Desertification remediation microbial agent and application thereof

Abstract

The invention belongs to the technical field of microorganisms, and relates to a desertification remediation fungicide and application thereof. The invention provides a desertification remediation microbial agent which is prepared by the following steps: respectively fermenting bacillus subtilis, geotrichum candidum and candida utilis to obtain bacterial liquids, and mixing the bacterial liquids. The bacillus subtilis, the geotrichum candidum and the candida utilis are synergistically matched, and when the bacillus subtilis, the geotrichum candidum and the candida utilis are applied to the desertification soil remediation process, the compressive strength and the shear strength of soil can be effectively improved, and the dosage of urea and calcium superphosphate is reduced.

Description

Technical Field

[0001] The present invention belongs to the technical field of microbiology and relates to a desertification restoration microbial agent and its application. Background Art

[0002] Desertification is a severe environmental problem faced globally. It not only leads to a significant decline in land productivity but also affects the sustainable development of human society. The causes of desertification are complex, including natural factors such as climate change and water resource shortage, as well as human factors such as overgrazing and unreasonable farming. These factors work together to cause soil structure damage, fertility decline, and vegetation reduction, ultimately forming desertification. Traditional methods for desertification control, such as afforestation and irrigation improvement, can improve the soil environment to a certain extent, but they are often costly and limited by water resources and geographical environment. Therefore, it is particularly important to find a low-cost, environmentally friendly, and sustainable method for desertified soil treatment. In recent years, microbial remediation technology has attracted much attention due to its unique advantages. As an important part of the soil ecosystem, microorganisms can have a positive impact on the soil environment through mechanisms such as decomposing organic matter, participating in nutrient cycling, and promoting plant growth. In the treatment of desertified soil, by utilizing the biological characteristics of microorganisms, the soil structure can be effectively improved, soil fertility can be enhanced, and vegetation restoration can be promoted, thereby achieving effective treatment of desertified soil.

[0003] ZL201510439948.1 discloses a Geotrichum candidum and its application in greenhouse soil remediation, which records that Geotrichum Candidum XHS0030B CGMCC NO.9435, in combination with Rhodopseudomonas palustris, Candida utilis, and Mortierella alpina, has the effect of greenhouse soil remediation. ZL201510439890.0 discloses a Bacillus subtilis and its application in greenhouse soil remediation, which records that Bacillus Subtilis XHS0035Kc CGMCC NO.9434, in combination with Pseudomonas fluorescens, Bacillus mucilaginosus, and Sporidiobolus salmonicolor, has outstanding technical effects in greenhouse soil remediation. However, there has been no literature report on the mixture of Geotrichum candidum XHS0030B and Bacillus subtilis XHS0035Kc for desertification restoration, and it is unknown whether there is a synergistic effect or an antagonistic effect when the two are mixed.

[0004] Although microbial remediation technology shows great potential in the field of soil treatment, its application in desertified soil treatment still faces some challenges. For example, the environment of desertified soil is harsh, making it difficult for microorganisms to survive and reproduce; at the same time, the microbial community structure in desertified soil is single, lacking microorganisms with efficient remediation functions. Therefore, how to screen and cultivate microorganisms adapted to the desertified soil environment and construct an effective microbial remediation system has become the focus of current research. Summary of the Invention

[0005] The purpose of the present invention is to construct a synergistic microbial remediation system to improve the compressive strength and shear strength of soil and reduce the dosage of urea and superphosphate. To this end, the present invention provides a desertification remediation bactericide and its application to meet this need in the art.

[0006] On the one hand, the present invention relates to a desertification remediation bactericide, which is obtained by fermenting Bacillus subtilis, Geotrichum candidum, and Candida utilis to obtain bacterial solutions respectively and then mixing them;

[0007] The Bacillus subtilis is Bacillus subtilis HS0035Kc, and the preservation number is CGMCC NO.9434;

[0008] The Geotrichum candidum is Geotrichum candidum XHS0030B, and the preservation number is CGMCC NO.9435;

[0009] The preservation number of Candida utilis is CICC NO.1314.

