A salt-alkali tolerant Streptomyces caulkinii strain and its application

Abstract

The present invention provides a salt- and alkali-tolerant strain of Streptomyces cavourensis and its applications. The strain, named Streptomyces cavourensis PY4, was deposited with the China Center for Type Culture Collection at Wuhan University in Wuhan, China on November 21, 2024, with a deposit number of CCTCC NO: M 20242608. This strain can still effectively degrade the herbicide acetochlor in a highly saline and alkali environment. After 15 days of application, it can reduce the residual acetochlor in the soil by 91.50%. It can be used to solve the problem of excessive acetochlor residues in soil and other environments, and has broad application prospects in the field of agricultural production.

Description

Technical Field

[0001] The invention relates to the field of pesticide degradation microorganisms and agriculture, and relates to a salt- and alkali-resistant Streptomyces caulkinii and an application thereof. Background Art

[0002] Any discussion of the prior art throughout the specification should not be considered as an admission that such prior art is widely known or forms part of the common general knowledge in the field.

[0003] Acetochlor is one of the three largest herbicide classes in my country in terms of production and usage volume. It is a broad-spectrum, highly effective amide pre-emergence herbicide primarily used to control grass weeds in wheat, corn, soybean, and peanut fields. Acetochlor has a short half-life in soil, generally eliminating the need for remediation. However, excessive and inappropriate application can easily maintain high levels, making it difficult to degrade in a timely manner. Acetochlor is highly permeable and can penetrate deep soil and groundwater, causing toxicity to humans and animals. It has been designated a Class B-2 carcinogen by the US Environmental Protection Agency. Therefore, degrading acetochlor residues in farmland is of great significance for agricultural production and food safety. Pesticide contamination remediation generally involves physical, chemical, and biological methods. Microbial remediation has gained attention for its effectiveness, short cycle times, ease of operation, and lack of secondary pollution. However, few reports exist on microbial remediation technologies for acetochlor that are adaptable to complex soil environments.

[0004] Patent CN 105062917 B discloses a strain of Rhodococcus sp. AC-1, which, when applied directly to soil, achieved a 90.1% degradation rate for acetochlor within seven days. Patent CN 116254188 B discloses a strain of Cupidesulfovibrio sp. SRB-6, which achieved a 97% degradation rate for acetochlor in paddy mud within 14 days under anaerobic conditions. The strains provided in these patents have high acetochlor degradation efficiencies under both normal and anaerobic conditions. However, no strains have been found to exhibit significant degradation effects in highly saline and alkaline soil environments. Summary of the Invention

[0005] The present invention aims to provide a salt- and alkali-tolerant acetochlor-degrading bacterial strain and its application. Specifically, it comprises:

[0006] First, a strain of Streptomyces caulkinii that can efficiently degrade acetochlor and its fermentation broth are provided. In particular, the strain maintains good degradation activity even in highly saline and alkaline environments, effectively resolving the existing problem of a lack of acetochlor-degrading bacteria that are adaptable to complex environments.

[0007] Secondly, a series of products containing the above-mentioned strains or their fermentation broths are provided, including suspensions, liquid fermentation preparations, powders, granules and other forms to meet the needs of different application scenarios.

[0008] A third aspect provides a method for degrading acetochlor using the strain or its fermentation liquid or product. This method helps degrade residual acetochlor in the environment. The method is applicable to, but not limited to, soil, water, sediments, or other contaminated areas, and has particularly good application results in saline-alkali soils.

[0009] Fourthly, the invention further provides the application of the strain in the degradation of acetochlor. The characteristics of the strain of the present invention expand the application range of the strain, making it suitable not only for ordinary farmland soil, but also for the degradation of acetochlor in saline-alkali soil, water bodies, sediments or other polluted areas.

[0010] The present invention addresses the existing technical problem of a lack of salt- and alkali-tolerant acetochlor-degrading bacteria, providing a novel solution for the bioremediation of residual acetochlor in saline-alkali farmland, water bodies, and other environments. This solution offers advantages such as high degradation efficiency, strong adaptability, and a wide range of applications. It significantly improves the degradation of residual acetochlor and has significant application value in agricultural production and environmental protection.

[0011] Specifically, the present invention provides the following technical solutions.

