Bacillus orientalis, microbial preparation and method for increasing content of calcium carbonate in granite residual soil

Abstract

The invention belongs to the technical field of microbial soil fixation, and particularly relates to bacillus orientalis, a microbial preparation and a method for increasing the content of calcium carbonate in granite residual soil. According to the invention, microorganisms capable of producing calcium carbonate are screened from granite residual soil, unknown bacteria are subjected to molecular identification, an acid-resistant strain is obtained through screening, and the content of calcium carbonate in the granite residual soil can be effectively increased.

Description

Technical Field

[0001] This application belongs to the technical field of microbial soil stabilization, and specifically relates to a Bacillus toyonensis, a microbial agent, and a method for increasing the calcium carbonate content in granite residual soil. Background Art

[0002] Granite residual soil is a type of soil formed after the weathering of granite, mainly composed of sand grains, silt, and clay. The particle size and proportion are affected by the degree of weathering. This soil is usually weakly acidic to neutral, and its mineral components mainly include quartz, feldspar, and mica. The nutrient content is relatively low, the water-holding capacity and air permeability are medium, and it has good drainage. Due to its particle composition and pore structure, granite residual soil may settle under load and is prone to geological disasters such as landslides under strong rainfall, which affects engineering construction.

[0003] Microbial-induced calcium carbonate precipitation technology (MICP) is a method that uses the metabolic activities of specific microorganisms to promote calcium carbonate precipitation under suitable environmental conditions. When microorganisms consume organic matter during their metabolism, ammonia is released and the carbon dioxide concentration in the environment is directly increased, resulting in an increase in the surrounding pH value, thereby forming calcium carbonate precipitation. This technology has broad application prospects, such as in the warranty and repair of building materials, soil improvement, water purification, and heavy metal removal, and has attracted much attention due to its environmental protection and economy.

[0004] However, currently, the application scenarios of this technology are mostly in cement and concrete. The optimal growth environment of the microorganisms used is mostly alkaline with a pH range of 8 - 9.5. At the same time, during the process of urease hydrolysis inducing calcium carbonate, as ammonia is continuously generated, the concentration of hydroxide ions increases, resulting in an increase in the pH value of the solution.

[0005] In addition, although there are existing records on the related applications of Bacillus in the field of soil stabilization, the recorded strains do not have acid tolerance characteristics. Therefore, it is particularly important to further study Bacillus and its applications in the field of granite soil stabilization. Summary of the Invention

[0006] Based on this, an embodiment of this application provides a Bacillus toyonensis ZQ-7-1 and a method for increasing the calcium carbonate content in granite residual soil.

[0007] This application provides a Bacillus toyonensis ZQ-7-1 on the one hand. It was deposited at the China General Microbiological Culture Collection Center on February 27, 2025, with the deposit number CGMCC No. 33676, address: No. 3, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences.

[0008] On the other hand, the present application provides a microbial agent, including the above-mentioned Bacillus toyonensis ZQ-7-1.

[0009] On the other hand, the present application provides a method for increasing the calcium carbonate content in granite eluvial soil, including:

[0010] The step of applying the above-mentioned Bacillus toyonensis ZQ-7-1 into granite eluvial soil.

[0011] In some embodiments, it includes: pouring the bacterial solution of Bacillus toyonensis ZQ-7-1 and a reaction solution into the granite eluvial soil for reaction.

[0012] In some embodiments, the reaction solution includes urea and a calcium salt.

[0013] In some embodiments, the concentration of urea in the reaction solution is 0.2 mol / L to 1.5 mol / L; the concentration of the calcium salt in the reaction solution is 0.2 mol / L to 1.5 mol / L.

[0014] In some embodiments, the calcium salt includes one or both of calcium chloride and calcium acetate.

[0015] In some embodiments, the viable count of the bacterial solution of Bacillus toyonensis ZQ-7-1 is 10 9 CFU / mL to 10 12 CFU / mL.

[0016] In some embodiments, the volume ratio of the bacterial solution of Bacillus toyonensis ZQ-7-1 to the mass of the granite eluvial soil is 1 mL:(9 - 20) g.

[0017] In some embodiments, the volume ratio of the bacterial solution of Bacillus toyonensis ZQ-7-1 to the reaction solution is 1:(5 - 25).

[0018] In some embodiments, the reaction duration is 24h to 72h.