[0010] Further, in the desertification remediation bactericide provided by the present invention, in the desertification remediation bactericide, the viable bacteria ratio of Bacillus subtilis, Geotrichum candidum, and Candida utilis is 1-20:1-10:1.

[0011] On the other hand, the present invention relates to the application of the desertification remediation bactericide in the remediation of desertified soil.

[0012] Further, in the application provided by the present invention, it includes: mixing the desertification remediation bactericide and the soil to be remediated and then standing for 2-8 h, then adding urea and superphosphate and mixing, and finally adding water to a water content of 20-50% to obtain a remediation agent; for every 1 m 3 The soil to be remediated is mixed with 300-400 g of the remediation agent to obtain the remediated soil.

[0013] Further, in the application provided by the present invention, by mass ratio, the ratio of the soil to be remediated, the desertification remediation bactericide, urea, and superphosphate is 1:1-2:1-2:0.5-1.5.

[0014] Further, in the application provided by the present invention, the desertification repair microbial agent improves the compressive strength of the soil.

[0015] Further, in the application provided by the present invention, the desertification repair microbial agent improves the shear strength of the soil.

[0016] Compared with the prior art, the technical solution provided by the present invention has at least the following beneficial effects or advantages:

[0017] The present invention constructs a synergistic microbial repair system by using specific Bacillus subtilis, Geotrichum candidum, and Candida utilis. Taking advantage of the strong growth and reproduction ability, fast growth rate, stable genetic characteristics of Bacillus subtilis HS0035Kc and Geotrichum candidum XHS0030B, and their specific effects on soil repair, Candida utilis is further introduced. Compared with using only single microbial agents or dual microbial agents for desertified soil repair, the repair rate is greatly improved, and the compressive strength and shear strength of the soil can be effectively improved. Specific Embodiments

[0018] Next, the technical solution of the present invention will be described in conjunction with the embodiments. However, the present invention is not limited to the following embodiments. The experimental methods and detection methods described in each embodiment are conventional methods unless otherwise specified; the reagents and materials described are commercially available unless otherwise specified. The %, as used in the following embodiments, is the mass percentage unless otherwise specified. The ratios described in the following embodiments are mass ratios unless otherwise specified.

[0019] Bacillus subtilis HS0035Kc and Geotrichum candidum XHS0030B are provided by Xinjiang Huisen Biotechnology Co., Ltd., and Candida utilis (CICC NO.1314) is provided by the Institute of Microbiology, Chinese Academy of Sciences.

[0020] Example 1

[0021] This example provides a preparation test for the Bacillus subtilis HS0035Kc bacterial solution.

[0022] (1) Inoculation: Prepare a solid medium for Bacillus Subtilis XHS0035Kc CGMCC NO.9434. After sterilization, strictly perform aseptic operation, transfer from the slant to the plate, and culture at 28°C for 3 days.

[0023] (2) Primary culture: Pick a single colony from the solid medium in step (1) and transfer it to a 50 mL conical flask containing liquid medium. Perform aseptic operation and culture at 28°C and 120 r / min for 24 h to 48 h.

[0024] (3) Secondary fermentation: Inoculate the primary cultured strain obtained in the above step into a 500 mL conical flask containing liquid medium, and perform aseptic operation. Incubate at 28 °C and 120 r / min for 24 h to 48 h.

[0025] (4) Quantification: Dilute the culture product with liquid medium to OD 600 = 1 to obtain the Bacillus subtilis HS0035Kc bacterial liquid for use.

[0026] The solid medium used for inoculation and primary culture is tryptone soy agar solid medium, which is prepared as follows: 15 g of tryptone, 5 g of soy peptone, 5 g of NaCl, 20 g of agar, make up the volume to 1000 mL with water, adjust the pH to 7.2, dispense, and sterilize to obtain the tryptone soy agar solid medium.

