Bacillus C8 for reducing chromium pollution of soil and application thereof

Abstract

The invention relates to bacillus C8 for reducing chromium pollution of soil and application thereof, belongs to the technical field of microorganisms, and provides bacillus (Bacillus sp.) C8 for reducing chromium pollution of soil, the bacillus C8 is preserved in China General Microbiological Culture Collection Center on October 12, 2024, the preservation address is No. 3, No.1 Yard, Beichen West Road, Chaoyang District, Beijing, and the preservation number is CGMCC NO.3669. The preservation number is CGMCC (China General Microbiological Culture Collection Center) The invention provides application of bacillus C8 in reducing the chromium content of soil. The bacillus C8 is used for reducing chromium pollution in soil, the production cost is low, the production method is simple, the chromium control effect is good, and the method can be applied to sunflower planting in chromium-polluted farmland in a large area.

Description

Technical Field

[0001] The invention belongs to the technical field of microorganisms, and in particular relates to a Bacillus C8 for reducing soil chromium pollution and an application thereof. Background Art

[0002] With the advancement of urbanization and industrialization, the problem of heavy metal pollution has become increasingly prominent. At present, soil is mainly polluted by heavy metals, and chromium (Cr) is the main heavy metal pollutant. Heavy metal pollution in cultivated soil has a serious negative impact on crop production. In addition, chromium can also be enriched in the human body through the food chain, endangering human health.

[0003] Studies have shown that existing methods for reducing soil chromium pollution include the comprehensive use of multiple technologies, such as ex situ leaching, direct reduction, and plant methods. For heavily chromium-contaminated soil, ex situ leaching is an effective treatment technology. By using different eluents (such as deionized water, citric acid, oxalic acid, EDTA, etc.), hexavalent chromium in the soil can be extracted under specific conditions and reduced to non-toxic trivalent chromium. The effect of this method is affected by the concentration of the eluent and the reaction temperature. The direct reduction method uses a reducing agent such as ferrous sulfate to reduce hexavalent chromium to trivalent chromium, and adjusts the pH value by quicklime to form a stable hydroxide precipitate, thereby achieving soil remediation. The plant method removes heavy metals from the soil through the absorption capacity of plants, including plant extraction technology, root filtration technology, etc. In addition, fertilization and additive methods are also effective remediation methods. By increasing the organic matter content in the soil, the soil's adsorption capacity for heavy metals is improved, thereby reducing its harm to plants. The additive method uses adhesives and soil conditioners to improve soil structure and reduce the mobility of heavy metals. For example, biochar, nanomaterials, glutamic acid, gibberellin acid and 5-aminolevulinic acid can reduce the toxicity of chromium to crops. However, the production costs of the above materials are relatively high and they are not completely suitable for large-scale application in farmland. Rhizospheric bacteria have been widely used in agricultural production in recent years due to their high reproduction coefficient and low cost. Summary of the invention

[0004] In order to solve the above technical problems, the present invention proposes a Bacillus C8 for reducing soil chromium pollution and its application. Bacillus C8 is used to reduce chromium pollution in the soil. The production cost is low, the production method is simple, the chromium prevention and control effect is good, and it can be widely used in sunflower planting in chromium-contaminated farmland.

[0005] To achieve the above-mentioned purpose, the present invention provides a Bacillus sp. C8 for reducing soil chromium pollution. The Bacillus sp. C8 was deposited in the General Microbiology Center of China Microbiological Culture Collection Committee on October 12, 2024, with the deposit address at No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, and the deposit number is: CGMCC NO.32191.

[0006] The present invention also provides the use of the Bacillus C8 in reducing the chromium content in soil.

[0007] The present invention also provides a preparation for reducing the chromium content in soil, wherein the preparation comprises the Bacillus C8.

[0008] Preferably, the effective viable count of Bacillus C8 in the preparation is 1.0×10 8 ~1.2×10 8 cfu / mL.

[0009] The present invention also provides a method for reducing chromium pollution in soil by using the Bacillus C8, comprising the following steps: 8 ~1.2×10 8 The Bacillus C8 bacterial solution with a cfu / mL was mixed with the soil at a ratio of 200-300mL:1-1.5kg.