[0012] In the first aspect of the present invention, a salt- and alkali-tolerant Streptomyces cavourensis PY4 is provided, which was deposited in the China Center for Type Culture Collection at Wuhan University, Wuhan, China on November 21, 2024, with a deposit number of CCTCC NO: M 20242608.

[0013] The strain has the following characteristics:

[0014] 1) Morphological Characteristics: A single colony of strain PY4 was streaked onto a NA plate, inverted in a constant temperature incubator, and incubated at 37°C for 48 hours. The colony was round, with neat edges, small and dense, dry and opaque, with a powdery white surface and easy to pick up. Microscopic observation revealed that the mycelium of strain PY4 was slender and wavy, with small, nearly spherical conidia produced at different locations on the hyphae.

[0015] 2) 16S rDNA gene sequence: as shown in SEQ ID NO: 1.

[0016] The strain exhibits excellent salt and alkali resistance and degradation capabilities:

[0017] 1) pH tolerance: The plant can grow normally in the pH range of 6-9 and has good ability to degrade acetochlor within this pH range, with a degradation rate exceeding 71% (3 days). The highest degradation rate is at pH 7, reaching 78.53%. Even in the high alkaline environment of pH 9, the degradation rate is still maintained at 73.26%.

[0018] 2) Salt tolerance: The plant can grow normally in a sodium chloride concentration range of 5-100 g / L and exhibits good acetochlor degradation activity under these conditions, with a degradation rate exceeding 73% (3 days). The highest degradation rate, reaching 85.42%, was achieved at 5 g / L. Even in a high-salt environment of 100 g / L, the degradation rate still reached 73.59%.

[0019] 3) Temperature adaptability: It can grow in the range of 20-37°C, has good activity in the range of 25-37°C, and has the highest degradation rate at 30°C, reaching 78.43% (3 days).

[0020] In a second aspect of the present invention, a fermentation broth of the salt- and alkali-tolerant Streptomyces calwinii described in the first aspect is provided, which is obtained by culturing the Streptomyces calwinii in a culture medium.

[0021] Among them, the culture medium can be selected from a culture medium containing a carbon source and a nitrogen source, such as: NB medium, ISP medium, LB medium, Gao's medium No. 1, etc. For example, in one embodiment, the culture medium composition includes 10 g / L peptone, 3 g / L beef extract powder, 5 g / L sodium chloride, and pH = 7.2.

[0022] In a third aspect of the present invention, a product is provided, which comprises the fermentation broth of the salt- and alkali-tolerant Streptomyces calwinii described in the first aspect or the salt- and alkali-tolerant Streptomyces calwinii described in the second aspect.

[0023] The product is a microbial agent and can be used in any of the following dosage forms: 1) suspension, 2) liquid fermentation preparation, 3) powder and 4) granule.

[0024] In the fourth aspect of the present invention, there is provided the use of the salt- and alkali-tolerant Streptomyces calviphyllum described in the first aspect or the fermentation broth of the salt- and alkali-tolerant Streptomyces calviphyllum described in the second aspect or the product described in the third aspect in the degradation of acetochlor.

[0025] The application environment includes but is not limited to soil, water, sediment or other polluted areas. The soil includes ordinary soil and saline-alkali soil.

[0026] In the fifth aspect of the present invention, a method for degrading acetochlor is provided, the method comprising: applying the fermentation broth of the salt- and alkali-tolerant Streptomyces calvus described in the first aspect or the salt- and alkali-tolerant Streptomyces calvus described in the second aspect or the product described in the third aspect to a material to be treated that is contaminated with acetochlor.

[0027] The term "substance to be treated" as used herein refers to any object contaminated with acetochlor and requiring treatment using the method of the present invention, including but not limited to soil, water, sediment, contaminated samples, or other contaminated carriers. Specifically, it refers to soil and water. The soil includes both ordinary soil and saline-alkali soil.

[0028] In some embodiments of the present invention, the initial concentration of viable bacteria of the salt-alkali tolerant Streptomyces caulkinii when applied to the object to be treated, especially soil or water, is 1×10 6 -1×10 8 CFU / mL; the application amount is 0.2-2% of the mass of soil or water.

[0029] In some embodiments of the present invention, the pH of the object to be treated, especially soil or water, is 6-9, and the water-soluble total salt content thereof is 0.1-20‰, preferably 0.1-10‰, and more preferably 0.1-8‰.

