Strain with dissimilatory nitrate reducing capacity and application thereof

Abstract

The invention belongs to the technical field of microbial system classification and environmental biology, and particularly relates to a strain with dissimilatory nitrate reducing capacity and application of the strain. The bacterial strain with the dissimilatory nitrate reducing capacity is named shewanella sp. AS1, is preserved in Guangdong Microbial Culture Collection Center (GDMCC) of Institute of Microbiology, Guangdong Academy of Sciences, located on the fifth floor, No. 59, No. 100 Courtyard, Xianlie Middle Road, Guangzhou City on May 19, 2025, and has the preservation number of GDMCC No: 66362. The AS1 strain has the outstanding capability of dissimilatory reduction of nitrate into ammonia, can gradually reduce nitrate (NO3 <->) into ammonium (NH4 < + >) in an anaerobic environment by using nitrate (NO3 <->) as a terminal electron acceptor, and has a wide application prospect in the aspects of improvement and research of various environmental problems caused by environmental pollution remediation and large loss of nitrogen.

Description

Technical Field

[0001] The invention belongs to the fields of microbial system classification and environmental biotechnology, and particularly relates to a strain with nitrate dissimilatory reduction ability and application thereof. Background Art

[0002] Mangroves are wetland woody plant communities composed primarily of evergreen trees or shrubs, growing in the intertidal zones of tropical and subtropical coasts. Mangrove ecosystems are the most distinctive wetland ecosystems, combining characteristics of both terrestrial and marine ecosystems. Their naturally alternating aerobic / anaerobic environment fosters a rich reservoir of extracellular respiratory microorganisms. Furthermore, mangroves play a vital role in purifying seawater, providing wind and wave protection, carbon sequestration, and maintaining biodiversity. Known as the "Guardian of the Coast" and the "Green Lungs of the Ocean," mangroves also serve as a crucial habitat for rare and endangered waterfowl.

[0003] Shewanella is a facultative anaerobic bacterium with extracellular respiration, and is widely distributed in nature. Currently, researchers have isolated different members of this genus from a variety of sources, including activated sludge, seawater, sediments, invertebrates, and fish. With the rapid growth of the global population, human activities are changing the global nitrogen cycle at an unprecedented rate. Among them, the large-scale burning of fossil fuels, the sharp increase in nitrogen demand in agriculture and industry, and the over-reliance on inorganic nitrogen fertilizers in crop production have caused global agricultural inorganic nitrogen use to soar to 110 TgN / year. What is worrying is that in agricultural production systems, nitrogen utilization efficiency is low. For every unit of active nitrogen used, only 4-14% can be effectively utilized, and nearly half of the active nitrogen is lost to the environment during fertilization. Large-scale nitrogen loss can cause various environmental problems: (1) Soil acidification is exacerbated, eutrophication of water bodies is frequent, and the resulting ecological disasters such as algal blooms or red tides seriously damage the health of aquatic ecosystems; (2) Excessive emissions of nitrogen oxides not only exacerbate global warming, but may also form acid rain, which in turn affects soil and water quality; (3) Excessive nitrogen input from agricultural fertilization and animal husbandry can inhibit the growth of certain plant species, reduce biodiversity, and thus seriously threaten the ecological environment. Studies have shown that some members of the genus Shewanella can convert nitrate into nitrite, and then convert nitrite into ammonia nitrogen. Therefore, it has great application potential in soil environments to prevent nitrogen loss.