[0010] The present invention also provides the use of the Bacillus C8 in alleviating plant chromium stress.

[0011] Preferably, the effective viable count of Bacillus C8 is 1.0×10 8 ~1.2×10 8 cfu / mL.

[0012] Preferably, the plant is sunflower.

[0013] The present invention also provides a preparation for alleviating chromium stress of sunflower, wherein the preparation comprises the Bacillus C8.

[0014] Preferably, the effective viable count of Bacillus C8 in the preparation is 1.0×10 8 ~1.2×10 8 cfu / mL.

[0015] Compared with the prior art, the present invention has the following advantages and technical effects:

[0016] The present invention provides a Bacillus C8 for reducing chromium pollution in soil. Bacillus C8 can reduce Cr in soil. 6+ The content of chromium can be reduced, thereby reducing the chromium hazard in the soil; the production cost of the Bacillus C8 of the present invention is low, the production method is simple, and the prevention and control effect of chromium pollution is good; planting sunflowers in chromium-contaminated soil treated with Bacillus C8 can alleviate the chromium stress of sunflowers, and Bacillus C8 can be widely used in the production of sunflowers in chromium-contaminated farmland. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0018] Figure 1 1 is a strain morphology diagram of Bacillus megaterium bacteria C1, Arthrobacter bacteria C6 and Bacillus C8, wherein A is a colony morphology diagram and B is a strain morphology diagram;

[0019] Figure 2 The experimental results for screening candidate strains resistant to chromium;

[0020] Figure 3 The determination of the effect of Bacillus C8 on the prevention and treatment of chromium pollution, where A is 0.03 μmol / mL Cr 6+ Chromium content in sunflower roots of the treatment groups, B is the growth status of sunflowers in the soil treatment group with a chromium content of 100 mg / kg, Cr represents the chromium pollution group, and Cr+C8 represents the Bacillus C8 treatment group. DETAILED DESCRIPTION

[0021] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0022] It should be understood that the terms described in the present invention are only for describing special embodiments and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. Each smaller range between the intermediate value in any stated value or stated range and any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0023] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0024] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to the skilled artisan. The present invention description and examples are exemplary only.

[0025] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0026] Example 1

[0027] 1. Acquisition of chromium-resistant candidate strains

[0028] (1) Take 1 g of soil containing 100 mg / kg K2CrO4 and add it to a test tube containing 10 mL of sterile water. The test tube is defined as 10 -1 , and mix to obtain soil suspension.

[0029] (2) Add 1 mL of soil suspension into a test tube containing 9 mL of sterile water, defined as 10 -2 ; And so on, get 10 -3 , 10 -4 , 10 -5 , 10 -6 and 10 -7 .

[0030] (3) Take 200 μL of the diluted suspension 10 -7 , spread on beef extract peptone solid medium (0.3% beef extract, 1% peptone, 0.5% sodium chloride, 1% agar), invert at 28°C for 3 days, pick single colonies from the low-concentration diluent plate, perform colony PCR sequencing with 16S primers, and obtain strain information by sequencing.

[0031] 2. Screening of chromium-resistant candidate strains (1) Through 16S alignment, three strains with higher duplication rates were screened for verification, namely Bacillus megaterium ( Priestia megaterium ) Bacteria MnS1200909 referred to as C1, Arthrobacter genus ( Arthrobacter sp. ) bacteria C6 and Bacillus spp. ( Bacillus sp. ) Bacteria 201705CJKOP-59, referred to as C8.

[0033] like Figure 1 Middle A and Figure 1 Shown in B are the morphological diagrams of the three strains C1, C6 and C8.

[0034] (2) The three strains were tested for their in vitro chromium resistance using a 0.03 μmol / mL Cr 6+ The three strains C1, C6 and C8 were resuspended in the solution. 600 The total effective viable count of bacteria is 1.0×10 8 ~1.2×10 8 cfu / mL) and cultured at 28°C, 180 rpm for 48 h to determine the Cr 6+ Solution concentration.