[0030] In some embodiments of the present invention, before applying to the object to be treated, especially soil, the method further includes adjusting the moisture in the object to be treated, especially soil, so that the moisture content of the object to be treated, especially soil, is adjusted to 60-80% (w / w) of its maximum water holding capacity.

[0031] In one embodiment of the present invention, the method comprises the following steps:

[0032] 1) When the treatment object is soil: before application, adjust the soil moisture content to 60-80% (w / w) of its maximum water holding capacity;

[0033] Applying the strain, the fermentation liquid of the strain or the product containing the strain to the soil; and maintaining appropriate soil moisture.

[0034] 2) When the treatment object is a water body: directly apply the strain, the fermentation liquid of the strain, or the product containing the strain to the water body.

[0035] In the above embodiment, preferred application conditions are:

[0036] 1) Strain activity: The concentration of viable bacteria at the time of application is 1×10 6 -1×10 8 CFU / mL.

[0037] 2) Application amount: 0.2-2% of the amount of the substance to be treated.

[0038] 3) Environmental conditions: pH 6-9, water-soluble total salt content 0.1-20‰, preferably 0.1-10‰, more preferably 0.1-8‰.

[0039] The water-soluble total salts mentioned in the present invention refer to inorganic salts that can dissolve in water, including chlorides, sulfates, carbonates, bicarbonates, nitrates, etc. For example: sodium chloride, sodium sulfate, calcium carbonate, etc.

[0040] The water-soluble total salt content of the present invention refers to the percentage or concentration of the total mass of all water-soluble salts in a sample (usually soil or water, etc.) to the dry mass of the sample.

[0041] Compared with the prior art, the advantages of the present invention include:

[0042] The Streptomyces caulkinii strain provided by the present invention has excellent salt-alkali tolerance and can maintain activity within a pH range of 6-9. Even under highly alkaline conditions of pH 9 and a high-salt environment with a sodium chloride concentration of 100 g / L, the strain can achieve a degradation rate of 75.16% for acetochlor within 3 days. This solves the problem of the prior art lacking degrading bacteria adapted to high-salinity and alkaline environments.

[0043] The degradation of acetochlor by Streptomyces kaulii in saline-alkali soil conforms to the first-order kinetic equation (y=10.74e (-0.1322t) ), its degradation half-life was only 5.24 days, 9.75 days shorter than that of the control group. This high degradation capacity significantly improved the treatment efficiency of residual acetochlor.

[0044] The strain of the present invention has a wide range of environmental adaptability. In addition to tolerating high pH and high salt, it can also maintain good degradation activity within the temperature range of 25-37°C. This excellent environmental adaptability enables it to function stably in complex practical application environments.

[0045] The degradation method provided by the present invention is simple to operate and the conditions of use are easy to control. By adjusting the moisture content of the material to be treated, such as soil, to 60-80% (w / w) of the maximum water holding capacity and applying the active bacteria at a dosage of 0.2%-2% with a concentration of 1×10 6 -1×10 8 CFU / mL of bacterial solution can achieve effective degradation of the treated object such as acetochlor in soil.

[0046] The technical solution of the present invention has a wide range of applications. This method is applicable not only to soil environments but also to the remediation of aquatic environments. In particular, the present invention provides an efficient and feasible solution for the remediation of saline-alkali soils.

[0047] The invention can be developed into various dosage forms, including suspensions, liquid fermentation preparations, powders and granules, etc., which are convenient for storage, transportation and practical application, and have good industrialization prospects.

[0048] In summary, the present invention not only provides a salt- and alkali-tolerant acetochlor-degrading bacterium with excellent performance, but also establishes a complete application technology system, providing a new technical solution for the bioremediation of acetochlor pollution in saline-alkali environments, which has important application value and promotion significance. BRIEF DESCRIPTION OF THE DRAWINGS

[0049] The drawings constituting part of the present application are provided to provide a further understanding of the present application. The illustrative embodiments and their descriptions of the present application are provided to explain the present application and do not constitute an undue limitation on the present application. The following describes the implementation scheme of the present application in detail in conjunction with the drawings, wherein:

[0050] Figure 1 The figure shows the peak pattern of acetochlor in high performance liquid chromatography (HPLC).

