Urea-producing arthrobacter degrading atrazine and application thereof

Abstract

The application belongs to the technical field of biological repair, and relates to a urea-producing Arthrobacter ADDB-13 capable of effectively degrading atrazine, which is preserved in the China Center for Type Culture Collection and has a preservation number of CCTCC NO: M20242925. The bacterial suspension obtained based on the strain is cultured in an inorganic salt culture medium added with 100 mg / L of atrazine for 70 h, and the atrazine degradation rate reaches 99.90%, so that the bacterial suspension can be used for repairing soil containing atrazine residues and promoting the growth of crops stressed by atrazine.

Description

Technical Field

[0001] This invention belongs to the field of bioremediation technology and relates to a urea-producing strain of Atrazine that can effectively degrade atrazine and its application. Background Technology

[0002] Atrazine is a triazine herbicide, chemically named 2-chloro-4-ethylamino-6-isopropylamino-1,3,5-triazine, widely used in the cultivation of broadleaf and grassy weeds in crops such as corn, rice, wheat, and soybeans. As a herbicide used worldwide for a long time, atrazine has advantages such as low price, high weeding efficiency, and the ability to control a wide variety of weeds. However, it also has a long half-life in soil, is difficult to degrade, and has strong migration properties, resulting in significant environmental pollution. Atrazine also has significant phytotoxicity on wheat, soybeans, sweet potatoes, and other crops sensitive to late-blooming growth, causing wheat leaves to wither and grains to be smaller; inhibiting soybean growth and reducing soybean yield; and causing sweet potato leaves to turn yellow and plants to be stunted. When plants absorb atrazine, it can be transferred through the food chain to aquatic animals, terrestrial animals, and humans, causing damage to the nervous, endocrine, immune, and genetic systems, and even cancer.

[0003] Microbial degradation is a recognized environmentally friendly method for removing pollutants. Many types of microorganisms can effectively degrade atrazine, including *Arthrobacter* spp. (…). Arthrobacter sp.), genus Citrullococcus ( Citricoccus sp.), Pseudomonas spp. Pseudomonas sp.), Acinetobacter spp. Acinetobacter (sp.), etc. According to statistics, the main pathways of microbial degradation of atrazine include dechlorination, dealkylation, hydroxylation, and ring cleavage, each of which is controlled by different degradation genes atzA to atzF.

[0004] In recent years, although scholars from various countries have screened out a variety of strains that can efficiently degrade atrazine, there are relatively few strains that are suitable for various environments and can be practically applied to soil. Therefore, it is still necessary to continuously enrich the strain library in order to produce microbial agents that can be applied to the field.

[0005] He Bipo et al. (Isolation and identification of atrazine-degrading bacterium SFAD3 and its soil remediation potential [J]) Plant protection Nursing Journal (2023, 50(4): 1096-1104.) reported a *Pseudomonas mendoza* SFAD3 strain for atrazine degradation, which achieved a 72.6% degradation rate of atrazine in contaminated soil after 30 days of culture; Lü Huiping (Isolation and Identification of Highly Efficient Atrazine Degrading Bacteria [J]. Chemical Technology and Development, 2017, 40(2): 22-24.) reported the atrazine degradation rate of Agrobacterium AT-2, Brucella HB-1, Staphylococcus HB-7, and Pseudomonas HW-4. The degradation efficiency was only about 40% after 24 hours, but reached about 90% after 3 days. Summary of the Invention

[0006] The purpose of this invention is to provide a strain of Ureaplasma urealyticum (Ureaplasma urealyticum). Paenarthrobacter ureafaciens The strains were selected to effectively degrade atrazine, providing microbial resources for the environmental remediation of triazine herbicides.

[0007] To achieve the above-mentioned objectives, this invention provides a strain of Ureaplasma urealyticum that effectively degrades atrazine. Specifically, this strain is Ureaplasma urealyticum ADBD-13, which was deposited on December 27, 2024, at the China Center for Type Culture Collection, Wuhan, China, with accession number CCTCC NO: M20242925.

[0008] The *Arthrobacter ureagenes* ADBD13 strain described in this invention was screened from cornfield soil in Pingyuan County, Dezhou City, Shandong Province. Based on morphological, physiological, biochemical, and molecular identification results, this strain was determined to belong to *Arthrobacter ureagenes* (…). Paenarthrobacter ureafaciens ).

