Escherichia coli Top 10-A and method for degrading chloroaniline by using Escherichia coli Top 10-A

Abstract

The invention discloses Escherichia coli Top 10-A and a method for degrading chloroaniline by using the Escherichia coli Top 10-A. The preservation number of the Escherichia coli Top 10-A is CCTCC (China Center for Type Culture Collection) NO: M20242002, and the Escherichia coli Top 10-A is preserved in China Center for Type Culture Collection on September 19, 2024. In the invention, the Escherichia coli Top 10-A with the preservation number of CCTCC NO: M 20242002 is separated from industrial sludge for the first time, has the morphological characteristics of short rod shape and gram staining negative, is confirmed as a new functional strain through molecular biological identification, and has uniqueness and repeatability. The strain can specifically grow by taking chloroaniline as a unique carbon and nitrogen source, additional organic nutrition is not needed, the culture cost is reduced, compared with a commercially available strain ATCC 25922, the 7-day degradation rate of the strain reaches 38.1% which is far higher than that of a control strain by less than 1%, the core problem that the existing microbial degradation efficiency is low is solved, the specific degradation advantage aiming at chloroaniline is shown, and the application prospect is wide. A brand new strain resource is provided for the development of a biodegradation agent.

Description

Technical Field

[0001] The present invention relates to the technical field of microorganisms, in particular to an Escherichia coli Top10-A and a method for degrading chloroaniline. Background Art

[0002] Chloroaniline substances, when in contact with the skin, can cause local irritation symptoms such as redness, swelling, pain, and blisters. In severe cases, they can cause burns and ulcers, affecting the skin's normal function. Inhalation of chloroaniline vapor or dust can cause severe respiratory irritation, leading to symptoms such as coughing, wheezing, and difficulty breathing. Inhalation of high concentrations can also cause chemical pneumonia, pulmonary edema, and even life-threatening conditions. Long-term exposure to chloroaniline substances can damage the blood system, leading to methemoglobinemia, which reduces the blood's oxygen-carrying capacity and causes tissue hypoxia, resulting in symptoms such as headaches, dizziness, fatigue, and cyanosis. These substances can be metabolized and transformed in the liver, causing toxic effects on liver cells, leading to liver damage and abnormal liver function, manifested by symptoms such as elevated liver enzymes and jaundice. Long-term accumulation can lead to serious liver diseases such as cirrhosis. Chloroaniline substances also pose a significant threat to the environment: if they enter water bodies, they can severely damage aquatic ecosystems. They are difficult to degrade in water and remain there for a long time, affecting water quality, turning the water black and smelly, reducing dissolved oxygen levels, leading to oxygen deprivation and death in aquatic organisms, disrupting the aquatic food chain, and affecting biodiversity. Once in the soil, chloroanilines adsorb on the surface of soil particles, affecting the soil's physical, chemical, and biological properties, inhibiting the activity of microorganisms, reducing soil fertility and self-purification capacity, and ultimately affecting plant growth and development, leading to reduced crop yields and quality.

[0003] At present, the methods for treating chloroaniline mainly include physical methods and chemical methods, but these methods have certain limitations.

[0004] 1. Adsorption: This can be considered a physical enrichment process and is often used as a pretreatment method, but it is difficult to achieve true complete degradation. The operation is relatively simple and can effectively reduce the concentration of chloroanilines. However, after adsorption saturation, the adsorption material must be treated to avoid secondary pollution.

[0005] 2. Membrane separation: This method achieves separation based on physical screening and membrane permeability. It offers good separation performance, can be operated at room temperature, exhibits no phase change, and is environmentally friendly. However, the membranes are expensive and can be contaminated during operation, impacting both treatment effectiveness and service life.

[0006] 3. Photocatalytic oxidation method: It involves physical light radiation and chemical reaction. The reaction conditions are mild and can be carried out at room temperature and pressure. It can deeply mineralize chloroaniline substances, but the activity and stability of the photocatalyst need to be further improved, and the light utilization rate is low.

[0007] 4. Electrochemical method: Degradation of pollutants is achieved through electrode reactions. This method has the advantages of simple operation, fast reaction speed, and no secondary pollution, but it has high energy consumption. The choice of electrode materials and service life have a significant impact on treatment effect and cost.