[0051] Figure 2 An acetochlor standard curve is shown.

[0052] Figure 3 The colony morphology of strain PY4 on NA plates is shown.

[0053] Figure 4 The bacterial morphology of strain PY4 under an optical microscope is shown.

[0054] Figure 5 The figure shows the effect of strain PY4 on the degradation kinetics of acetochlor in saline-alkali soil.

[0055] Figure 6 The graph shows the degradation rates of acetochlor in each saline-alkali soil treatment group at different times in Example 6.

[0056] Figure 7 The graph shows the degradation rates of acetochlor in each treatment group of normal soil in Example 6 at different times. DETAILED DESCRIPTION

[0057] The present application will be further described in conjunction with specific examples. It should be understood that these examples are intended to illustrate the present application only and are not intended to limit the scope thereof. Experimental methods for which specific conditions are not specified in the examples are generally performed under conventional conditions or conditions recommended by the manufacturer.

[0058] Unless otherwise defined, all technical terms and scientific terms used in this application should have the meaning familiar to those skilled in the art. Unless otherwise specified, the reagents or raw materials used in this application can be obtained by conventional means and used in accordance with conventional methods or product specifications in this area. In addition, any content similar to or equivalent to the methods or materials described can be applied to the methods of this application. The preferred embodiments and materials described in this application are for illustrative purposes only.

[0059] Example 1 Screening of strains

[0060] 1. Test materials:

[0061] (1) The emulsion acetochlor was obtained from Nantong Jiangshan Pesticide Chemical Co., Ltd. under the trade name of Heness. The acetochlor content was 90%, and the emulsion was dissolved in acetone to a stock concentration of 50 g / L.

[0062] (2) Soil sample collection: A total of more than 60 soil samples were collected for the experiment, which were taken from the sludge at the sewage outlets of Huayang Pesticide Factory, Baofeng Pesticide Factory and Zhongnong United Pesticide Factory in Tai'an City, Shandong Province.

[0063] (3) Test culture medium

[0064] MSM liquid culture medium: ammonium sulfate 1.0 g / L, potassium dihydrogen phosphate 0.5 g / L, dipotassium hydrogen phosphate 1.5 g / L, magnesium sulfate heptahydrate 0.1 g / L, sodium chloride 1.0 g / L.

[0065] MSM solid medium: Add 20g / L agar to the MSM liquid medium.

[0066] NB medium: peptone 10 g / L, beef extract powder 3 g / L, sodium chloride 5 g / L, pH = 7.2.

[0067] NA medium: Add 20 g / L agar to NB medium.

[0068] 2. Test methods:

[0069] (1) Enrichment culture of strains

[0070] 5g of soil was added to 50mL of MSM liquid culture medium with an acetochlor concentration of 50mg / L, a sodium chloride concentration of 30g / L, and a pH of 7, and placed in a shaking incubator at 37°C and 180r / min for enrichment culture. After 7d, 5mL of fermentation liquid was transferred to 50mL of MSM liquid culture medium with an acetochlor concentration of 100mg / L, a sodium chloride concentration of 50g / L, and a pH of 8 for enrichment culture. After 7d, 5mL of fermentation liquid was transferred to 50mL of MSM liquid culture medium with an acetochlor concentration of 200mg / L, a sodium chloride concentration of 100g / L, and a pH of 9. After further culture for 7d, 1mL of supernatant was taken and washed according to 10-1 , 10 -2 , 10 -3 , 10 -4 , 10 -5 The diluted solutions were spread on MSM solid culture medium plates containing 200 mg / L acetochlor, 100 g / L sodium chloride, and pH 9, and placed in an incubator at 37°C for inverted culture.

[0071] (2) Isolation and purification of pesticide-degrading bacteria: After a single colony grows on the enriched and acclimated dilution plate, use an inoculation loop to pick a single colony with fast growth and regular colonies, streak it on the corresponding MSM solid agar medium plate, culture it at 37°C, repeat the streak 2-3 times until a single colony with good growth grows, and store it on a slant using NA medium.