[0009] Further strain performance testing confirmed that the screened Arthrobacter ureagenesis ADBD-13 has a significant ability to degrade atrazine and can be used as an atrazine degrading agent.

[0010] Furthermore, the Ureaplasma urealyticum ADBD-13 described in this invention can be specifically applied to the remediation of soil containing atrazine residues.

[0011] Furthermore, the *Arthrobacter urealyticum* ADBD-13 described in this invention also has the effect of promoting the growth of crops under atrazine stress, including but not limited to corn or wheat.

[0012] Based on the aforementioned effects of *Arthrobacter ureagenesis* ADBD-13, this invention further provides a *Arthrobacter ureagenesis* bacterial suspension that degrades atrazine, specifically prepared with an inorganic salt culture medium containing 1 × 10⁻⁶ *Arthrobacter ureagenesis* ADBD-13. 9 CFU / mL bacterial suspension.

[0013] Specifically, the *Arthrobacter ureaplasma* ADBD-13 bacterial suspension is prepared by inoculating selected single colonies of *Arthrobacter ureaplasma* ADBD-13 into LB liquid medium, incubating at 30°C and 200 rpm for 16–20 h with shaking, centrifuging the bacterial suspension, collecting the precipitate, washing with inorganic salt medium, and then preparing a 1×10⁻⁶ concentration using inorganic salt medium. 9CFU / mL bacterial suspension.

[0014] The inorganic salt culture medium consists of: 0.45g KH2PO4, 1.79g K2HPO4, 0.4g NaCl, 0.2g MgSO4·7H2O, diluted with distilled water to 1000mL, pH 7.0.

[0015] The *Arthrobacter ureagenesis* suspension prepared by this invention can be used as an atrazine degrading agent.

[0016] Furthermore, the *Arthrobacter ureagenesis* suspension can be used as a soil remediation agent for atrazine residues in the bioremediation of soils containing atrazine residues.

[0017] Furthermore, the *Arthrobacter ureagenesis* suspension described in this invention also has a bioremediation effect on wheat and corn seedlings in soil containing atrazine residues.

[0018] Experiments have shown that soil remediated with the *Arthrobacter urealyticum* suspension described in this invention can effectively promote the growth of atrazine-stressed wheat and corn seedlings.

[0019] More specifically, the present invention preferably uses the Ureaplasma urealyticum suspension after a 50-fold volume dilution (i.e., a final concentration of 2% of the original suspension, v / v).

[0020] This invention screened and obtained a strain of Ureaplasma urealyticum ADBD-13 that can effectively degrade atrazine. The degradation rate of atrazine in inorganic salt culture medium reached 99.90%, and the degradation rate in soil containing atrazine residue reached 70.54% after 7 days of treatment. It has significant biodegradation effect on atrazine and can effectively remediate soil with atrazine residue.

[0021] This invention also experimentally demonstrated that atrazine-containing soil has a stress effect on the growth of wheat and corn crops, and that Ureaplasma urealyticum ADBD-13 can effectively repair the stress effect of atrazine on wheat and corn seedlings. Attached Figure Description

[0022] Figure 1 This is a phylogenetic tree of Arthrobacter ureaplasm ADBD-13 constructed based on the 16S rRNA sequence.

[0023] Figure 2 The growth curve and degradation ability of Ureaplasma urealyticum ADBD-13 in inorganic salt liquid culture medium are shown.

[0024] Figure 3 The atrazine degradation rate and turbidity of *Arthrobacter ureagenesis* ADBD-13 at different initial pH values ​​are shown.

[0025] Figure 4The atrazine degradation rate and turbidity of *Arthrobacter ureagenesis* ADBD-13 at different shaking incubation temperatures are shown.

[0026] Figure 5 The atrazine degradation rate and turbidity of *Ureaplasma urealyticum* ADBD-13 at different initial atrazine concentrations are shown.

[0027] Figure 6 This is a graph showing the change in atrazine residue in soil over time after treatment with Ureaplasma urealyticum ADBD-13. Implementation

[0028] The specific embodiments of the present invention will be further described in detail below with reference to the accompanying drawings and examples. The following examples are only used to more clearly illustrate the technical solutions of the present invention, so that those skilled in the art can fully understand and utilize the present invention.