[0008] Microbial degradation, currently the primary method for degrading chloroanilines, offers the following advantages: 1. Environmentally friendly: Compared to physical and chemical methods, microbial degradation generally does not produce secondary pollution. Microorganisms can convert chloroanilines into carbon dioxide, water, and harmless small molecules, achieving harmless treatment without the toxic or harmful byproducts or residual substances that some chemical treatment methods produce. 2. Low cost: Microbial degradation generally does not require high temperatures, high pressures, or strong oxidants, resulting in relatively low operating costs. Once the microbial community is successfully cultivated, it only requires suitable environmental conditions and necessary nutrients for sustained degradation, eliminating the need for large amounts of chemical agents and energy. 3. High specificity: Different microorganisms have a high degree of specificity for recognizing and degrading chloroanilines with specific structures. By screening and cultivating specific microbial strains capable of specifically degrading target chloroaniline pollutants, it is possible to identify or cultivate specific microorganisms for different types of chloroaniline pollution. 4. In situ treatment: Microbial degradation can be carried out at the site of contamination, eliminating the need to transport the pollutants to specialized treatment facilities, reducing transportation costs and risks. For example, in soil or water contaminated with chloroanilines, degrading agents can be directly added or bioaugmentation can be used to promote in situ microbial degradation of chloroanilines. 5. Resource Recovery Potential: The microbial degradation of chloroanilines can, in some cases, lead to resource recovery. For example, some microorganisms may produce economically valuable metabolites during the degradation process, or release certain elements from chloroanilines through degradation, which can be further recycled. 6. Strong Sustainability: Microorganisms have the ability to self-replicate and renew. As long as environmental conditions are favorable, microbial populations can continue to grow and reproduce, maintaining their degradation capabilities. Furthermore, microorganisms can gradually adapt to environmental changes during evolution, potentially developing enhanced degradation capabilities through natural selection and genetic variation, making the degradation process more sustainable. 7. Complete Degradation: Microorganisms can gradually decompose chloroanilines through a series of enzymatic reactions, ultimately achieving complete mineralization and converting them into inorganic substances, fundamentally eliminating the toxicity and hazards of the pollutants, rather than simply converting one form of the pollutant into another.

[0009] Although microorganisms can degrade PCA, their efficiency in complex environments remains low. Further efforts are needed to identify and cultivate microbial strains with PCA degradation capabilities, or to construct microbial communities and optimize microbial metabolic pathways to improve PCA degradation efficiency. Therefore, identifying strains capable of PCA degradation is of great significance. Summary of the Invention

[0010] In order to overcome the above-mentioned defects of the prior art, the embodiments of the present invention provide an Escherichia coli Top 10-A and a method for degrading chloroanilines, which solve the problems of the existing chloroanilines being highly harmful, the existing treatment methods being limited, and the low efficiency of microbial degradation.

[0011] To achieve the above object, the present invention provides the following technical solutions:

[0012] An Escherichia coli Escherichiacoli Top10-A, the strain has a preservation number of CCTCC NO: M20242002, and was deposited in the China Center for Type Culture Collection on September 19, 2024.

[0013] Preferably, the strain has the morphological characteristics of a short rod with blunt ends, Gram-negative staining, and an approximately spherical rod shape under an electron microscope.

[0014] Preferably, the invention relates to a use of Escherichia coli Top10-A in the preparation of a chloroaniline degradation agent.

[0015] Preferably, a method for degrading chloroaniline by Escherichia coli Top10-A comprises the following steps:

[0016] S1: inoculating the Escherichia coli according to claim 1 into a mineral salt medium (MSB) containing chloroaniline as the sole carbon source, nitrogen source and energy source;

[0017] S2: Cultivating the strain in the culture medium to allow the strain to degrade chloroaniline.

[0018] Preferably, the liquid culture medium formula of the inorganic salt culture medium (MSB) is: 1gNa2HPO4, 0.5gKH2PO4, 0.03gMgSO4·7H2O, 0.05g yeast powder, 5mL trace element solution per liter of distilled water, and the pH value is 7.0±0.2.

[0019] Preferably, the concentration of chloroaniline in the culture medium is 200-400 mg / L, preferably 300 mg / L.

[0020] Preferably, step S1 includes a strain activation step before inoculation: the Escherichia coli is cultured in MSB liquid culture medium at 28±2°C and 120±10 rpm on a shaking platform for 48 hours, and after obtaining the activated liquid, it is inoculated into a culture medium containing chloroaniline at an inoculation rate of 5%-15% (preferably 10%).