[0072] Example 2 Rescreening of strains

[0073] 1. Test method:

[0074] (1) Experimental treatment

[0075] The 11 strains selected in Example 1 were cultured in NB medium at 37°C and 180 rpm in shake flasks until the logarithmic growth phase. Then, a 2% inoculum was inoculated into MSM liquid medium containing 200 mg / L acetochlor, 100 g / L sodium chloride, and pH 9. The culture was shaken in the dark at 37°C and 180 rpm for 3 days, with a blank control. Three replicates were performed for each strain. The residual acetochlor content in the fermentation broth was quantified by HPLC at the initial and final stages of the experiment. The degradation rate was calculated according to the following formula:

[0076] Acetochlor degradation rate (%) = (C0-C0) / C0×100%.

[0077] Wherein, C0 is the initial acetochlor content in the fermentation broth, in mg / L; C is the acetochlor content in the fermentation broth at the end of the experiment, in mg / L.

[0078] (2) Extraction and detection method of acetochlor in fermentation broth

[0079] Extraction method: Take 2 mL of the above fermentation broth respectively into a 10 mL plastic centrifuge tube, add 2 g of sodium sulfate and 4 mL of acetonitrile for extraction, shake in a shaker at 180 rpm for half an hour, then ultrasonicate for 10 minutes, continue to centrifuge at 4000 rpm for 5 minutes, take the upper organic phase, pass it through a 0.22 μm filter membrane, and then determine it by high performance liquid chromatography (HPLC).

[0080] Liquid chromatography conditions

[0081] The mobile phase was acetonitrile:water (70:30, V / V), the chromatographic column was InertSustain AQ-C18 (4.6 mm × 250 mm, 5 μm), the UV detection wavelength was 205 nm, the flow rate was 0.8 mL / min, the injection volume was 20 μL, and the column temperature was 25°C.

[0082] The retention time of acetochlor is 9.531 min, and the peak is as follows Figure 1 shown.

[0083] (3) Standard curve of acetochlor

[0084] Prepare a 1000mg / L acetochlor mother solution with acetone, and then dilute the mother solution with acetonitrile to form a series of standard sample solutions (0mg / L, 50mg / L, 100mg / L, 200mg / L, 300mg / L). Then, analyze the solution on HPLC. Draw a standard curve with the concentration of acetochlor as the horizontal axis and the peak area of ​​the target compound as the vertical axis. The results are as follows: Figure 2 As can be seen from the figure, the concentration of acetochlor has a good correlation with the peak area, and its linear regression equation is y=141356x, and the correlation coefficient R 2 =0.9991.

[0085] 2 Test results

[0086] Table 1 Degradation rate of acetochlor by each strain (%, n=3)

[0087]

[0088] As shown in Table 1, among the 11 strains screened in Example 1, one strain had an acetochlor degradation rate exceeding 70% in a high saline-alkali environment. This strain was numbered PY4, and its acetochlor degradation rate was 75.16%.

[0089] Example 3 Identification of bacterial species

[0090] (1) Colony morphological characteristics

[0091] Streak a single colony of strain PY4 onto a NA medium plate, place the plate upside down in a constant temperature incubator at 37°C, and culture for 48 hours. The colony will be round, with neat edges, small and dense, dry and opaque, with a powdery and white surface, and easy to pick up (see Figure 3 ).

[0092] (2) Bacterial morphological characteristics

[0093] The strain PY4 was stained with crystal violet staining solution. Under an optical microscope, the mycelium was observed to be slender and wavy, and small, nearly spherical conidia were produced at different sites of the mycelium (see Figure 4 ).

[0094] (3) Molecular identification

[0095] The bacterial genome was extracted by referring to the operating procedures of the Bacterial Genome Extraction Kit (Solaibo). Using the genome as a template, the bacterial 16S rDNA universal primers 27F and 1492R were used to amplify the bacterial 16S rDNA sequence, which was then sent to Sangon Biotech (Shanghai) Co., Ltd. for sequencing. Comparison with BLAST in NCBI showed that the 16S rDNA sequence of strain PY4 was 99.86% similar to the 16S rDNA sequence of Streptomyces cavourensis 1AS2a, indicating that the strain is Streptomyces cavourensis. The strain was named Streptomyces cavourensis PY4 (hereinafter referred to as strain PY4) and was deposited in the China Center for Type Culture Collection at Wuhan University, Wuhan, China on November 21, 2024, with the deposit number CCTCC NO: M20242608.