[0027] The liquid medium used for primary culture and secondary fermentation is tryptone soy agar liquid medium, which is prepared as follows: 15 g of tryptone, 5 g of soy peptone, 5 g of NaCl, make up the volume to 1000 mL with water, adjust the pH to 7.2, dispense, and sterilize to obtain the tryptone soy agar liquid medium.

[0028] Example 2

[0029] This example provides a preparation test for the Geotrichum candidum XHS0030B bacterial liquid.

[0030] (1) Inoculation: Prepare the solid medium of Geotrichum candidum XHS0030B. After sterilization, perform strict aseptic operation, transfer from the slant to the plate, and incubate at 28 °C for 3 days.

[0031] (2) Primary culture: Pick a single colony from the solid medium in step (1) and transfer it to a 50 mL conical flask containing liquid medium. Perform aseptic operation and incubate at 25 °C and 120 r / min for 24 h to 48 h.

[0032] (3) Secondary fermentation: Inoculate the primary cultured strain into a 500 mL conical flask containing liquid medium, and perform aseptic operation. Incubate at 25 °C and 120 r / min for 36 h to 72 h.

[0033] (4) Quantification: Dilute the culture product with liquid medium to OD 600 = 1 to obtain the Geotrichum candidum XHS0030B bacterial liquid for use.

[0034] The solid medium used for inoculation and primary culture is potato solid medium, which is prepared as follows: Wash and peel the potatoes, take 200 g and cut into small pieces, add 1000 mL of water, boil for half an hour, and make up the water. Add 10 g of agar to the filtrate, boil and dissolve, then add 20 g of sucrose, make up the water, dispense, and sterilize to obtain the potato solid medium.

[0035] The liquid medium used for primary culture and secondary fermentation is potato liquid medium, which is prepared as follows: Wash and peel the potatoes, take 200 g and cut them into small pieces, add 1000 mL of water, boil for half an hour, and then make up the water. After adding 20 g of sucrose to the filtrate and boiling until dissolved, make up the water, dispense, and sterilize to obtain the potato liquid medium.

[0036] Example 3

[0037] This example provides a preparation test of Candida utilis liquid.

[0038] (1) Inoculation: Prepare a solid medium of Candida utilis (CICC NO.1314), and after sterilization, strictly perform aseptic operation, transfer from the slant to the plate, and culture at 28 °C for 3 days.

[0039] (2) Primary culture: Pick a single colony from the solid medium in step (1) and transfer it to a 50 mL conical flask containing the liquid medium, perform aseptic operation, and culture at 28 °C and 120 r / min for 24 h to 48 h.

[0040] (3) Secondary fermentation: Inoculate the primary culture strain into a 500 mL conical flask containing the liquid medium, perform aseptic operation, and culture at 28 °C and 120 r / min for 36 h to 72 h.

[0041] (4) Quantification: Dilute the culture product with the liquid medium to OD 600 = 1 to obtain the Candida utilis liquid for use.

[0042] The solid medium used for inoculation and primary culture is malt extract solid medium, which is prepared as follows: Take a number of barley malts, crush them, add 4 times the weight of the malt in water, stir evenly, saccharify in an incubator at 60 °C for 4 h, take out and filter to obtain malt extract; after measuring the volume and concentration (Bé degrees of 5 °) of this malt extract, add 15 g of agar to every 1000 mL of malt extract, boil until dissolved, dispense, and sterilize to obtain the malt extract solid medium.

[0043] The liquid medium used for primary culture and secondary fermentation is malt extract liquid medium, which is prepared as follows: Take a number of barley malts, crush them, add 4 times the weight of the malt in water, stir evenly, saccharify in an incubator at 60 °C for 4 h, take out and filter to obtain malt extract; after measuring the volume and concentration (Bé degrees of 5 °) of this malt extract, dispense, and sterilize to obtain the malt extract liquid medium.