[0019] In some embodiments, the bacterial solution of Bacillus toyonensis ZQ-7-1 includes the fermentation broth of Bacillus toyonensis ZQ-7-1.

[0020] In some of these embodiments, the steps for preparing the fermentation broth include: inoculating a single colony of Bacillus toyonensis ZQ-7-1 into a fermentation medium for fermentation.

[0021] In some of these embodiments, the fermentation conditions include: a temperature of 22°C to 42°C and a time of 12 h to 72 h.

[0022] In some of these embodiments, the fermentation medium includes 10 g / L to 20 g / L of peptone, 2 g / L to 8 g / L of beef extract, and 4 g / L to 10 g / L of sodium chloride, with a pH of 5.5 to 7.5.

[0023] In some of these embodiments, the fermentation is carried out under oscillating conditions at a rotational speed of 150 rpm to 250 rpm.

[0024] This application provides a method for screening microorganisms capable of producing calcium carbonate from granite residual soil, molecularly identifying unknown bacteria, screening an acid-tolerant strain, and effectively increasing the calcium carbonate content in granite residual soil. In addition, the microorganisms used in this application are cultures of microorganisms that naturally exist in the environment and do not cause pollution to the environment. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] To more clearly illustrate the technical solutions in the embodiments of this application and more fully understand this application and its beneficial effects, the following will briefly introduce the drawings required for use in the description of the embodiments. Obviously, the drawings in the following description are only some embodiments of this application, and those skilled in the art can obtain other drawings based on these drawings without creative efforts.

[0026] Figure 1 The 16S rRNA phylogenetic tree of Bacillus toyonensis ZQ-7-1 provided in an embodiment of this application;

[0027] Figure 2 The scanning electron microscope photograph of Bacillus toyonensis ZQ-7-1 provided in an embodiment of this application;

[0028] Figure 3 The urease activity of Bacillus toyonensis ZQ-7-1 provided in an embodiment of this application under different pH medium conditions;

[0029] Figure 4The change in pH of granite eluvial soil collected from different regions before and after treatment with the bacterial solution of Bacillus toyonensis ZQ-7-1 and the reaction solution provided by an embodiment of the present application;

[0030] Figure 5 The calcium carbonate content of granite eluvial soil collected from different regions after treatment with the bacterial solution of Bacillus toyonensis ZQ-7-1 and the reaction solution provided by an embodiment of the present application. Specific Embodiments

[0031] The present application will be further described in detail below in conjunction with the embodiments and examples. It should be understood that these embodiments and examples are only used to illustrate the present application and not to limit the scope of the present application. The purpose of providing these embodiments and examples is to make the understanding of the disclosed content of the present application more thorough and comprehensive. It should also be understood that the present application can be implemented in many different forms and is not limited to the embodiments and examples described herein. Those skilled in the art can make various changes or modifications without departing from the connotation of the present application, and the equivalent forms obtained also fall within the protection scope of the present application. In addition, in the following description, a large number of specific details are given to provide a more thorough understanding of the present application. It should be understood that the present application can be implemented without one or more of these details.

[0032] Unless otherwise defined, all technical and scientific terms used in the present application have the same meaning as commonly understood by those skilled in the technical field to which the present application belongs.

[0033] Terms

[0034] Unless otherwise stated or there is a contradiction, the terms or phrases used herein have the following meanings:

[0035] As used herein, the alternative scopes of the terms "and / or", "or / and", and "and / or" include any one of two or more related listed items, as well as any and all combinations of the related listed items. The aforesaid any and all combinations include combinations of any two related listed items, any more related listed items, or all related listed items. It should be noted that when at least three items are connected by at least two conjunctive combinations selected from "and / or", "or / and", and "and / or", it should be understood that in this application, this technical solution undoubtedly includes the technical solution connected by "logical AND", and undoubtedly also includes the technical solution connected by "logical OR". For example, "A and / or B" includes three parallel solutions: A, B, and A + B. Another example, the technical solution of "A, and / or, B, and / or, C, and / or, D" includes any one of A, B, C, and D (that is, the technical solution connected by "logical OR"), and also includes any and all combinations of A, B, C, and D, that is, it includes combinations of any two or any three of A, B, C, and D, and also includes the combination of the four items A, B, C, and D (that is, the technical solution connected by "logical AND").