[0021] Preferably, the control parameters during the culture process are as follows:

[0022] Temperature: 28±2℃;

[0023] Dissolved oxygen content: 20%-30% (preferably 25%);

[0024] pH value: maintained at 7.0 ± 0.2 by phosphate buffer.

[0025] Preferably, the degradation rate of chloroaniline by the Escherichia coli is ≥38.1% within 7 days and ≥39.0% within 15 days, and is significantly higher than that of the control strain Escherichia coli ATCC25922.

[0026] Preferably, when the inorganic salt culture medium is a solid culture medium, 15 g / L agar powder is added for the isolation and purification or slant preservation of the strain, and the concentration of chloroaniline in the solid culture medium is 200-400 mg / L.

[0027] The present invention provides an Escherichia coli Top10-A and a method for degrading chloroaniline, and its technical effects and advantages are as follows:

[0028] 1. This invention, for the first time, isolated Escherichia coli Top 10-A from industrial sludge with the preservation number CCTCC NO: M 20242002. Its morphological characteristics are short rods and Gram-negative. It was confirmed as a new functional strain through molecular biological identification, which is unique and reproducible. This strain can specifically grow with chloroaniline as the only carbon and nitrogen source, without the need for additional organic nutrients, simplifying the cultivation cost. Compared with the commercially available strain ATCC 25922, its 7-day degradation rate reached 38.1%, far exceeding the control strain's <1%, solving the core problem of low degradation efficiency of existing microorganisms, showing the specific degradation advantage of chloroaniline, and providing a new strain resource for the development of biodegradants.

[0029] 2. This invention, by limiting key parameters such as 300 mg / L chloroaniline concentration, 28°C culture temperature, 25% dissolved oxygen content and pH 7.0, constructs the optimal degradation conditions for the strain. The degradation rate reaches 38.1% in 7 days and stabilizes at more than 39.0% in 15 days, achieving stable degradation. At the same time, the flexible design of the concentration range (200-400 mg / L) and inoculation size (5%-15%) adapts to the fluctuations in pollutant concentrations and degradation needs of different scales in actual polluted environments, thereby improving the universality of the method. Solid culture medium (containing 15 g / L agar powder) supports the isolation, purification and long-term preservation of strains, forming a complete technical chain of "screening-activation-degradation-preservation", meeting the requirements for strain preparation and storage in industrial production, and promoting the application of technology.

[0030] 3. Compared to the secondary pollution risk of physical adsorption methods and the high energy consumption of chemical oxidation methods, this invention utilizes microbial degradation technology to convert chloroanilines into harmless small molecules through bacterial metabolism, achieving harmless treatment and conforming to the trend of green environmental protection. The culture medium contains only inorganic salts, trace yeast powder, and target pollutants, reducing raw material costs by over 50% compared to traditional culture media. It also eliminates the need for harsh conditions such as high temperature and high pressure, significantly reducing equipment and operating costs. The strain can be directly added to contaminated soil or water for in-situ degradation, avoiding the transportation costs and risks of pollutant transfer. It is suitable for large-scale environmental remediation, offering both environmental and economic benefits, and provides a low-cost, sustainable solution for the treatment of chloroaniline pollutants. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 This is a culture diagram of Escherichia coli Top 10-A of the present invention;

[0032] Figure 2 This is an electron microscope image of a stained Escherichia coli Top 10-A strain of the present invention;

[0033] Figure 3 This is a comparison diagram of a method for degrading chloroaniline by Escherichia coli Top 10-A of the present invention. DETAILED DESCRIPTION

[0034] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts shall fall within the scope of protection of the present invention.

[0035] It should be noted that, in this document, relational terms such as first and second, etc. are only used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any actual relationship or order between these entities or operations. Moreover, the terms "include", "comprise" or any other variations thereof are intended to cover non-exclusive inclusion, so that a process, method, article or apparatus that includes a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements inherent to such process, method, article or apparatus. In the absence of further restrictions, the elements defined by the sentence "include..." do not exclude the existence of other identical elements in the process, method, article or apparatus that includes the elements.

[0036] The present invention provides an Escherichia coli Top 10-A and a method for degrading chloroaniline. The technical solution is described in detail below with reference to examples.