[0029] However, the present invention can be implemented in many other ways different from those described in the following embodiments, and those skilled in the art can make similar improvements without departing from the spirit of the present invention. Therefore, the scope of protection of the present invention is not limited to the specific embodiments disclosed below.

[0030] Unless otherwise defined, all technical and scientific terms used in this invention have the same meaning as commonly understood by those skilled in the art. The terminology used in this specification is merely for describing particular embodiments and is not intended to limit the invention.

[0031] The term "and / or" as used in this invention includes any and all combinations of one or more of the associated listed items.

[0032] The terms “multiple,” “various,” “multiple times,” “multiple groups,” etc., used in this invention, unless otherwise specified, refer to a quantity greater than or equal to 2; “more than” includes the number itself, such as “two or more” including two, three or more kinds.

[0033] The term "preferred" as used in this invention is only used to describe a more effective implementation or embodiment, and does not constitute a limitation on the scope of protection of this invention.

[0034] Unless otherwise specified, the production processes, experimental methods, or testing methods involved in the embodiments of this invention are all conventional methods in the prior art, and their names and / or abbreviations are all conventional names in the field, which are very clear and distinct in the relevant application areas. Those skilled in the art can understand the conventional process steps based on the names and apply the corresponding equipment, and implement them according to conventional conditions or conditions recommended by the manufacturer, or by referring to experimental methods known in the field.

[0035] The various instruments, equipment, raw materials or reagents used in the embodiments of this invention are not subject to any special restrictions on their source. They are all conventional products that can be purchased through regular commercial channels and can be prepared according to conventional methods known to those skilled in the art.

[0036] The strain *Arthrobacter ureagenes* ADBD-13 used in this embodiment of the invention was isolated from soil samples from a cornfield in Pingyuan County, Dezhou City, Shandong Province.

[0037] Specifically, 10g of soil sample was added to 100mL of inorganic salt medium containing 10mg / L atrazine, and cultured at 30℃ with shaking at 180r / min. Every 7 days, the culture was transferred at a rate of 10% to inorganic salt medium with atrazine concentrations progressively increased to 20mg / L, 30mg / L, 50mg / L, and 100mg / L. Finally, the enriched culture at an atrazine concentration of 100mg / L was serially diluted to 10^10 concentrations. -1 10 -2 10 -3 10 -4 10 -5 For each concentration of atrazine, take 100 μL and spread it onto a solid inorganic salt culture medium plate containing 100 mg / L atrazine. Place the plates in a 30°C incubator for incubation.

[0038] After culturing for 3 to 5 days, colonies with hydrolysis zones were picked, streaked to isolate and purify them, and five strains that can effectively degrade atrazine, namely ADBG-2, ADBG-3, ADBG-5, ADBD-13 and ADBD-14, were screened out.

[0039] After 35 days of acclimatization, the five degrading bacteria selected from the cornfield all grew transparent hydrolysis zones on inorganic salt solid culture medium containing 100 mg / L atrazine. The diameters of the transparent hydrolysis zones were 5.0 mm, 4.0 mm, 3.0 mm, 6.0 mm, and 5.5 mm, respectively. Among them, ADBD-13 had the largest hydrolysis zone diameter and the best degradation effect. Therefore, it was selected as an atrazine degrading strain, and its degradation characteristics were determined.

[0040] Atrazine solution: Weigh 1g of atrazine technical grade, add 90mL of methanol to dissolve, and make up to 100mL to obtain an atrazine stock solution with a final concentration of 10000mg / L. Dilute to the required concentration according to the ratio.

[0041] The preparation methods for various culture media used in the embodiments of the present invention are as follows:

[0042] Inorganic salt (minimal medium, MM) liquid culture medium: KH2PO4 0.45g, K2HPO4 1.79g, NaCl 0.4g, MgSO4·7H2O 0.2g, distilled water to a final volume of 1000mL, pH adjusted to 7.0.

[0043] LB (Luria-Bertani) liquid medium: 10g tryptone, 5g yeast extract, 10g NaCl, distilled water to a final volume of 1000mL, pH adjusted to 7.0.

[0044] Inorganic salt and LB solid medium are obtained by adding 2% agar to the liquid medium.