[0036] In this application, the terms "multiple", "diverse", "multiple times", "multiple elements", etc. refer to a quantity greater than or equal to 2 if not otherwise specified. For example, "one or more" means one or greater than or equal to two.

[0037] As used herein, "its combination", "any combination thereof", "any combination mode thereof", etc. include all suitable combination modes of any two or more than two items in the listed items.

[0038] In this article, the "suitable" in "suitable combination mode", "suitable mode", "any suitable mode", etc. is subject to being able to implement the technical solution of this application, solve the technical problems of this application, and achieve the expected technical effects of this application.

[0039] In this application, terms such as "further", "even further", "especially", etc. are used for descriptive purposes, indicating differences in content, but should not be construed as limiting the protection scope of this application.

[0040] In this application, "optionally", "optional", "optional" mean that it can be either present or absent, that is, it refers to any one of the two parallel solutions of "present" or "absent". If "optional" appears multiple times in a technical solution, without special instructions and without contradictions or mutual restrictions, each "optional" is independent of each other.

[0041] In this application, for the technical features described in an open-ended manner, it includes a closed technical solution composed of the listed features, and also includes an open technical solution containing the listed features.

[0042] In this application, when it comes to numerical intervals (i.e., numerical ranges), unless otherwise specified, the selectable numerical values are considered continuous within the above numerical intervals, and include the two numerical endpoints of the numerical range (i.e., the minimum value and the maximum value), as well as each numerical value between these two numerical endpoints. Unless otherwise specified, when the numerical interval only refers to integers within the numerical interval, it includes the two endpoint integers of the numerical range, as well as each integer between the two endpoints. In this article, it is equivalent to directly listing each integer. For example, when t is an integer selected from 1 to 10, it means that t is any integer selected from the integer group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10. In addition, when multiple ranges are provided to describe features or characteristics, these ranges can be combined. In other words, unless otherwise specified, the ranges disclosed in this article should be understood to include any and all sub-ranges subsumed therein.

[0043] The temperature parameter in this application, unless otherwise specified, allows both constant temperature treatment and variation within a certain temperature range. It should be understood that the constant temperature treatment allows the temperature to fluctuate within the accuracy range controlled by the instrument. Fluctuation within ranges such as ±5°C, ±4°C, ±3°C, ±2°C, ±1°C is allowed.

[0044] In this application, %(w / w) and wt% both represent weight percentage, %(v / v) refers to volume percentage, and %(w / v) refers to mass-volume percentage.

[0045] All documents mentioned in this application are cited as references in this application, just as if each document was cited separately as a reference. Unless it conflicts with the inventive purpose and / or technical solution of this application, the cited documents involved in this application are cited for all contents and all purposes. When this application involves cited documents, the definitions of relevant technical features, terms, nouns, phrases, etc. in the cited documents are also cited together. When this application involves cited documents, the examples and preferred methods of the relevant technical features cited can also be incorporated as references into this application, but only to the extent that this application can be implemented. It should be understood that when the cited content conflicts with the description in this application, this application shall prevail or be amended adaptively according to the description of this application.

[0046] Currently, the optimal growth environment of urease-producing microorganisms is mostly in the range of slightly alkaline pH 8 - 9.5. At the same time, during the process of urease hydrolysis inducing calcium carbonate, as ammonia is continuously generated and the concentration of hydroxide ions increases, the pH value of the solution rises. However, granite residual soil is acidic. The alkaline substances (such as NH3) produced by the hydrolysis of existing urease-producing microorganisms will gradually change the local pH, which may inhibit the microbial activity or damage the original properties of the soil, and the compatibility with granite residual soil is not high.

[0047] Based on this, on the one hand, the present application provides a Bacillus toyonensis ZQ-7-1, which was deposited at the General Microbiology Center of the China Committee for Culture Collection of Microorganisms on February 27, 2025, with the deposit number CGMCC No. 33676, and the address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences.

[0048] On the other hand, the present application provides a method for increasing the calcium carbonate content in granite residual soil, including:

[0049] The step of pouring Bacillus toyonensis ZQ-7-1 into granite residual soil.

[0050] The Bacillus toyonensis ZQ-7-1 provided by the present application has acid tolerance characteristics, is more suitable for production in granite residual soil, and can effectively promote the calcium carbonate content in granite residual soil.

[0051] On the other hand, the present application provides a microbial preparation, including the Bacillus toyonensis ZQ-7-1 described above.