[0004] Mangrove ecosystems harbor a rich resource of bacterial strains capable of dissimilatory nitrate reduction, but their utilization is low and has yet to be fully explored. Currently, relatively few studies have examined the pure culture diversity of Shewanella species in mangrove ecosystems. The only pure culture strains of Shewanella isolated from mangroves are a humus-reducing bacterium (Shewanella sp.) W3 discovered by Wu Peng et al. ("Isolation and Humic Substance-Reducing Properties of Shewanella sp. W3 from Mangrove Wetlands." Environmental Science 31.4 (2010): 1041-1046) and a new species of Shewanella mangrovi isolated by Liu et al. ("Shewanella mangrovi sp. nov., an acetaldehyde-degrading bacterium isolated from mangrove sediment." (2015): 2630-2634). Therefore, further research is needed to identify Shewanella species with excellent dissimilatory nitrate reduction abilities. Summary of the Invention

[0005] To overcome the shortcomings of the above-mentioned existing technologies, the present invention isolated and screened a new species of Shewanella from mangrove sediments in Futian District, Shenzhen City, Guangdong Province, and named Shewanella sp. AS1. This bacterium has the ability of dissimilatory nitrate reduction and has broad application prospects in the research of environmental pollution remediation and the treatment of various environmental problems caused by large-scale nitrogen loss.

[0006] In order to achieve the above object, the technical solution adopted by the present invention is:

[0007] In a first aspect, the present invention provides a Shewanella sp. AS1 strain, which was deposited in the Guangdong Provincial Microbiological Culture Collection Center on May 19, 2025, with a deposit number of GDMCC No: 66362; the complete 16S rDNA sequence of the Shewanella AS1 strain is shown in SEQ ID No: 1.

[0008] The Shewanella sp. AS1 strain of the present invention was isolated and screened from mangrove sediment in Futian District, Shenzhen City, Guangdong Province. The strain appears as a pink, rounded form with neat edges and a smooth, moist surface on 2216E solid culture medium. Observation under a transmission electron microscope revealed that the strain is rod-shaped, approximately 0.61 to 0.78 μm wide and 1.03 to 1.90 μm long, lacking peritrichous flagella. The Shewanella sp. AS1 strain is a facultative anaerobic, Gram-negative bacterium. It is positive for catalase, oxidase, and Tween 80 hydrolysis, but negative for urease activity, unable to hydrolyze starch and casein. It is positive for β-glucosidase and can metabolize arabinose and N-acetyl-glucosamine as substrates. Drug sensitivity testing showed that the AS1 strain is sensitive to chloramphenicol, erythromycin, gentamicin, neomycin, ciprofloxacin, and norfloxacin, but resistant to clindamycin, lincomycin, and carbenicillin. The genome of Shewanella sp. AS1 strain was used as a template and amplified with universal primers 27F and 1492R for 16S rDNA. The obtained 16S rDNA sequence was compared with the EzBioCloud database for homology. The results showed that the strain with the highest homology to this strain was Shewanella insulae JBTF-M18. T The similarity was 96.4%. A phylogenetic tree was constructed using MEGAX software based on the 16s rRNA sequence of the strain, preliminarily identifying the strain as a new species of the genus Shewanella. The strain can grow in 2216E medium at temperatures ranging from 4 to 40°C, with an optimal growth temperature of 30°C. It can tolerate a NaCl concentration range of 1 to 10% (w / v), with an optimal NaCl concentration of 3% (w / v). The pH range for growth is 5 to 9, with an optimal pH of 6. Furthermore, Shewanella sp. AS1 was identified using API 20NE reagent strips, indicating that the strain is capable of nitrate reduction. Furthermore, further genomic analysis of the strain revealed data supporting the ability to perform dissimilatory nitrate reduction to ammonia.

[0009] The second aspect of the present invention also provides the use of Shewanella sp. AS1 strain in the dissimilatory reduction of nitrate to ammonia.

[0010] The third aspect of the present invention further provides the use of Shewanella sp. AS1 strain in environmental pollution remediation and / or soil improvement.

[0011] Preferably, the environmental pollution remediation includes removing excess nitrates from water bodies and reducing greenhouse gas emissions; and the soil improvement is to repair nitrate-contaminated soil or inhibit soil nitrogen loss.

[0012] Preferably, the suitable growth temperature of Shewanella AS1 strain is 4-40°C.

[0013] Preferably, the NaCl concentration range that the Shewanella AS1 strain tolerates is 1-10% by mass.