[0045] Unless otherwise specified, the amounts of raw material components and measurement parameters such as temperature and time involved in the embodiments of the present invention may have slight deviations within the range of weighing or measurement accuracy, and acceptable deviations caused by instrument testing accuracy or operational accuracy are allowed. Example

[0046] Example 1

[0047] Soil samples were collected from cornfields in Pingyuan County, Dezhou City, Shandong Province. 10g of soil sample was added to 100mL of inorganic salt liquid culture medium containing 10mg / L atrazine. After thorough mixing, the mixture was cultured at 30℃ and 180r / min with shaking.

[0048] Every 7 days, the culture was transferred at a rate of 10% to an inorganic salt liquid medium with increasing atrazine concentrations of 10 mg / L, 20 mg / L, 30 mg / L, 50 mg / L, and 100 mg / L.

[0049] Take the enriched culture with an atrazine concentration of 100 mg / L and serially dilute it to 10⁻⁶. -1 10 -2 10 -3 10 -4 10 - 5 Five concentrations of atrazine were used, with 100 μL of each concentration spread onto inorganic salt solid culture plates containing 100 mg / L atrazine. The plates were placed in a 30°C incubator and incubated for 3–5 days. Colonies with hydrolysis zones were picked, streaked to isolate and purify the bacteria, and strains that could effectively degrade atrazine were screened. The strain with the best degradation effect, ADBD-13, was selected.

[0050] Example 2

[0051] The isolated and purified strain ADBD-13 was streaked onto LB agar. Single colonies were picked; the colonies were viscous and moist, round, opaque, and pale yellow in color, with smooth and intact edges. Microscopic observation revealed that the colonies were rod-shaped and without spores.

[0052] The selected strain ADBD-13 was molecularly identified, and its genome was sequenced by Sangon Biotech (Shanghai) Co., Ltd. using next-generation sequencing technology.

[0053] Based on the genome sequencing results, RNAmmer was used to predict the 16S rRNA sequence of the bacterial species. The predicted results were compared with the NCBI nucleotide database using the BLAST program, and a phylogenetic tree was constructed using MEGA 11.0. The average nucleotide similarity (ANI) index between the tested strain and 20 reference strains was calculated using FastANI and a heatmap was generated. The species of the strain was determined by combining the predicted 16S rRNA sequence and the ANI index.

[0054] A comparison with the NCBI nucleotide database showed that strain ADBD-13 and Paenarthrobacter ureafaciens strain NC is in a unified branch, with 99.53% similarity. Based on the heatmap results of strain ADBD-13 and 20 reference strains, strain ADBD-13 and... Paenarthrobacter ureafaciens The ANI index of AAC22 (GCA-011393315.2) is as high as 98.09%, exceeding 95%.

[0055] Based on the 16S rRNA phylogenetic tree of strain ADBD-13 and the ANI results with the reference strain, strain ADBD-13 was finally identified as *Arthrobacter ureaplasmosis* (Ureaplasma urealyticum). Paenarthrobacter ureafaciens A phylogenetic tree of 16S rRNA was constructed using the neighbor-joining method for strain ADBD-13, as shown below. Figure 1 As shown.

[0056] Example 3

[0057] The growth curve and degradation ability of the screened strain ADBD-13 were determined.

[0058] Single colonies of strain ADBD-13 were inoculated into LB liquid medium and incubated with shaking for 16–24 h. After centrifugation at 4500 rpm for 10 min at 4 °C, the bacterial sludge was collected and repeatedly washed three times with inorganic salt liquid medium. Inorganic salt liquid medium was then added to adjust the colony concentration to 1 × 10⁻⁶. 9 The concentration of cfu / mL was used to obtain ADBD-13 bacterial culture.

[0059] The ADBD-13 bacterial suspension was added to 50 mL of inorganic salt liquid medium containing 100 mg / L atrazine to achieve a concentration of 2%. An uninoculated inorganic salt liquid medium containing atrazine served as a negative control. Samples were taken at 1.5, 3, 6, 9, 12, 15, 17.5, 20, 30, 50, 70, 90, 110, 130, and 150 h, and the OD values ​​of the bacterial suspension were measured. 600 The values ​​and atrazine concentrations in the culture medium were determined, and the experiment was repeated three times.

[0060] OD 600 The values ​​were determined using a micro spectrophotometer.

[0061] Atrazine concentration was determined by high performance liquid chromatography (HPLC). Specifically, atrazine was extracted from the bacterial solution with dichloromethane, evaporated to dryness, diluted to volume with chromatographic grade methanol, filtered through a 0.22 µm filter membrane, and the atrazine content was determined by external standard method using HPLC.