[0052] In some of these embodiments, it includes: applying the bacterial solution and reaction solution of Bacillus toyonensis ZQ-7-1 to granite residual soil and carrying out a reaction.

[0053] In some of these embodiments, the reaction solution includes urea and a calcium salt.

[0054] Under the action of urease produced by microorganisms, urea is decomposed into ammonium ions and carbon dioxide, and at the same time, the pH around the bacteria increases. In the presence of calcium ions, water-insoluble calcium carbonate salts are generated, further binding the surrounding sand grains together.

[0055] In some of these embodiments, the concentration of urea in the reaction solution is 0.2 mol / L to 1.5 mol / L; the concentration of the calcium salt in the reaction solution is 0.2 mol / L to 1.5 mol / L.

[0056] For example, the concentration of urea is 0.2 mol / L, 0.3 mol / L, 0.4 mol / L, 0.5 mol / L, 0.6 mol / L, 0.7 mol / L, 0.8 mol / L, 0.9 mol / L, 1.0 mol / L, 1.1 mol / L, 1.2 mol / L, 1.3 mol / L, 1.4 mol / L or 1.5 mol / L and any value in between.

[0057] For example, the concentration of the calcium salt is 0.2 mol / L, 0.3 mol / L, 0.4 mol / L, 0.5 mol / L, 0.6 mol / L, 0.7 mol / L, 0.8 mol / L, 0.9 mol / L, 1.0 mol / L, 1.1 mol / L, 1.2 mol / L, 1.3 mol / L, 1.4 mol / L or 1.5 mol / L, and any value in between.

[0058] In some of these embodiments, the calcium salt includes one or both of calcium chloride and calcium acetate.

[0059] In some of these embodiments, the viable count of the bacterial solution of Bacillus toyonensis ZQ-7-1 is 10 9 CFU / mL to 10 12 CFU / mL. For example, 10 9 CFU / mL, 10 10 CFU / mL, 10 11 CFU / mL or 10 12 CFU / mL, and any value in between. In some of these embodiments, the volume ratio of the bacterial solution of Bacillus toyonensis ZQ-7-1 to the mass of the granite residual soil is 1 mL:(9 - 20) g. For example, the volume ratio of the bacterial solution to the mass of the granite residual soil is 1:9, 1:10, 1:11, 1:12, 1:13, 1:14, 1:15, 1:16, 1:17, 1:18, 1:19 or 1:20, and any value in between.

[0060] In some of these embodiments, the volume ratio of the bacterial solution of Bacillus toyonensis ZQ-7-1 to the reaction solution is 1:(5 - 25). For example, the volume ratio of the bacterial solution to the reaction solution is 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 1:11, 1:12, 1:13, 1:14, 1:15, 1:16, 1:17, 1:18, 1:19, 1:20, 1:21, 1:22, 1:23, 1:24 or 1:25, and any value in between.

[0061] In some of these embodiments, the duration of the reaction is 24h - 72h. For example, the reaction duration is 24h, 26h, 28h, 30h, 32h, 34h, 36h, 38h, 40h, 42h, 44h, 46h, 48h, 50h, 52h, 54h, 56h, 58h, 60h, 62h, 64h, 66h, 68h, 70h or 72h, and any value in between.

[0062] In some of these embodiments, the bacterial liquid includes the fermentation broth of Bacillus toyonensis ZQ-7-1.

[0063] In some of these embodiments, the steps for preparing the fermentation broth include:

[0064] Inoculating a single colony of Bacillus toyonensis ZQ-7-1 into a fermentation medium for fermentation.

[0065] In some of these embodiments, the conditions for fermentation include: the temperature is 22°C to 42°C, and the time is 12 h to 72 h.

[0066] For example, the temperature is 22°C, 23°C, 24°C, 25°C, 26°C, 27°C, 28°C, 29°C, 30°C, 31°C, 32°C, 33°C, 34°C, 35°C, 36°C, 37°C, 38°C, 39°C, 40°C, 41°C or 42°C and any value in between.

[0067] For example, the fermentation time is 12 h, 14 h, 16 h, 18 h, 20 h, 22 h, 24 h, 26 h, 28 h, 30 h, 32 h, 34 h, 36 h, 38 h, 40 h, 42 h, 44 h, 46 h, 48 h, 50 h, 52 h, 54 h, 56 h, 58 h, 60 h, 62 h, 64 h, 66 h, 68 h, 70 h or 72 h and any value in between.