[0035] like Figure 2 As shown, only in the C8 strain treatment group, Cr 6+ The concentration of Cr in the other two strain treatment groups decreased significantly. 6+ The concentration hardly changed, and finally the C8 strain was identified as a candidate strain resistant to chromium.

[0036] The C8 strain is a Bacillus sp. bacterium. After 16S sequencing, it was determined that the strain has a high homology with C8.

[0037]

[0038] III. Determination of the effect of strain C8 on preventing and treating chromium pollution

[0039] (1) Use an inoculation loop to dip a small amount of C8 strain preserved in glycerol, spread it on beef extract peptone medium, and culture it upside down at 28°C for 1 day.

[0040] (2) Take a single colony and place it in 5 mL of beef extract peptone liquid medium, culture it at 28°C, 200 rpm for 1 day; then inoculate 5 mL of the bacterial solution into 500 mL of beef extract peptone liquid medium, culture it at 28°C, 200 rpm for 1 day, and centrifuge the cultured bacterial solution to remove the supernatant.

[0041] (3) Using Cr at a concentration of 0.03 μmol / mL 6+ Resuspend the above-removed bacterial solution with 100mg / kg (1kg soil contains 100mg of potassium dichromate) solution to make the bacterial solution OD 600 is 0.6; then cultured at 28℃, 200rpm for 1d, the centrifuged bacteria were resuspended in chromium solution and directly watered the sunflower for 2d, and the control group was the above two concentrations of potassium dichromate solution without bacterial solution.

[0042] The chromium content was determined by ICP-MS, referring to the determination method of multiple elements in food in GB 5009.268-2016 National Food Safety Standard.

[0043] The results are as follows Figure 3 Middle A and Figure 3 As shown in B, after 2 days of sunflower treatment, the Cr concentration of 0.03 μmol / mL was measured in the presence of C8 strain (Cr+C8) and without C8 strain (Cr). 6+ The results showed that the chromium absorption in the sunflower roots of the experimental group with bacterial solution was significantly reduced; the leaves of the sunflowers in the 100 mg / kg treatment group had a drooping trend regardless of whether the bacteria were added or not, showing a toxic phenomenon; however, in the control group without bacteria, some sunflowers showed obvious wilting and death, indicating that the C8 strain of the present invention has the effect of significantly reducing the chromium content in the soil and helping plants resist chromium toxicity.

[0044] The embodiments described above are only descriptions of the preferred modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary technicians in this field should all fall within the protection scope determined by the claims of the present invention.

Claims

1. A Bacillus sp. C8 for reducing chromium pollution in soil, characterized in that: The Bacillus C8 was deposited in the General Microbiology Center of the China Microbiological Culture Collection Administration on October 12, 2024, with the deposit address at No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and the deposit number is: CGMCC NO.32191.

2. Use of Bacillus C8 as claimed in claim 1 in reducing the chromium content in soil.

3. A preparation for reducing the chromium content in soil, characterized in that: The preparation comprises the Bacillus C8 according to claim 1.

4. The preparation according to claim 3, characterized in that The effective viable count of Bacillus C8 in the preparation is 1.0×10 8 ~1.2×10 8 cfu / mL.

5. A method for reducing chromium pollution in soil using the Bacillus C8 of claim 1, characterized in that: The following steps are included: the effective viable count is 1.0×10 8 ~1.2×10 8 The Bacillus C8 bacterial solution with a cfu / mL was mixed with the soil at a ratio of 200-300mL:1-1.5kg.

6. Use of Bacillus C8 as claimed in claim 1 in alleviating plant chromium stress.

7. The use according to claim 6, characterized in that: The effective viable count of Bacillus C8 is 1.0×10 8 ~1.2×10 8 cfu / mL.

8. The use according to claim 6, characterized in that: The plant is sunflower.

9. A preparation for alleviating chromium stress in sunflower, characterized in that: The preparation comprises the Bacillus C8 according to claim 1.

10. The preparation according to claim 9, characterized in that The effective viable count of Bacillus C8 in the preparation is 1.0×10 8 ~1.2×10 8 cfu / mL.

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