[0062] High performance liquid chromatography (HPLC) conditions: Hypersil ODS2 column (4.6 mm × 250 mm, 5 µm); column temperature 30 °C; mobile phase methanol:water (0.1% formic acid) = 70:30 (V / V); wavelength 225 nm; flow rate 1 mL / min; injection volume 20 µL.

[0063] Based on the measurement results, the degradation rate of atrazine was calculated using the formula: Degradation rate (%) = (Initial concentration of atrazine - Final concentration of atrazine) / Initial concentration of atrazine × 100%.

[0064] Figure 2 In the study, strain ADBD-13, using atrazine as its sole energy source, exhibited good growth in an inorganic salt liquid medium containing 100 mg / L atrazine. Initially, it experienced a lag phase of approximately 10 hours, during which the turbidity of the bacterial solution remained almost unchanged, and the atrazine concentration decreased slowly. From 10 to 30 hours, the turbidity of the bacterial solution increased rapidly, indicating that strain ADBD-13 was in the logarithmic growth phase. From 30 to 130 hours, strain ADBD-13 was in the stationary phase. After 130 hours, strain ADBD-13 entered the decline phase.

[0065] According to Figure 2 The atrazine concentration curves show that the atrazine concentration decreases rapidly during the logarithmic growth phase of strain ADBD-13, indicating that strain ADBD-13 is most sensitive to the effects at this time. Therefore, 20 hours was chosen as the study time for degradation characteristics. Furthermore, the atrazine degradation rate of strain ADBD-13 reached its highest level of 99.90% at 70 hours.

[0066] Obviously, compared with the atrazine degradation rate of 88.90% by Pseudomonas SFAD3 and 90% by Agrobacterium AT-2 in the background art, the strain ADBD-13 of the present invention can degrade atrazine more efficiently.

[0067] Example 4

[0068] The degradation characteristics of strain ADBD-13 were analyzed from three aspects: initial pH value, shaking incubation temperature, and initial concentration of atrazine.

[0069] The initial pH values ​​were set to 5.0, 6.0, 7.0, 8.0 and 9.0, the shaking incubation temperatures were 10℃, 20℃, 30℃, 40℃ and 50℃, and the initial atrazine concentrations were 10mg / L, 25mg / L, 50mg / L, 100mg / L and 200mg / L.

[0070] Add 2% ADBD-13 bacterial solution (1×10⁻⁶) 9 Add (cfu / mL) to an atrazine-containing inorganic salt liquid culture medium adjusted to the test conditions, and incubate at 150 rpm for 20 h. Then, detect the atrazine content.

[0071] According to the formula [( C 0+ C x ) / C The degradation rate of strain ADBD-13 was calculated as 0% × 100%. Where: C 0 represents the final concentration of atrazine in the control sample; C x This represents the final concentration of atrazine in the bacterial strain sample.

[0072] Specific results are as follows Figure 3 , Figure 4 , Figure 5 As shown, the initial pH value, shaking temperature, and initial concentration of atrazine all affected the degradation effect of strain ADBD-13.

[0073] Although strain ADBD-13 achieved the highest degradation rate of atrazine (89±0.4%) at an initial pH of 8.0, it was also effective in degrading atrazine at pH values ​​ranging from 5 to 9, with degradation rates exceeding 75% in all these pH ranges. Figure 3 This indicates that strain ADBD-13 has strong tolerance to acidic and alkaline environments.

[0074] Incubation temperature was the most influential factor on the degradation characteristics of strain ADBD-13. 30℃ was clearly the optimal incubation temperature for strain ADBD-13, at which the degradation rate of atrazine reached 80±0.5%. Figure 4 ).

[0075] Figure 5 This indicates that as the initial concentration of atrazine increases, the degradation rate of atrazine by strain ADBD-13 gradually decreases. This may be because the increased concentration of atrazine increases its cytotoxicity to the degrading bacteria, leading to a decrease in the degradation capacity of the bacteria.

[0076] Example 5

[0077] Single colonies of strain ADBD-13 were inoculated into 250 mL of LB liquid medium and fermented at 30°C and 200 rpm for 20 h. The precipitate was collected, washed three times repeatedly with inorganic salt medium, and then inorganic salt medium was added again to adjust the colony concentration in the solution to 1 × 10⁻⁶. 9 ADBD-13 bacterial suspension was prepared by using cfu / mL.