[0014] The fourth aspect of the present invention further provides a microbial agent having the ability of dissimilatory nitrate reduction, wherein the microbial agent uses Shewanella sp. AS1 strain as a main active ingredient.

[0015] Preferably, the Shewanella AS1 strain in the bacterial agent is in a logarithmic growth phase.

[0016] Preferably, the concentration of Shewanella AS1 strain in the bacterial agent is OD 600 =0.8-1.5.

[0017] Preferably, the microbial agent further comprises microbiologically acceptable excipients, which must be compatible with the microbial cells to ensure safety and efficiency.

[0018] More preferably, the excipients include carriers (such as humus, bentonite; used to adsorb bacteria for easy storage and application), protective agents (such as trehalose, phosphates; used to maintain activity and resist damage from the external environment), synergists (such as alginic acid, enzyme preparations; used to synergistically enhance the function of the bacterial agent), and dosage form adjuvants (such as wetting agents, adhesives; used to optimize the application form).

[0019] Compared with the prior art, the present invention has the following beneficial effects:

[0020] The present invention provides a new species of Shewanella with the ability to dissimilatory reduction of nitrate to ammonia, named Shewanella sp. AS1. The bacterium was isolated and screened from mangrove sediments in Futian District, Shenzhen City, Guangdong Province. It was deposited in the Guangdong Microbial Culture Collection Center (GDMCC) at the Institute of Microbiology, Guangdong Academy of Sciences, Building 59, 5th Floor, No. 100 Xianlie Middle Road, Guangzhou City on May 19, 2025, with a deposit number of GDMCC No: 66362. The bacterium not only has broad application prospects in the research of microbial dissimilatory reduction of nitrate to ammonia. Moreover, since the bacterium can utilize nitrate (NO3 - ) as the terminal electron acceptor, which is gradually reduced to ammonium (NH4 +), so it also has broad application prospects in the research of environmental pollution remediation and the treatment of various environmental problems caused by large-scale nitrogen loss. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 This is the colony morphology of Shewanella sp. AS1 strain.

[0022] Figure 2 This is a transmission electron micrograph of Shewanella sp. strain AS1.

[0023] Figure 3 This is the phylogenetic tree of Shewanella sp. AS1 strain based on 16S rRNA sequence.

[0024] Figure 4 This is the sequence map of Shewanella sp. AS1 strain based on the whole genome.

[0025] Figure 5 Schematic diagram of the dissimilatory reduction of nitrate to ammonia by Shewanella sp. AS1 strain.

[0026] Figure 6 This is a graph showing the changes in nitric nitrogen concentration monitored during the cultivation of Shewanella sp. AS1 strain in LB liquid culture medium containing nitrate. DETAILED DESCRIPTION

[0027] The following is a further description of specific embodiments of the present invention. It should be noted that the description of these embodiments is intended to facilitate understanding of the present invention and does not constitute a limitation of the present invention. In addition, the technical features involved in the various embodiments of the present invention described below may be combined with each other as long as they do not conflict with each other.

[0028] The experimental methods in the following examples are conventional methods unless otherwise specified, and the experimental materials used in the following examples are commercially available unless otherwise specified.

[0029] Example 1: Isolation and identification of Shewanella sp. AS1 strain

[0030] (1) Preparation of culture medium

[0031] 2216E liquid medium: 5 g / L tryptone, 1 g / L yeast extract, 25 g / L artificial seawater, pH = 7.6-7.8. Solid medium was prepared by adding 2% (w / v) agar powder to the above fresh medium.