[0096] The 16S rDNA nucleotide sequence of the strain is shown in SEQ ID NO: 1.

[0097] Example 4 Study on the degradation characteristics of acetochlor by strain PY4 under different environments

[0098] 1. Test method:

[0099] Different pH value tests: strain PY4 was cultured in NB medium to the logarithmic growth phase. Then, 2% of the inoculum was inoculated into MSM liquid medium at pH 6, 7, 8, and 9, respectively. The acetochlor concentration of the medium was 200 mg / L and the sodium chloride content was 100 g / L. The culture was shaken at 37°C and 180 rpm for 3 days. The degradation rate was then measured.

[0100] Salt content test: strain PY4 was cultured in NB medium to the logarithmic growth phase. Then, at a 2% inoculum size, it was inoculated into MSM liquid medium containing sodium chloride contents of 5, 20, 40, 60, 80, and 100 g / L, respectively. The pH of the medium was 9, and the acetochlor content was 200 mg / L. The culture was shaken at 37°C and 180 rpm for 3 days, and the degradation rate was measured.

[0101] Different temperature tests: strain PY4 was cultured in NB medium to the logarithmic growth phase, and then inoculated at a 2% inoculum into MSM liquid medium with a pH of 9, a sodium chloride content of 100 g / L, and an acetochlor content of 200 mg / L. The cells were placed in a shaker at 25, 30, 35, and 37°C, respectively, and cultured at 180 rpm for 3 days. The degradation rate was then measured.

[0102] 2. Test results:

[0103] Table 2 Effect of pH on the degradation rate of acetochlor (%, n=3)

[0104] pH 6 7 8 9 Degradation rate% 71.95±2.48 78.53±2.68 74.54±2.29 73.26±1.97

[0105] Table 3 Effect of salt concentration on the degradation rate of acetochlor (%, n=3)

[0106]

[0107] Table 4 Effect of temperature on the degradation rate of acetochlor (%, n=3)

[0108] Temperature 25 30 35 37 Degradation rate% 73.14±2.17 78.43±2.42 75.11±2.26 74.19±2.55

[0109] Table 2 shows that strain PY4's optimal pH for acetochlor degradation is 7, with a degradation rate of 78.53%. However, the degradation rate did not decrease significantly with increasing pH, reaching 74.54% and 73.26% at pH 8 and 9, respectively. Table 3 shows that within the salt concentration range of 5-100 g / L, the acetochlor degradation rate of strain PY4 decreased slightly with increasing salt concentration, but still reached 73.59% at a salt concentration of 100 g / L. Therefore, strain PY4 exhibits high salt tolerance during acetochlor degradation. Table 4 shows that strain PY4's optimal temperature for acetochlor degradation is 30°C, with a degradation rate of 78.43%. The degradation rate decreased slightly with increasing or decreasing temperature, but remained above 70%.

[0110] Example 5 Effects of strain PY4 on the degradation kinetics of acetochlor in saline-alkali soil

[0111] 1 Experimental design:

[0112] The saline-alkali soil for testing was collected from Wudi County, Binzhou City, with a pH of 8.02 and a water-soluble total salt content of 6.97‰. 200g of soil sample was weighed into a conical flask. Commercially available acetochlor was added to each sample to a concentration of 10mg / kg, and the mixture was thoroughly mixed. Sterile water was added to adjust the soil moisture content to approximately 70% of its maximum field water holding capacity. Strain PY4 was cultured in NB medium until the logarithmic growth phase. The treated group was inoculated with 10% of the PY4 seed solution. The same volume of NB medium was added as a control. The jars were sealed with sterile, breathable film, and incubated in a 25°C incubator. Each treatment was repeated three times, maintaining a constant moisture content. Samples were collected on days 0, 3, 6, 9, 12, 15, 18, and 21 after treatment, dried at 65°C, and finely ground through a 60-mesh sieve for subsequent acetochlor content determination.

[0113] Extraction and determination of acetochlor in soil: Weigh 10 g of soil sample and add it into a 50 mL centrifuge tube, add 10 g of sodium sulfate and 25 mL of acetonitrile for extraction, place it in a shaker at 180 r / min for 30 minutes, then ultrasonicate for 10 minutes, and then centrifuge it at 4000 r / min for 5 minutes. Take the supernatant and filter it through a 0.22 μm organic filter membrane. The residual amount of acetochlor in the soil can be determined by HPLC.