[0068] In some of these embodiments, the fermentation medium includes 10 g / L to 20 g / L of peptone, 2 g / L to 8 g / L of beef extract, and 4 g / L to 10 g / L of sodium chloride, with a pH of 5.5 to 7.5.

[0069] For example, the concentration of peptone is 10 g / L, 11 g / L, 12 g / L, 13 g / L, 14 g / L, 15 g / L, 16 g / L, 17 g / L, 18 g / L, 19 g / L or 20 g / L and any value in between.

[0070] For example, the concentration of beef extract is 2 g / L, 3 g / L, 4 g / L, 5 g / L, 6 g / L, 7 g / L or 8 g / L and any value in between.

[0071] In some of these embodiments, the conditions for fermentation further include shake culture.

[0072] In some of these embodiments, the rotation speed of the shake culture is 150 rpm to 250 rpm.

[0073] For example, the rotational speeds are 150 rpm, 155 rpm, 160 rpm, 165 rpm, 170 rpm, 175 rpm, 180 rpm, 185 rpm, 190 rpm, 195 rpm, 200 rpm, 205 rpm, 210 rpm, 215 rpm, 220 rpm, 225 rpm, 230 rpm, 235 rpm, 240 rpm, 245 rpm or 250 rpm, as well as any value in between.

[0074] The embodiments of the present application will be described in detail below in conjunction with examples. It should be understood that these examples are only used to illustrate the present application and not to limit the scope of the present application. For the experimental methods without specific conditions in the following examples, the guidance given in the present application shall be preferentially referred to, and it can also be carried out according to the experimental manuals or conventional conditions in the art, or according to the conditions recommended by the manufacturer, or by referring to the experimental methods known in the art.

[0075] In the following specific embodiments, for the measurement parameters of raw material components, if there is no special description, there may be slight deviations within the weighing accuracy range. For temperature and time parameters, acceptable deviations caused by instrument test accuracy or operation accuracy are allowed.

[0076] It should be understood that in various embodiments of the present application, the magnitudes of the serial numbers of the above processes do not mean the sequence of execution. The execution sequence of each process should be determined according to its function and internal logic, and should not constitute any limitation to the implementation process of the embodiments of the present application.

[0077] Example 1

[0078] I. Isolation and screening of urease-producing and acid-tolerant strains

[0079] 1. Strain collection location: The urease-producing strains provided in this application were isolated from Huaiji County, Zhaoqing City, Guangdong Province.

[0080] 2. Screening process

[0081] In this example, the test samples were first enriched and cultured at 37°C under the condition of 3 mol / L high-concentration urea for 24 h, and then various microbial vegetative cells that could not tolerate and utilize high-concentration urea were killed.

[0082] In addition, since bacteria have urease, which can decompose urea to produce a large amount of ammonia, making the culture medium alkaline and showing red. Taking advantage of this feature, the present application then gradient-diluted the treated culture solution, spread it on a urease screening culture plate, cultured it at 37°C, picked the strains that turned the culture medium red, and isolated single colonies by streaking. The obtained microorganisms were urease-producing microorganisms.

[0083] Since this embodiment needs to take into account the acid tolerance of the strain, when preparing the urease screening plate, the concentration of urea added must be strictly controlled. After filtering and sterilizing the urea solution, it is gradually added to the urea agar basal medium until the color just turns red, and then the addition is stopped. At this time, the pH of the medium is detected to be 7.2.

[0084] Next, the acid tolerance of the selected strains is screened. The strains with urease-producing activity selected in the previous round are inoculated into the fermentation medium with a pH of 5.5 and cultured at 37 °C for 24 hours.

[0085] Among them, the components of the fermentation medium are peptone 10 g / L, beef extract 3 g / L, sodium chloride 5 g / L, and the pH is 5.5.

[0086] 3. Screening Results

[0087] Select the strain with the highest monomeric enzyme activity and identify it by the 16S rDNA method. It is named Bacillus toyonensis ZQ-7-1, and the results are as Figure 1 shown.

[0088] Furthermore, its morphology is observed by scanning electron microscopy, and the results are as Figure 2 shown.