[0032] (2) Isolation of strains

[0033] 2 g of mangrove sediment sample collected from the Mangrove Ecological Protection Area, Futian District, Shenzhen City, Guangdong Province (22°30′-22°32′N, 113°56′-114°3′E) was weighed and placed in 10 mL of sterile saline and shaken vigorously. After standing, the supernatant was taken for gradient dilution (10 -1 ~10 -5 ), take 100 μL of dilution (10 -3 ~10 -5 ) was evenly spread on 2216E solid culture medium and inverted in a constant temperature incubator at 30°C for 7 days. After a single colony grew on the plate, the colony morphology was observed, and single colonies of different morphologies were randomly selected with an inoculation loop and inoculated onto a new plate for further purification. The purified colonies were then inoculated into fresh 2216E liquid culture medium and placed in a constant temperature shaking incubator (30°C, 180rpm) for 24 hours. After the obtained strain was streaked on 2216E solid culture medium and incubated at 30°C for 48 hours, the single colonies formed on the plate were pink and round with neat edges and a smooth and moist surface ( Figure 1 At the same time, under the transmission electron microscope, the bacteria were observed to be rod-shaped, about 0.61 to 0.78 μm wide and 1.03 to 1.90 μm long, with peritrichous flagella ( Figure 2 The isolated strain was then labeled AS1 and inoculated into 2216E liquid culture medium for subsequent experiments. Furthermore, the strain was stored at -80°C with 10% (v / v) glycerol for long-term storage.

[0034] (3) 16S rDNA identification of strain AS1

[0035] use Genomic DNA was extracted using a genomic DNA purification kit (Promega, USA) and quantified using a TBS-380 fluorometer (Turner Biosystems, Sunnyvale, CA). TMThe Rapid DNA-Seq kit and Illumina platform were used for library construction and library testing. Subsequently, the qualified library was sequenced on both ends using the Illumina HiSeqX10 sequencer, with a sequencing end length of 150bp to obtain the original data sequence. Some low-quality reads in the original sequence were trimmed using fastp software (https: / / github.com / OpenGene / fastp, version 0.20.0), and then sequence assembly was performed using SOAPdenovo 2 software. The genome of the AS1 strain was then used as a template, and the 16SrDNA universal primers 27F (agagtttgatcctggctca) and 1492R (ggttaccttg ttacgactt) were used for amplification to obtain its 16SrDNA sequence (as shown in SEQ ID NO.1). Finally, the sequence was uploaded to the EzBioCloud database, and the results showed that the strain had the highest homology with the strain Shewanella insulae JBTF-M18. T The 16S rRNA similarity of the strain was the highest, at 96.4%, which was lower than the threshold of 98.7% for identifying new species (for the definition of the threshold of new species, see the reference "Beye, Mamadou, et al." Careful use of 16SrRNA gene sequence similarity values ​​for the identification of Mycobacterium species." New microbes and new infections 22(2018):24-29."). To this end, the phylogenetic tree (such as Figure 3 As shown in Figure 3 , strain AS1 was found to be distinct from other Shewanella clades, forming an independent branch. Therefore, it was preliminarily identified as belonging to the genus Shewanella sp. and named Shewanella sp. AS1.

[0036] 16S rDNA sequence of Shewanella sp. AS1 (SEQ ID NO. 1):

[0037]

[0038] (4) Analysis of physiological and biochemical characteristics of Shewanella sp. AS1

[0039] In the Gram staining experiment, the bacterial solution is first smeared on a clean glass slide, allowed to dry naturally, fixed by flame 2-3 times, and then observed under an optical microscope after primary staining with crystal violet, mordanting with iodine solution, decolorizing with ethanol, and counter-staining with safranin. The growth experiment of the strain is as follows: the Shewanella sp. AS1 strain cultured to the logarithmic growth phase is inoculated into 2216E culture medium at a 1% inoculum, and the test strain is grown for one week at different temperatures (0, 4°C, 10°C, 15°C, 20°C, 25°C, 30°C, 37°C, 42°C). The NaCl tolerance growth test of the strain is also carried out in 2216E culture medium containing 0-12% (w / v) NaCl (NaCl concentration interval of 1%, w / v). At the same time, in order to determine the growth pH range of the strain in 2216E culture medium, the pH range is set to 3-10 (pH is separated by 1 pH unit). Three parallel samples are set for the above experiments, and samples are taken every 12 hours to measure the OD 600 Enzyme activity assays were performed on 2216E solid medium containing hydrogen peroxide, starch, casein, and Tween 80 (see the reference "Dong, X. & Cai, M. (2001). Manual of Systematic and Determinative Bacteriology. Beijing: Academic Press."). Other physiological and biochemical characteristics were assessed using the API 20NE kit (BioMérieux, France). Antimicrobial susceptibility testing was also performed using the disc method.