[0044] Example 4

[0045] This example provides a desertified soil remediation test.

[0046] The desertified soil used in the experiment was collected from the sandy land of Naiman Banner, Horqin, and used as the soil to be repaired. The Bacillus subtilis HS0035Kc bacterial solution, Geotrichum candidum XHS0030B bacterial solution, and Candida utilis bacterial solution used in the following experiments were provided by Examples 1 to 3.

[0047] Experimental group 1: Take the Bacillus subtilis HS0035Kc bacterial solution, Geotrichum candidum XHS0030B bacterial solution, and Candida utilis bacterial solution and mix them as a desertification repair bactericide. Mix the desertification repair bactericide and the soil to be repaired evenly, let it stand at room temperature (28 °C) indoors for 2 h, then add urea and superphosphate and mix evenly, and add water to adjust the water content to 20% to obtain a repair agent; in the preparation of the repair agent, the volume ratio of the Bacillus subtilis HS0035Kc bacterial solution, Geotrichum candidum XHS0030B bacterial solution, and Candida utilis bacterial solution in the desertification repair bactericide is 1:1:1, and the mass ratio of the soil to be repaired, the desertification repair bactericide, urea, and superphosphate is 1:1:1:0.5; for every 1 m 3 Mix 300 g of the repair agent with the soil to be repaired to obtain the repaired soil.

[0048] Experimental group 2: Take the Bacillus subtilis HS0035Kc bacterial solution, Geotrichum candidum XHS0030B bacterial solution, and Candida utilis bacterial solution and mix them as a desertification repair bactericide. Mix the desertification repair bactericide and the soil to be repaired evenly, let it stand at room temperature (28 °C) indoors for 6 h, then add urea and superphosphate and mix evenly, and add water to adjust the water content to 30% to obtain a repair agent; in the preparation of the repair agent, the volume ratio of the Bacillus subtilis HS0035Kc bacterial solution, Geotrichum candidum XHS0030B bacterial solution, and Candida utilis bacterial solution in the desertification repair bactericide is 10:5:1, and the mass ratio of the soil to be repaired, the desertification repair bactericide, urea, and superphosphate is 1:1.5:1.5:1; for every 1 m 3 Mix 350 g of the repair agent with the soil to be repaired to obtain the repaired soil.

[0049] Experimental group 3: Take the Bacillus subtilis HS0035Kc bacterial solution, Geotrichum candidum XHS0030B bacterial solution, and Candida utilis bacterial solution and mix them as a desertification repair bactericide. Mix the desertification repair bactericide and the soil to be repaired evenly, let it stand at room temperature (28 °C) indoors for 8 h, then add urea and superphosphate and mix evenly, and add water to adjust the water content to 50% to obtain a repair agent; in the preparation of the repair agent, the volume ratio of the Bacillus subtilis HS0035Kc bacterial solution, Geotrichum candidum XHS0030B bacterial solution, and Candida utilis bacterial solution in the desertification repair bactericide is 20:10:1, and the mass ratio of the soil to be repaired, the desertification repair bactericide, urea, and superphosphate is 1:2:2:1.5; for every 1 m 3 Mix 400 g of the repair agent with the soil to be repaired to obtain the repaired soil.

[0050] Positive control group 1: This control group is the same as experimental group 3, except that the desertification restoration microbial agent does not contain Geotrichum candidum XHS0030B bacterial liquid and Candida utilis bacterial liquid.

[0051] Positive control group 2: This control group is the same as experimental group 3, except that the desertification restoration microbial agent does not contain Candida utilis bacterial liquid.

[0052] Positive control group 3: This control group is the same as experimental group 3, except that the desertification restoration microbial agent does not contain Geotrichum candidum XHS0030B bacterial liquid.

[0053] Positive control group 4: This control group is the same as experimental group 3, except that the desertification restoration microbial agent does not contain Bacillus subtilis HS0035Kc bacterial liquid.

[0054] Blank control group: No additional treatment is given to this control group.