[0114] 2Test results:

[0115] like Figure 5 As shown in the figure, the degradation dynamics of acetochlor by strain PY4 conforms to the first-order kinetic equation. The first-order kinetic equation (C = C0e -kt ) fitting, after fitting, it can be seen that the degradation half-life (T 1 / 2 =ln2 / k) were 14.99d and 5.24d respectively (see Table 5). Kinetic analysis showed that its degradation followed the first-order kinetic law, indicating that strain PY4 promoted the degradation of acetochlor in saline-alkali soil and that the degradation ability of the strain was stable and efficient.

[0116] Table 5 Kinetic parameters of acetochlor degradation by strain PY4

[0117] deal with Degradation kinetic equation Correlation coefficient k <![CDATA[Half-life T 1 / 2 > <![CDATA[R 2 ]]> CK <![CDATA[y=10.74e (-0.04624t) ]]> 0.04624 14.99 0.9921 PY4 <![CDATA[y=10.74e (-0.1322t) ]]> 0.1322 5.24 0.9974

[0118] Example 6 Field effect test of strain PY4

[0119] 1 Experimental design:

[0120] Effect test on saline-alkali soil:

[0121] In the saline-alkali soil of Bincheng District, Binzhou, the degradation effect of strain PY4 on acetochlor was tested. The soil pH was 7.93 and the water-soluble total salt content was 5.84‰. After spraying acetochlor evenly on the soil, it was ploughed and left to stand for 1 day to allow the acetochlor and soil to mix and reach a stable state. After that, the bacterial agent was applied (that is, the strain PY4 was cultured in NB medium for 4 days, the spores were collected by centrifugation, and then resuspended with sterile water to prepare spore suspensions of different concentrations) for remediation tests. The theoretical acetochlor pollution value of the soil is 5 mg / kg, and the initial acetochlor content was determined to be 5.37 mg / kg. The following treatment groups were set up: CK: no bacterial agent was added; T1: the concentration of the added bacterial agent was 1×10 8 CFU / mL; T2: the concentration of added bacterial agent is 1×10 7 CFU / mL; T3: the concentration of added bacterial agent is 5×10 6 CFU / mL. The plot area of ​​each treatment group was 24m 2 (6m × 4m), with three replicates per treatment group. Soil samples from the 0-20cm topsoil layer were collected using the five-point sampling method on days 3, 7, and 15 after inoculant application. After drying at 65°C, they were ground and passed through a 60-mesh sieve for subsequent determination of acetochlor content. The extraction and determination of acetochlor in soil were performed as in Example 5. The degradation rate was calculated using the following formula:

[0122] Acetochlor degradation rate (%) = (C0-C t ) / C0×100%.

[0123] Where, C0 is the initial content of acetochlor in the soil, unit is mg / kg; C t It is the content of acetochlor in the soil at time t, in mg / kg.

[0124] Normal soil effect test:

[0125] The degradation of acetochlor by strain PY4 was tested in normal soil from Ningyang County, Tai'an City. The soil had a pH of 7.44 and a water-soluble total salt content of 0.85‰. Acetochlor was evenly sprayed on the soil and then plowed. After standing for 1 day to allow the acetochlor and soil to mix and reach a stable state, a microbial agent was applied for remediation. The theoretical acetochlor contamination value in the soil is 5 mg / kg, and the initial acetochlor content was determined to be 5.10 mg / kg. The following treatment groups were set up: CK: no microbial agent added; T: microbial agent added at a concentration of 1×10 7 CFU / mL. The plot area of ​​each treatment group was 48m 2 Three replicates were set up for each treatment group. Soil samples from the 0-20 cm topsoil layer were collected using the five-point sampling method on days 3, 7, and 15 after inoculant application. After drying at 65°C, they were ground and passed through a 60-mesh sieve for subsequent determination of acetochlor content. The determination method was the same as above.