[0078] Example 6

[0079] 100g of soil was weighed, sieved and sterilized, and then atrazine was added until the atrazine content in the soil was 50mg / kg. The remediation effect of the ADBD-13 bacterial suspension prepared in Example 5 on soil containing atrazine residue was investigated.

[0080] The treated soil was divided into two groups. One group was treated with ADBD-13 bacterial suspension to achieve a concentration of 10 in the soil. 6 CFU / g, and another group was treated with sterile distilled water. Both groups of soil were placed in sterile conical flasks, and water was sprayed daily to maintain soil moisture.

[0081] Samples were taken on days 1, 2, 3, 4, 5, 6, and 7 to detect atrazine residues in the soil. The samples were added to 50 mL of methanol, sonicated for 40 min, centrifuged at 12000 rpm for 15 min, and the supernatant was collected, evaporated to dryness at 35℃, diluted to volume with chromatographic grade methanol, filtered through a 0.22 µm filter, and analyzed. Each experiment was repeated three times.

[0082] according to Figure 6 The curve showing the change of atrazine residue over time indicates that sterile distilled water could only reduce the atrazine concentration in the soil from 50 mg / kg to 49.12 mg / kg after 7 days, with a degradation rate of only 1.76%. However, after treatment with ADBD-13 bacterial suspension, the atrazine concentration in the soil decreased to 14.73 mg / kg after 7 days, with a degradation rate of 70.54%. This demonstrates that ADBD-13 bacterial suspension has a significant biodegradation effect on atrazine and can effectively remediate atrazine-contaminated soil.

[0083] Example 7

[0084] This embodiment verifies the restorative effect of ADBD-13 bacterial suspension on wheat and corn seedlings under atrazine stress in soil.

[0085] Weigh 250g of soil into a flowerpot, add distilled water to moisten the soil and mix thoroughly. The soil is then randomly divided into four groups: blank group, control group, stress group and treatment group.

[0086] In this study, no atrazine was added to the blank group and the control group, while 2.5 µL of atrazine at a concentration of 10 g / L was added to the stress group and the treatment group, so that the atrazine concentration in the soil was 1 mg / kg; and 2.5 mL of ADBD-13 bacterial suspension was added to the control group and the treatment group.

[0087] Select plump, similarly sized germinated wheat and corn seeds, and plant 6 seeds per pot in soils with different treatments. Grow for 14 days at 27°C, with light and dark cycles of 16h and 8h respectively, relative humidity of 70%, and light intensity of 5500 lux.

[0088] Each treatment was repeated 5 times. In each repetition, 5 plants were randomly selected. The plant height, root length, fresh weight and dry weight of wheat and corn were recorded. The results are listed in Tables 1 and 2.

[0089]

[0090]

[0091] As shown in Tables 1 and 2, in the pot experiment of atrazine stress, the plant height, root length, fresh weight, and dry weight of wheat and maize in the stress group were reduced by at least 32% compared with the control group. The control group, which received only the bacterial suspension, had little effect on the four growth indicators of wheat and maize seedlings, showing virtually no difference from the control group. However, when the bacterial suspension was mixed with atrazine in the treatment group, the four growth indicators of wheat and maize seedlings increased by at least 21% and 29%, respectively, compared with those under stress. This indicates that the use of ADBD-13 bacterial suspension can effectively remediate the stress caused by atrazine in the soil on wheat and maize seedlings.

[0092] Example 8

[0093] This embodiment verifies the restorative effect of ADBD-13 bacterial suspension on atrazine stress in wheat and maize seedlings in inorganic salt liquid culture medium.

[0094] Select plump, similarly sized germinated wheat and corn seeds and plant them separately on foam boards in a space cup. Immerse the roots in different treatment solutions and grow them for 14 days at 27℃, with light and dark cycles of 16h and 8h respectively, relative humidity of 70%, and light intensity of 5500 lux. Record the growth indicators of the whole wheat and corn plants, such as plant height, root length, fresh weight, and dry weight.

[0095] The specific compositions of the different treatment solutions are as follows:

[0096] A: Blank group, inorganic salt liquid culture medium; B: Control group, inorganic salt liquid culture medium with 2% ADBD-13 bacterial suspension added; C: Stress group, inorganic salt liquid culture medium containing 100 mg / L atrazine; D: Treatment group, inorganic salt liquid culture medium containing 100 mg / L atrazine and 2% ADBD-13 bacterial suspension added.