[0055] The compressive strength and shear strength of the restored soil were measured at 48h, 96h, 144h, and 192h after restoration, respectively. The test results are shown in Table 1.

[0056] Table 1: Strength of the Restored Soil

[0057]

[0058] As can be seen from Table 1, in this experiment, by comparing different desertified soil remediation schemes, the remediation effect of a desertification remediation bactericide composed of Bacillus subtilis HS0035Kc bacterial liquid, Geotrichum candidum XHS0030B bacterial liquid and Candida utilis bacterial liquid on desertified soil was explored. The experimental results showed that compared with the blank control group, the compressive strength and shear strength of the soil after remediation in all experimental groups and the positive control group were improved, indicating that the addition of the bactericide had a positive effect on soil remediation. Among the experimental groups, with the extension of the standing time and the increase of the bactericide ratio, the water content and dosage of the remediation agent, the improvement amplitude of the soil strength was different, but generally showed an increasing trend with time. The compressive strength and shear strength between Experimental Groups 1-3 were better than those of the positive control group. In the positive control group, the combination without Geotrichum candidum XHS0030B bacterial liquid and Candida utilis bacterial liquid performed the worst, indicating that these two bacterial liquids played an important role in enhancing the efficiency of soil remediation; the combination without Bacillus subtilis HS0035Kc bacterial liquid performed better in the initial stage, but the strength growth was slow over time, indicating that Bacillus subtilis HS0035Kc bacterial liquid was crucial for the long-term remediation effect of the soil. To sum up, a desertification remediation bactericide composed of Bacillus subtilis HS0035Kc bacterial liquid, Geotrichum candidum XHS0030B bacterial liquid and Candida utilis bacterial liquid mixed in a certain proportion can effectively improve the compressive strength and shear strength of desertified soil under the conditions of appropriate standing time, water content and dosage of the remediation agent, providing an effective biological remediation method for desertified soil remediation.

[0059] As described above, the basic principles, main features and advantages of the present invention are preferably described. The above embodiments and the description are only for describing the preferred embodiments of the present invention. The present invention is not limited by the above embodiments. Without departing from the spirit and scope of the present invention, various changes and improvements made by those of ordinary skill in the art to the technical solution of the present invention shall fall within the protection scope determined by the present invention.

Claims

1. A desertification remediation bacterial agent, characterized in that: The desertification restoration bacterial agent is obtained by fermenting Bacillus subtilis, Geotrichum candidum and Candida utilis to obtain bacterial liquids respectively, and then mixing them; The Bacillus subtilis is Bacillus subtilis HS0035Kc, with a deposit number of CGMCCNO.9434; The candidum is candidum XHS0030B, and its deposit number is CGMCC NO.9435; The deposit number of the Candida utilis is CICC NO.1314.

2. The desertification remediation bacterial agent according to claim 1, characterized in that: In the desertification remediation bacterial agent, the ratio of live bacteria of Bacillus subtilis, Geotrichum candidum and Candida utilis is 1-20:1-10:

1.

3. Use of the desertification remediation bacterial agent according to claim 1 or 2 in desertification soil remediation.

4. The use according to claim 3, characterized in that: include: The desertification restoration bacteria agent and the soil to be restored are mixed and left to stand for 2 to 8 hours, and then urea and superphosphate are added and mixed, and finally water is added to obtain a restoration agent with a water content of 20 to 50%. 3 The soil to be repaired is mixed with 300-400 g of the repair agent to obtain repaired soil.

5. The use according to claim 4, characterized in that: Calculated by mass ratio, the ratio of the soil to be repaired, the desertification repair bacterial agent, urea and superphosphate is 1:1-2:1-2:0.5-1.

5.

6. The use according to claim 3, characterized in that: The desertification remediation bacterial agent improves the compressive strength of the soil.

7. The use according to claim 3, characterized in that: The desertification remediation bacterial agent improves the shear strength of the soil.

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