[0089] The 16s DNA sequence of Bacillus toyonensis ZQ-7-1 is as shown in SEQ ID NO.1:

[0090]

[0091] II. Cultivation of Bacillus toyonensis ZQ-7-1

[0092] Prepare a fermentation medium, where the components of the fermentation medium are 10 g / L peptone, 3 g / L beef extract, 5 g / L sodium chloride, and the pH is 5.5.

[0093] Fill 100 mL of the fermentation medium into a 500 mL culture flask for sterilization, inoculate Bacillus toyonensis ZQ-7-1 into the fermentation medium, and culture it with a shaker at 37 °C. The rotation speed of the shaker is 200 rpm.

[0094] The results show that Bacillus toyonensis ZQ-7-1 has certain acid tolerance activity. In the range of pH 4 - 7, it can survive and grow and has a relatively high urease activity, as Figure 3 shown.

[0095] Example 2

[0096] This example provides a method for increasing the calcium carbonate content in granite residual soil.

[0097] I. Induction process

[0098] 1. Ferment and culture Bacillus toyonensis ZQ-7-1 in Example 1 in a fermentation medium at 37 °C for 60 hours to prepare a bacterial solution.

[0099] Among them, the components of the fermentation medium are 10 g / L peptone, 3 g / L beef extract, 5 g / L sodium chloride, and the pH is 5.5.

[0100] 2. Select granite residual soil samples

[0101] The used granite residual soil is taken from three transmission tower bases in Huaiji County, Zhaoqing City, Guangdong Province (the samples are represented by ZQ-3; ZQ-7; ZQ-8 respectively), Fogang County, Qingyuan City (the samples are represented by FG-5; FG-7 respectively), and Wangzishan Forest Park, Huadu District, Guangzhou City (the samples are represented by GZ-2; GZ-5; GZ-7; GZ-9; GZ-11 respectively).

[0102] 3. Prepare a reaction solution

[0103] The reaction solution for inducing mineralization to generate calcium carbonate contains 1 mol / L urea and 1 mol / L calcium chloride.

[0104] 4. Reaction process

[0105] Mix the bacterial liquid cultured in Step 1 with the above 9 types of granite residual soils respectively, with the volume ratio of the bacterial liquid to the soil being 1:15; then add the reaction solution, with the volume ratio of the bacterial liquid to the reaction solution being 1:10. Stir the bacterial liquid, the reaction solution and the granite residual soil evenly, and after standing for 72 hours, measure the pH and calcium carbonate content of the soil.

[0106] II. Determination of the pH of Granite Residual Soils Collected from Different Regions

[0107] Put about 10 g of soil sample into 50 mL of ionized water, stir evenly and then stand for 24 hours. Decant the supernatant to obtain the soil extract. Insert an electronic pH meter into the extract, and record the pH value displayed on the screen after the reading stabilizes.

[0108] The results are as Figure 4 shown. The granite residual soils collected from different locations in three regions are all slightly acidic, with the pH ranging from 5.7 to 6.8.

[0109] After being treated with the bacterial liquid of Bacillus toyonensis ZQ-7-1 and the reaction solution, due to the action of microbial urease, urea in the reaction solution is hydrolyzed to produce ammonium ions, and the pH of the soil samples all increases to a certain extent, but basically remains within the neutral range, and has no significant impact on the soil acid-base environment of the granite residual soil.

[0110] III. Determination of the Calcium Carbonate Content in Granite Residual Soils Collected from Different Regions

[0111] The calcium carbonate in the granite residual soil reacts with a certain amount of excessive hydrochloric acid standard solution, and the remaining acid is titrated with sodium hydroxide standard solution. Calculate the calcium carbonate content in the soil based on the amount of the hydrochloric acid standard solution consumed.

[0112] The specific calculation process includes: Weigh 5.00 g of soil sample and place it in a 100 mL beaker. Add 20.00 mL of 0.5 mol / L hydrochloric acid standard solution, cover it with a watch glass, stir with a glass rod to expel the generated carbon dioxide. After cooling, transfer the solution to a 100 mL volumetric flask, rinse the soil sample and the beaker with water, and then dilute to the scale with water and shake well. Pipette 50.00 mL of the clearer solution into a 150 mL conical flask, add a few drops of phenolphthalein indicator, and titrate the remaining hydrochloric acid with 0.25 mol / L sodium hydroxide until the solution turns slightly pink and does not fade within 30 s.