[0040] Identification results are as follows: Shewanella sp. strain AS1 is a Gram-negative bacterium that can grow in temperatures ranging from 4 to 40°C, with an optimal growth temperature of 30°C. It can tolerate a NaCl concentration range of 1 to 10% (w / v), with an optimal growth concentration of 3% (w / v). Its pH range is 5 to 9, with an optimal pH of 6. It is positive for oxidase and catalase activities and can hydrolyze Tween 80, but not starch or casein. Analysis using API 20NE reagent strips revealed that strain AS1T is capable of nitrate reduction and is positive for β-glucosidase, metabolizing arabinose and N-acetyl-glucosamine. Antimicrobial susceptibility testing revealed that the bacterium is sensitive to rifampicin, neomycin, erythromycin, gentamicin, chloramphenicol, polymyxin B, penicillin, ciprofloxacin, norfloxacin, and ofloxacin, but is resistant to novobiocin, clindamycin, carbenicillin, kanamycin, and lincomycin. Shewanella sp. AS1 strain and its closely related model strain Shewanella insulae JBTF-M18 T The morphological and physiological and biochemical characteristics of the samples are compared in Table 1.

[0041] In addition, Shewanella sp. AS1 strain and type strain Shewanella insulae JBTF-M18 T The strains also exhibited the following characteristics: both were Gram-negative and positive for catalase, oxidase, and Tween 80 hydrolysis, but negative for urease activity. In antimicrobial susceptibility testing, both strains were sensitive to chloramphenicol, erythromycin, gentamicin, neomycin, ciprofloxacin, and norfloxacin, but resistant to clindamycin, lincomycin, and carbenicillin.

[0042] Table 1: Comparison of morphological and physiological and biochemical characteristics of Shewanella sp. AS1 strain and its related strains

[0043]

[0044]

[0045] Note: + indicates positive, w indicates weakly positive, - indicates negative, S: sensitive, R: resistant, NR indicates not reported.

[0046] a: The data of model bacteria are from the following literature:

[0047] S. Park, IKKim, W. Kim, J.-H. Yoon. (2020) Shewanella insulae sp. nov., isolated from a tidal flat. Int. J. Syst. Evol. Microbiol. 70, 3872-3877.

[0048] (5) Fatty acid characteristics analysis of Shewanella sp. AS1 strain

[0049] The Shewanella sp. AS1 strain and the model bacteria Shewanella insulae JBTF-M18 T (Purchased from the Agricultural Microorganism Culture Collection Center of the National Institute of Agro-biotechnology of Korea, KACC) entrusted the Xiamen Marine Microorganism Culture Collection Management Center to identify the fatty acid content. Sample pre-treatment was as follows: the strain was cultured in 2216E medium to the late logarithmic growth phase, then centrifuged at 6000g for 10min at room temperature to collect the bacteria, washed the bacteria once with physiological saline, placed in a freeze dryer for freeze drying, and sent for testing after obtaining 1g of freeze-dried bacteria. Fatty acid detection was carried out according to the MIDI standard (Shorock Microbial Identification System, Version 6.0B), and the fatty acids were subjected to the steps of saponification, methylation, extraction and washing in sequence. The experimental instrument used was a gas chromatograph (Agilent6850, USA), and identification was carried out using the TSBA6.0 database of the microbial identification system.

