero-carbon denitrification of water using conductive bacteria. The method is basically the same as that in Example 2, except that zero-valent iron is not added in this comparative example.

[0062] Figure 2This is a comparison chart showing the denitrification effects of three different treatment methods: Example 2 (N2 group), Example 3 (N3 group), and Comparative Example 1 (N1 group). It can be seen that the control group (N1 group) without added zero-valent iron has almost no denitrification performance, while the group with added ball-milled zero-valent iron (N3 group) can achieve complete removal of approximately 25 ppm nitrate nitrogen within 6 days. The group with added micron-sized zero-valent iron (N2 group) can achieve removal of approximately 20 ppm nitrate nitrogen, with the remaining approximately 8 ppm nitrogen accumulating as ammonia nitrogen.

[0063] Example 4

[0064] This embodiment describes a method for achieving zero-carbon denitrification of water by coupling conductive bacteria with zero-valent iron. The method is essentially the same as in Embodiment 3, except that the inoculation ratio of *S. oneylon* MR-1 to *Paracoccus denitrifyingus* in the polluted water is 1:1; the OD of *S. oneylon* MR-1... 600 (MR-1) = 0.08; the OD of the denitrifying paracoccus 600 (Pd) = 0.08.

[0065] Figure 3 This is a diagram showing the denitrification effect in water when the inoculation ratio of Oneida Shewanella MR-1 to Denitrifying Paracoccus is 2:1 in Example 3. Figure 4 This is a graph showing the denitrification effect in water when the inoculation ratio of *S. onenedashewanella* MR-1 to *Paracoccus denitrifyingus* is 1:1, as shown in Example 4. It can be observed that, under the same conditions, increasing the inoculation ratio of *S. onenedashewanella* MR-1 can effectively improve the denitrification rate and efficiency of the denitrification system.

[0066] Example 5

[0067] This embodiment presents a method for achieving zero-carbon denitrification of water by coupling conductive bacteria with zero-valent iron. The method specifically includes the following steps:

[0068] For the remediation of nitrate-contaminated water, a 100 mL aqueous solution with a nitrate nitrogen concentration of 70 ppm was used as simulated wastewater. 0.1 g of ball-milled zero-valent iron was added to the solution, and *Shewanella onedii* MR-1 was added according to OD... 600 =0.08 for inoculation, and the inoculation amount (wet weight) of anaerobic denitrification sludge is 5 kg / m³. 3 After purging oxygen from the reaction flask with argon gas, the flask was sealed and placed in a constant-temperature shaker at 30°C and 30 rpm. Samples were taken periodically, and the concentrations of nitrate, nitrite, and ammonium were determined using ion chromatography and converted to nitrate nitrogen, nitrite nitrogen, and ammonia nitrogen concentrations. Among these:

[0069] The nitrate nitrogen aqueous solution consists of 250 mg / L NaHCO3, 25 mg / L NaH2PO4, and 5 mmol / L KNO3;

[0070] The zero-valent iron used in this embodiment is ball-milled zero-valent iron, which is prepared by ball milling 100-mesh commercial micron zero-valent iron at 500 rpm for 4 hours. The average particle size of the ball-milled zero-valent iron is 1.31 μm.

[0071] Comparative Example 2

[0072] This comparative example of a method for achieving zero carbon denitrification in water using conductive bacteria is basically the same as that in Example 6, except that: no ball-milled zero-valent iron and conductive bacteria Oneida Shewanella MR-1 were added in this comparative example.

[0073] Comparative Example 3

[0074] The method for achieving zero carbon denitrification in water using conductive bacteria in this comparative example is basically the same as that in Example 6, except that the conductive bacteria Oneida Shewanella MR-1 were not added in this comparative example.

[0075] Comparative Examples 2 and 3 above were set up as control experiments with only denitrified sludge and a mixed system of denitrified sludge and ball milled zero-valent iron.

[0076] Figure 5 This is a comparative graph showing the denitrification effects of three different treatment methods: Example 5 (N3 group), Comparative Example 2 (N1 group), and Comparative Example 3 (N2 group). It can be seen that the control group (N1 group) with only denitrifying sludge showed no significant denitrification performance within 2 days. However, after adding ball-milled zero-valent iron (N2 group), approximately 20 ppm of nitrate nitrogen was removed within 2 days. When ball-milled zero-valent iron and the conductive bacteria *Shewanella onychomycosis* MR-1 were added simultaneously (N3 group), the denitrification performance of the denitrifying sludge was significantly enhanced, achieving complete removal of approximately 65 ppm of nitrate nitrogen within 2 days.

Claims

1. A method for achieving zero-carbon denitrification of water by coupling conductive bacteria with zero-valent iron, characterized in that: The method specifically includes the following steps: The polluted water containing nitrate nitrogen is placed in a reactor. Then, conductive bacteria and zero-valent iron are added to the polluted water according to the specified ratio. Next, denitrification model microorganisms or anaerobic denitrification sludge are added. Finally, the oxygen in the reactor is removed and the reactor is sealed. The reactor is then subjected to constant temperature and vibration at 20-35℃ for 2-20 days. No additional organic carbon source is added to the reactor. The ratio of zero-valent iron to polluted water is 0.5-2 g / L. The conductive bacteria are *Schizella onychomycosis* MR-1; the denitrification model microorganism is *Paracococcus denitrifyingus*; when using the denitrification model microorganism for treatment, the concentration ratio of *Schizella onychomycosis* MR-1 to *Paracococcus denitrifyingus* in the polluted water is 2:1; when using anaerobic denitrification sludge for treatment, the inoculum amount of the anaerobic denitrification sludge, on a wet weight basis, is 5 kg / m³. 3 The Onenaeida Shewanella MR-1 OD 600 =0.

08.

2. The method according to claim 1, characterized in that: The concentrations of nitrate nitrogen in the polluted water bodies were all between 28 and 70 ppm.

3. The method according to claim 1, characterized in that: The ratio of zero-valent iron to polluted water is 1 g / L.

4. The method according to claim 1, characterized in that: The zero-valent iron includes commercial micron-sized zero-valent iron or ball-milled zero-valent iron.

5. The method according to claim 4, characterized in that: The commercial micron-sized zero-valent iron is 100-300 mesh reducing iron powder.

6. The method according to claim 4, characterized in that: The average particle size of the ball-milled zero-valent iron is 0.5-2 micrometers.

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