[0126] 2 Test results

[0127] Depend on Figure 6 It can be seen that different bacterial agent concentrations have significantly different effects on the degradation of acetochlor in saline-alkali soil. The degradation rates of acetochlor in each treatment group are: T2>T1>T3>CK, indicating that the optimal concentration of strain PY4 for degradation of acetochlor in saline-alkali soil is 1×10 7 CFU / mL. As time went on, the degradation rates of the treatment groups with different concentrations of the inoculant all showed an upward trend, indicating that strain PY4 colonized the saline-alkali soil and decomposed acetochlor as a carbon source for its growth metabolism. On the 3rd and 7th days, the degradation rate of the T1 treatment group was significantly higher than that of the T3 treatment group, but by the 15th day, the difference in degradation rates between the two treatment groups was small, indicating that 1×10 8 CFU / mL concentration of the bacterial agent showed a significant advantage in the degradation of acetochlor from 0 to 7 days, and on the 15th day, it was significantly better than the concentration of 5×10 6 The degradation advantage of the CFU / mL treatment group was no longer obvious. On the 15th day, the degradation rate of acetochlor in the T2 treatment group reached 91.50%, which was 137.97% higher than that in the CK treatment group.

[0128] Depend on Figure 7 It can be seen that 1×10 7 The CFU / mL inoculant had a significant degradation effect on acetochlor in normal soil. On the 7th and 15th days, the degradation rates of the T treatment group reached 88.02% and 97.13%, respectively, which were 128.92% and 75.45% higher than those of the CK group.

[0129] The above description is merely a preferred embodiment of the present application and is not intended to limit the scope of the present application. Although the present application has been described in detail with reference to the above embodiments, those skilled in the art may, after reading this description, make various modifications to the technical solution or replace some of the technical features with equivalents. Any modification, equivalent replacement, or improvement made within the spirit and principles of the present application shall be deemed to fall within the scope of protection of the present application.

Claims

1. A salt- and alkali-tolerant Streptomyces caulkinii strain, characterized in that: It was named Streptomyces caulkinii ( Streptomyces cavourensis ) PY4, which was deposited in the China Center for Type Culture Collection, Wuhan University, Wuhan, China on November 21, 2024, with the deposit number being CCTCC NO: M 20242608.

2. The fermentation liquid of the salt-alkali tolerant Streptomyces caulkinii according to claim 1, characterized in that The Streptomyces cauliferus is cultured in a culture medium to obtain the product.

3. The fermentation liquid of the salt-alkali tolerant Streptomyces caulkinii according to claim 2, wherein The culture medium is selected from NB medium, ISP medium, LB medium and Gao's medium No.

1.

4. A product comprising the fermentation broth of the salt- and alkali-tolerant Streptomyces calvorus according to claim 1 or the salt- and alkali-tolerant Streptomyces calvorus according to any one of claims 2 to 3.

5. The product according to claim 4, characterized in that The product is a microbial agent, including suspension, liquid fermentation preparation, powder and granule.

6. Use of the salt- and alkali-tolerant Streptomyces calvorus according to claim 1, the fermentation broth of the salt- and alkali-tolerant Streptomyces calvorus according to any one of claims 2 to 3, or the product according to any one of claims 4 to 5 in the degradation of acetochlor.

7. The use according to claim 6, characterized in that The degrading of acetochlor includes degrading acetochlor in the environment, and the environment includes soil and water.

8. The use according to claim 7, characterized in that The soil includes ordinary soil and saline-alkali soil.

9. A method for degrading acetochlor, characterized in that: The method comprises: applying the salt-alkali tolerant Streptomyces calvorus according to claim 1 or the fermentation broth of the salt-alkali tolerant Streptomyces calvorus according to any one of claims 2-3 or the product according to any one of claims 4-5 to the object to be treated that is contaminated with acetochlor.

10. The method according to claim 9, characterized in that When applied to the treated material, the initial concentration of viable bacteria of salt-alkali tolerant Streptomyces caulkinii was 1×10 6 -1×10 8 CFU / mL; the application amount is 0.2-2% of the amount of the substance to be treated.

11. The method according to claim 9, characterized in that The pH of the object to be treated is 6-9, and the water-soluble total salt content thereof is 0.1-20‰.

12. The method according to claim 9, characterized in that Before being applied to the object to be treated, the method further includes adjusting the moisture content of the object to be treated so that the moisture content of the object to be treated is adjusted to 60-80% of its maximum water holding capacity, w / w.

13. The method according to claim 9, characterized in that The objects to be treated include soil, water and sediment.

Images