[0050] The fatty acid analysis results in Table 2 show that the major fatty acids (>5%) of Shewanella sp. AS1 strain are C15:0-iso (23.5%), Summed Feature 3 (14.8%), C17:1ω8c3 (14.6%), C13:0iso (9.2%), and C16:0 (8.3%). T Compared with the model strain, the fatty acid content ratio is roughly similar, but the C16:0 content of Shewanella sp. AS1 strain is significantly higher than that of the model strain, while the C13:0-iso 3OH content is significantly lower than that of the model strain. The above results prove that the Shewanella sp. AS1 strain of the present invention is different from the model strain Shewanella insulae JBTF-M18 TTable 2: Shewanella sp. AS1 strain and its related strain Shewanella insulae JBTF-M18 T Comparative results of fatty acid analysis

[0051]

[0052] In the table, SummedFeature a Represents two or three fatty acid groups that cannot be separated by gas-liquid chromatography, SummedFeature 1:iso-C15:1-H / C13:03-OH; SummedFeature 3:C16:1ω7c / C16:1ω6c; SummedFeature 8:C18:1ω7c / C18:1ω6c.

[0053] (6) Molecular taxonomic status of Shewanella sp. AS1 strain

[0054] To further determine the molecular taxonomic status of Shewanella sp. AS1, the genome of AS1 was analyzed in comparison with other bacteria in the genus Shewanella. The ANI calculator and Genome-to-Genome Distance Calculator (GGDC) server of the EzBiocloud data platform were used to perform a genome sequence-based phylogenetic analysis of Shewanella sp. AS1. The results showed that ( Figure 4 ), strain AS1 is located in a branch distinct from other bacteria and belongs to the genus Shewanella. Digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) were also calculated between the genomes of Shewanella sp. AS1 and its closely related species (see Table 3). The results showed that the dDDH values ​​of strain AS1 ranged from 19.8% to 21.3%, well below the conspecific similarity threshold of 70%, and the ANI values ​​were also below the 95% threshold. This result strongly suggests that Shewanella sp. AS1 is a new, independent species.

[0055] Table 3: Calculated values ​​of ddDH and ANI between Shewanella sp. AS1 strain and its closely related species

[0056]

[0057] In summary, combining morphological, physiological and biochemical characteristics, fatty acid chemical classification and molecular biology classification techniques, it can be determined that Shewanella sp. AS1 strain is a new species of the genus Shewanella.

[0058] The Shewanella sp. AS1 strain was deposited in the Guangdong Microbial Culture Collection (GDMCC) located at the Institute of Microbiology, Guangdong Academy of Sciences, 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou on May 19, 2025, with the deposit number GDMCC No: 66362.

[0059] Example 2: Ability of Shewanella sp. AS1 strain to dissimilatory reduction of nitrate to ammonia under anaerobic conditions

[0060] The Shewanella sp. AS1 strain was inoculated into LB liquid medium and cultured in a constant temperature shaker (180 rpm, 30°C) until the logarithmic growth phase (OD 600 =0.8~1.0), then high-speed centrifugation (6000rpm, 10min), remove the supernatant, resuspend the bacteria with sterile 0.9% NaCl, repeat the above steps twice, thoroughly wash the residual culture medium of the bacteria, and finally resuspend the bacterial solution with sterile culture medium to OD600≈1 to obtain the bacterial solution. Add 80mL of nitrate reduction medium to a 100mL penicillin bottle, fill it with high-purity nitrogen to deoxygenate for more than 45min, seal it with butyl rubber stopper and aluminum cap, and sterilize it at 121℃ for 20min. After the culture medium cools to room temperature, inoculate the bacterial suspension into anaerobic culture medium at a 1% inoculation amount and place it in a 30℃ incubator for culture. Take samples on time and measure NO3 at 220nm and 275nm wavelengths according to the national standard HJ / T 346-2007. - The concentration of NO2 was determined at a wavelength of 540 nm using a Griess kit (Feijing Biotechnology, Fuzhou, China). - The concentration was measured using a Nessler kit (Guangdong Huankai Biotechnology Co., Ltd.) to determine the ammonia nitrogen content. This was used to monitor the ability of Shewanella sp. AS1 strain to dissimilatoryly reduce nitrate to ammonia.