[0037] Strain information

[0038] Escherichia coli Top 10-A (deposit number: CCTCC NO: M 20242002) was identified by morphological observation and 16S rRNA gene sequencing. Its morphological characteristics are: short rod shape with blunt ends, Gram-negative staining, and approximately spherical club shape under electron microscopy. It was deposited in the China Center for Type Culture Collection on September 19, 2024.

[0039] Example 1

[0040] refer to Figure 1-3 This embodiment provides an Escherichia coli Top 10-A and a method for degrading chloroaniline, which are used for strain screening and identification. The specific implementation content includes:

[0041] Implementation purpose:

[0042] Verify the isolation process of the strain, confirm its morphological characteristics and preservation information, and support the uniqueness of the strain in claims 1-2.

[0043] Implementation steps:

[0044] (1) Sludge sampling and enrichment culture: 10 g of sludge sample from the sewage treatment plant was inoculated into 100 mL of MSB liquid culture medium containing 200 mg / L chloroaniline (formula: 1 g Na2HPO4, 0.5 g KH2PO4, 0.03 g MgSO4·7H2O, 0.05 g yeast powder, 5 mL trace element solution, pH 7.0 per liter). The culture was shaken at 30°C and 200 rpm for 7 days until the culture medium became turbid.

[0045] (2) Subculture and acclimatization: The enrichment solution was transferred to fresh MSB medium (containing 200 mg / L chloroaniline) at a 1% inoculum volume, and repeated three times to screen out strains that use chloroaniline as the sole carbon and nitrogen source;

[0046] (3) Streak isolation: Spread the acclimation solution on MSB solid medium (containing 200 mg / L chloroaniline) containing 15 g / L agar powder, incubate at 28°C for 48 h, pick a single colony and purify by continuous streaking until a single morphological colony is obtained;

[0047] (4) Morphological and molecular identification: Gram staining showed negative short rods, and electron microscopy showed a rod-like shape ( Figure 1); genomic DNA was extracted, and the 16S rRNA gene was amplified and sequenced. The alignment results showed that the homology with Escherichia coli was ≥99%, and it was named Escherichia coli Top 10-A.

[0048] Implementation effect: The target strain was successfully isolated and obtained, and its morphological characteristics were consistent with claim 2. The preservation information was complete and could be uniquely identified by the preservation number CCTCC NO: M 20242002.

[0049] Example 2

[0050] This embodiment provides an Escherichia coli Top 10-A and a method for degrading chloroaniline, which is used for degrading chloroaniline under optimal degradation conditions. The specific implementation content includes:

[0051] Implementation purpose:

[0052] The core degradation method of the verification clearly defines the degradation efficiency under the conditions of 300 mg / L concentration, 28°C, 25% dissolved oxygen content, and pH 7.0.

[0053] Implementation steps:

[0054] (1) Strain activation: Frozen Top 10-A was inoculated into 50 mL of MSB liquid medium and cultured in a shaking incubator at 28°C and 120 rpm for 48 h to obtain an activation solution;

[0055] (2) Degradation culture: The activation solution was inoculated at 10% of the inoculum into 100 mL of MSB culture medium containing 300 mg / L chloroaniline (the formula is the same as in Example 1) and placed in a tabletop fermenter. The control parameters were as follows:

[0056] Temperature: 28°C

[0057] Dissolved oxygen content: adjusted to 25% by stirring rate;

[0058] pH value: automatically add 1M NaOH / HCl to maintain 7.0;

[0059] (3) Sampling and testing: Samples were collected on the 1st, 3rd, 5th, 7th, 10th, and 15th day of culture, and the supernatant was collected after centrifugation. The concentration of chloroaniline was determined by liquid chromatography (HPLC), and the degradation rate was calculated:

[0060]

[0061] Implementation effect: The degradation rate changes over time as shown in the following table:

[0062] Time (days) 1 3 5 7 10 15 Degradation rate% 12.9 22.7 30.6 38.1 38.9 39.0

[0063] The results showed that it could be effectively degraded in the concentration range of 200-400 mg / L, and the degradation rate was the highest at 300 mg / L.

[0064] Example 3

[0065] This embodiment provides an Escherichia coli Top 10-A and a method for degrading chloroaniline, which is used to determine the effect of chloroaniline concentration on degradation rate. The specific implementation includes:

[0066] Implementation purpose: To verify the validity of the concentration range (200-400 mg / L) described in claim 6 and determine the optimal concentration.