[0097] All treatment solutions were incubated at 30°C and 150 rpm for 5 days and then diluted 10 times. Sterile distilled water was added during the experiment to maintain moisture.

[0098] Each group was treated with 6 wheat and 6 corn seeds, and the treatment was repeated 5 times. In each repetition, 5 plants were randomly selected to record data.

[0099]

[0100]

[0101] As shown in Tables 3 and 4, the growth indicators of wheat and corn seedlings grown in different treatment solutions for 14 days showed significant changes. The four growth indicators of wheat and corn seedlings treated with ADBD-13 bacterial suspension and atrazine were all lower than those of the control group to some extent. However, the atrazine treatment reduced the four growth indicators of wheat seedlings by 20%, 46%, 43%, and 61%, respectively, and the four growth indicators of corn seedlings by 43%, 61%, 58%, and 46%, respectively. In contrast, the ADBD-13 bacterial suspension treatment only reduced the fresh weight of wheat seedlings by 13%, while the other growth indicators of wheat and corn seedlings remained almost unchanged.

[0102] However, in the treatment groups simultaneously treated with ADBD-13 bacterial suspension and atrazine, compared to the control group treated with atrazine alone, all four growth indicators of wheat and corn seedlings showed significant improvement. Specifically, the four growth indicators for wheat seedlings increased by 24%, 57%, 70%, and 91%, respectively, while those for corn seedlings increased by 27%, 105%, 32%, and 34%, respectively. In particular, the fresh weight reached the level of the control group, and the plant height exceeded that of the control group, indicating a certain growth-promoting effect. This suggests that ADBD-13 bacterial suspension can significantly repair the stress caused by atrazine residues in water on wheat and corn seedlings, and the repair effect is better than that in soil. This is because during hydroponic repair, most of the atrazine has been converted into cyanuric acid, which poses a less threatening threat to wheat and seedlings.

[0103] The above embodiments illustrate several specific and detailed implementations of the present invention, but should not be construed as limiting the scope of protection of the present invention. It should be noted that those skilled in the art can make various substitutions, modifications, or improvements without departing from the principles and spirit of the present invention, all of which should be included within the scope of protection of the present invention.

Claims

1. A strain of atrazine-degrading Arthrobacter ureagenesis ADBD-13 was deposited at the China Center for Type Culture Collection on December 27, 2024, with accession number CCTCC NO: M20242925.

2. A suspension of *Arthrobacter ureagenesis* that degrades atrazine, wherein the suspension contains *Arthrobacter ureagenesis* ADBD-13 as described in claim 1.

3. The *Arthrobacter urealyticum* suspension according to claim 2, characterized in that... The bacterial suspension was prepared using an inorganic salt culture medium at a concentration of 1×10⁻⁶. 9 CFU / mL bacterial suspension.

4. The *Arthrobacter ureagenesis* suspension according to claim 3, characterized in that... Single colonies of *Arthrobacter ureaplasma* ADBD-13 were inoculated into LB liquid medium and cultured at 30°C with shaking at 200 rpm for 16–20 h. The bacterial culture was then centrifuged, the precipitate was collected, washed with inorganic salt medium, and then prepared into a 1×10⁻⁶ concentration using inorganic salt medium. 9 CFU / mL bacterial suspension.

5. The *Arthrobacter urealyticum* suspension according to claim 3 or 4, characterized in that... The inorganic salt culture medium consists of: 0.45g KH2PO4, 1.79g K2HPO4, 0.4g NaCl, 0.2g MgSO4·7H2O, diluted with distilled water to 1000mL, pH 7.

0.

6. The use of the *Arthrobacter urealyticum* ADBD-13 as described in claim 1 in the degradation of atrazine.

7. The application of the Ureaplasma urealyticum ADBD-13 of claim 1 in the remediation of soil containing atrazine residues.

8. The application of the *Arthrobacter urealyticum* ADBD-13 of claim 1 in promoting the growth of atrazine-stressed crops, wherein the crop is maize or wheat.

9. The use of the *Arthrobacter urealyticum* suspension according to claim 2 in the preparation of atrazine degrading agents.

10. The use of the *Arthrobacter urealyticum* suspension according to claim 2 in the preparation of atrazine-containing soil remediation agents.

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