[0113] The results are as Figure 5As shown, after treatment with the bacterial solution of Bacillus toyonensis ZQ-7-1 and the reaction solution, under the action of microbial urease, urea in the reaction solution is hydrolyzed to produce ammonium ions, and carbonate ions are produced by hydrolysis and react with calcium ions in the reaction solution to produce calcium carbonate. Calcium carbonate is generated in different samples collected from different regions.

[0114] The generation of calcium carbonate induced by Bacillus toyonensis ZQ-7-1 indicates its potential to improve the engineering properties of granite residual soil.

[0115] The above-described embodiments merely represent several implementation manners of the present application, which are convenient for understanding the technical solutions of the present application specifically and in detail, but should not be construed as a limitation on the scope of patent protection of the application. It should be noted that for those of ordinary skill in the art, without departing from the concept of the present application, several modifications and improvements can still be made, and these all belong to the protection scope of the present application. In addition, it should be understood that after reading the above teachings of the present application, those skilled in the art can make various changes or modifications to the present application, and the equivalent forms obtained also fall within the protection scope of the present application. It should also be understood that the technical solutions obtained by those skilled in the art through logical analysis, reasoning or limited experiments based on the technical solutions provided in the present application are all within the protection scope of the appended claims of the present application. Therefore, the protection scope of the patent of the present application shall be subject to the content of the appended claims, and the specification can be used to interpret the content of the claims.

Claims

1. A Bacillus toyonensis ZQ-7-1, which was deposited on February 27, 2025 at the General Microbiology Center of the China Committee for Culture Collection of Microorganisms, with the deposit number CGMCC No. 33676, address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences.

2. A microbial preparation, characterized in that, Including the Bacillus toyonensis ZQ-7-1 described in claim 1.

3. A method for increasing the calcium carbonate content in granite residual soil, characterized in that, Including: The step of applying the Bacillus toyonensis ZQ-7-1 described in claim 1 or the microbial preparation described in claim 2 into the granite residual soil.

4. The method for increasing the calcium carbonate content in granite residual soil according to claim 3, wherein Including: Pouring the bacterial solution of the Bacillus toyonensis ZQ-7-1 and the reaction solution into the granite residual soil for reaction.

5. The method for increasing the calcium carbonate content in granite residual soil according to claim 4, characterized in that The reaction solution includes urea and calcium salt; The calcium salt includes one or both of calcium chloride and calcium acetate; The concentration of the urea in the reaction solution is 0.2 mol / L to 1.5 mol / L; the concentration of the calcium salt in the reaction solution is 0.2 mol / L to 1.5 mol / L.

6. The method for increasing the calcium carbonate content in granite residual soil according to claim 4, wherein The viable count of the Bacillus toyonensis ZQ-7-1 bacterial liquid is 10 9 CFU / mL to 10 12 CFU / mL; and / or, The mass ratio of the volume of the bacterial solution of the Bacillus toyonensis ZQ-7-1 to the mass of the granite residual soil is 1 mL:(9 - 20) g; and / or, The volume ratio of the bacterial solution of the Bacillus toyonensis ZQ-7-1 to the reaction solution is 1:(5 - 25).

7. The method for increasing the calcium carbonate content in granite residual soil according to claim 4, characterized in that, The duration of the reaction is 24h to 72h.

8. The method for increasing the calcium carbonate content in granite residual soil according to any one of claims 3 to 7, characterized in that, The bacterial solution of the Bacillus toyonensis ZQ-7-1 includes the fermentation broth of the Bacillus toyonensis ZQ-7-1; The preparation steps of the fermentation broth include: inoculating the Bacillus toyonensis ZQ-7-1 into the fermentation medium for fermentation to prepare the fermentation broth; The conditions of the fermentation include: the temperature is 22 °C to 42 °C, and the time is 12 h to 72 h.

9. The method for increasing the calcium carbonate content in granite residual soil according to claim 8, characterized in that, The fermentation medium includes 10 g / L to 20 g / L of peptone, 2 g / L to 8 g / L of beef extract, and 4 g / L to 10 g / L of sodium chloride, with a pH of 5.5 to 7.

5.

10. The method for increasing the calcium carbonate content in granite residual soil according to claim 8, characterized in that, The fermentation is carried out under the shaking condition with a rotation speed of 150 rpm to 250 rpm.

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