[0067] Implementation steps:

[0068] (1) Three parallel experiments were conducted. The concentrations of chloroaniline in the culture medium were 200 mg / L, 300 mg / L (control), and 400 mg / L, respectively. The remaining conditions were the same as those in Example 2 (inoculation amount 10%, 28° C., 25% dissolved oxygen, pH 7.0). The degradation rate was detected after 7 days of culture.

[0069] Implementation effect: The effect of chloroaniline concentration on degradation rate is shown in the following table:

[0070] Concentration (mg / L) 200 300 400 Degradation rate (%) 35.2 38.1 36.5

[0071] Example 4

[0072] This example provides an Escherichia coli Top 10-A and a method for degrading chloroaniline, and describes the effect of inoculation amount on degradation efficiency. The specific implementation includes:

[0073] Implementation purpose: To verify the rationality of the inoculum amount range (5%-15%) described in claim 7 and to determine the optimal inoculum amount.

[0074] Implementation steps:

[0075] (1) Three experiments were conducted with inoculum concentrations of 5%, 10% (control), and 15%. The remaining conditions were the same as those in Example 2 (300 mg / L concentration, 28° C., 25% dissolved oxygen, pH 7.0). The degradation rate was detected after 7 days of culture.

[0076] Implementation effect: The effect of chloroaniline concentration on degradation rate is shown in the following table:

[0077] Inoculation amount (%) 5 10 15 Degradation rate (%) 32.3 38.1 37.8

[0078] The results showed that 5%-15% inoculation amount was feasible, and the degradation efficiency was optimal at 10%.

[0079] Example 5

[0080] This embodiment provides an Escherichia coli Top 10-A and a method for degrading chloroaniline, which are used for separation, purification and preservation of solid culture medium. The specific implementation content includes:

[0081] Purpose of the experiment:

[0082] Verify the application of the solid culture medium (containing 15g / L agar powder) described in claim 10 in strain isolation, purification and preservation.

[0083] Experimental procedures

[0084] (1) Isolation and purification: The enriched liquid of Example 1 was spread on MSB solid medium (containing 15 g / L agar powder) containing 300 mg / L chloroaniline, cultured at 28°C for 48 h, and a single colony was picked and streaked three times to obtain a pure culture.

[0085] (2) Slant preservation:

[0086] The purified strain was inoculated into MSB solid slant medium (containing 200 mg / L chloroaniline and 15 g / L agar powder), stored at 4°C, and revived after 3 months. There was no significant decrease in the degradation ability.

[0087] Experimental results: Solid culture medium can effectively separate single colonies ( Figure 2 ), and the degradation rate of the slant-preserved strain after recovery was consistent with that of the fresh strain (7-day degradation rate 38.0%), supporting the use of the solid culture medium in claim 10.

[0088] Comparative Example 1

[0089] This comparative example provides a comparison of the degradation effects of the control strain, and the specific comparison includes:

[0090] Implementation purpose: To verify the technical effect of "significantly higher than the control strain ATCC 25922" as stated in claim 9.

[0091] Implementation steps: Using Escherichia coli ATCC 25922 (purchased from the American Type Culture Collection), a degradation experiment was carried out according to the method of Example 2 (300 mg / L concentration, 28° C., 25% dissolved oxygen, pH 7.0). The degradation rate was measured after culturing for 7 days.

[0092] Implementation effect: The control strain ATCC25922 had almost no change in chloroaniline concentration within 7 days, and the degradation rate was less than 1%, which was significantly lower than the 38.1% of the Top10-A strain, directly proving the specific degradation advantage of the strain of the present invention.

[0093] Description of the detection method:

[0094] HPLC conditions: Column: C18 column (4.6 × 250 mm, 5 μm);

[0095] Mobile phase: methanol: water = 60:40 (v / v);

[0096] Flow rate: 1.0 mL / min;

[0097] Detection wavelength: 230nm;

[0098] Injection volume: 20 μL.

[0099] Dissolved oxygen detection: An online dissolved oxygen electrode (Mettler-Toledo) was used for real-time monitoring, and the dissolved oxygen content was controlled by adjusting the stirring rate (200-800 rpm).

[0100] pH control: Automatically add 1M NaOH or HCl solution to maintain the pH value at 7.0±0.2.

[0101] Uniqueness of the strain: Through the screening and identification in Example 1, it was proved that Top 10-A is a new strain that can be uniquely identified by its deposit number and morphological characteristics and has not been disclosed in the prior art.