[0061] The LB liquid medium is formulated as follows: 10 g / L tryptone, 5 g / L yeast extract, 10 g / L sodium chloride, pH = 7.4, and the solvent is water. The nitrate-reducing medium (DM) is formulated as follows (1 L): 1.8 g KNO3, 7.0 g K2HPO4, 3.0 g KH2PO4, 0.05 g FeSO4·7H2O, 0.1 g MgSO4·7H2O, 10.0 g sodium lactate, pH = 7.0-7.2, and the solvent is water. High-purity nitrogen is used to deoxygenate the medium for more than 45 minutes, followed by sealing with a butyl rubber stopper and an aluminum cap. Finally, the medium is sterilized by autoclaving at 121°C for 20 minutes.

[0062] Figure 5 、 6 The results showed that Shewanella sp. AS1 strain can utilize nitrate (NO3 - ) as the terminal electron acceptor, which is gradually reduced to ammonium (NH4 + ), indicating that Shewanella sp. AS1 has the ability to dissimilatory reduce nitrate to ammonia in a nitrate-reducing medium. Specifically, after culturing Shewanella sp. AS1 to the logarithmic growth phase and reacting in a nitrate-reducing system for 30 hours, it completely consumes nitrate nitrogen and converts it to ammonia nitrogen. The concentration of the reduced ammonia nitrogen can reach approximately 50 mg / L, and the efficiency of the reduced ammonia nitrogen can reach 70%.

[0063] In summary, the Shewanella sp. AS1 strain of the present invention has an outstanding ability to dissimilatory reduction of nitrate to ammonia, and can utilize nitrate (NO3 - ) as the terminal electron acceptor, which is gradually reduced to ammonium (NH4 + ), and has broad application prospects in the research on environmental pollution remediation and the management of various environmental problems caused by large-scale nitrogen loss.

[0064] The embodiments of the present invention are described in detail above, but the present invention is not limited to the described embodiments. It is apparent to those skilled in the art that various changes, modifications, substitutions, and variations of these embodiments may be made without departing from the principles and spirit of the present invention, and the changes still fall within the scope of protection of the present invention.

Claims

1. A Shewanella sp. AS1 strain, characterized in that: The Shewanella AS1 strain was deposited in the Guangdong Provincial Microbial Culture Collection Center on May 19, 2025, with a deposit number of GDMCC No: 66362; the complete 16S rDNA sequence of the Shewanella AS1 strain is shown in SEQ ID No:

1.

2. Use of the Shewanella sp. AS1 strain according to claim 1 in the dissimilatory reduction of nitrate to ammonia.

3. Use of the Shewanella sp. AS1 strain according to claim 1 in environmental pollution remediation and / or soil improvement.

4. The use according to claim 3, characterized in that The environmental pollution remediation includes removing excess nitrates from water bodies and reducing greenhouse gas emissions; the soil improvement is to repair nitrate-contaminated soil or inhibit soil nitrogen loss.

5. The use according to any one of claims 2 to 4, characterized in that: The optimum growth temperature for Shewanella AS1 strain is 4-40℃.

6. The use according to any one of claims 2 to 4, characterized in that: The NaCl concentration range that Shewanella AS1 strain tolerates is 1-10% by mass fraction.

7. A microbial agent having nitrate dissimilatory reduction ability, characterized in that: The microbial agent uses the Shewanella sp. AS1 strain described in claim 1 as a main active ingredient.

8. The microbial agent having nitrate dissimilatory reduction ability according to claim 7, characterized in that: The Shewanella AS1 strain in the inoculum was in the logarithmic growth phase.

9. The microbial agent having nitrate dissimilatory reduction ability according to claim 7, characterized in that: The concentration of Shewanella AS1 strain in the inoculum was OD 600 =0.8-1.

5.

10. The microbial agent having nitrate dissimilatory reduction ability according to claim 7, characterized in that: The microbial agent further comprises microbiologically acceptable excipients.

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