[0102] Method effectiveness: Examples 2-4 verify the optimal combination of culture medium formula, concentration, inoculation amount, and culture conditions in the degradation method, forming a complete process parameter system to support the multi-level protection of claims 4-8.

[0103] Superiority of effect: The degradation rate data of Example 2 and the control experiment of Comparative Example 1 directly prove that the strain of the present invention is significantly superior to the existing strain in degradation efficiency, meeting the requirements of creativity.

[0104] Application integrity: Example 5 covers the separation and preservation of solid culture media, extending the scope of protection of the claims to the entire process of strain cultivation, enhancing the practicality and industrial value of the technical solution.

[0105] Those skilled in the art will appreciate that the modules and algorithm steps of the various examples described in conjunction with the embodiments disclosed herein can be implemented with electronic hardware, or a combination of computer software and electronic hardware. Whether these functions are performed in hardware or software depends on the specific application and design constraints of the technical solution. Professional and technical personnel can use different methods to implement the described functions for each specific application, but such implementation should not be considered to be beyond the scope of this application.

[0106] In addition, each functional module in each embodiment of the present application may be integrated into one processing module, or each module may exist physically separately, or two or more modules may be integrated into one module.

[0107] The above is only a specific implementation method of the present application, but the scope of protection of the present application is not limited to this. Any technician familiar with this technical field can easily think of changes or replacements within the technical scope disclosed in this application, which should be included in the scope of protection of the present application. Therefore, the scope of protection of the present application should be based on the scope of protection of the claims.

[0108] Finally: The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. An Escherichia coli Top10-A, characterized in that The deposit number of the strain is CCTCC NO: M20242002, and it was deposited in the China Center for Type Culture Collection on September 19, 2024.

2. The Escherichia coli Top10-A according to claim 1, wherein The strain has the morphological characteristics of being short rod-shaped, blunt-ended at both ends, Gram-negative in gram staining, and approximately spherical rod-shaped under an electron microscope.

3. Use of the Escherichia coli Top10-A according to claim 1 in preparing a chloroaniline degradation agent.

4. A method for degrading chloroaniline using Escherichia coli Top10-A, characterized in that: The following steps are involved: S1: inoculating the Escherichia coli according to claim 1 into a mineral salt medium (MSB) containing chloroaniline as the sole carbon source, nitrogen source and energy source; S2: Cultivating the strain in the culture medium to allow the strain to degrade chloroaniline.

5. The method for degrading chloroaniline by using Escherichia coli Top10-A according to claim 4, characterized in that: The liquid culture medium formula of the inorganic salt culture medium (MSB) is as follows: 1g Na2HPO4, 0.5g KH2PO4, 0.03g MgSO4·7H2O, 0.05g yeast powder, 5mL trace element solution per liter of distilled water, and the pH value is 7.0±0.

2.

6. The method for degrading chloroaniline using Escherichia coli Top10-A according to claim 5, characterized in that: The concentration of chloroaniline in the culture medium is 200-400 mg / L, preferably 300 mg / L.

7. The method for degrading chloroaniline using Escherichia coli Top10-A according to claim 4, characterized in that: Step S1: Before inoculation, the strain activation step is included: the Escherichia coli is cultured in MSB liquid culture medium at 28±2°C and 120±10 rpm on a shaking platform for 48 hours, and after obtaining the activated liquid, it is inoculated into a culture medium containing chloroaniline at an inoculum amount of 5%-15% (preferably 10%).

8. The method for degrading chloroaniline using Escherichia coli Top10-A according to claim 4, wherein: The control parameters during the culture process are as follows: Temperature: 28±2℃; Dissolved oxygen content: 20%-30% (preferably 25%); pH value: maintained at 7.0 ± 0.2 by phosphate buffer.

9. The method for degrading chloroaniline using Escherichia coli Top10-A according to claim 4, wherein: The degradation rate of chloroaniline by the Escherichia coli is greater than or equal to 38.1% within 7 days and greater than or equal to 39.0% within 15 days, and is significantly higher than that of the control strain Escherichia coli ATCC25922.

10. The method for degrading chloroaniline using Escherichia coli Top10-A according to claim 4, characterized in that: When the inorganic salt culture medium is a solid culture medium, 15 g / L agar powder is added for separation and purification of the strain or slant preservation. The concentration of chloroaniline in the solid culture medium is 200-400 mg / L.

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