Application of penicillium kluyverovii in improvement of saline-alkali soil
By preparing a compound microbial agent containing extracts of Penicillium clarkii, Sphingomonas sphingosine monocytogenes, and Artemisia annua, the problem of unsatisfactory results of Penicillium clarkii in the improvement of saline-alkali soil was solved, and the improvement of saline-alkali soil and the increase of crop yield were achieved.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-12-31
- Publication Date
- 2026-03-17
Smart Images
Figure CN121674069A_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The application provides application of paecilomyces fumosaeruleus in improved saline-alkali soil and relates to the technical field of soil improvement. BACKGROUND
[0002] In the vast world of microorganisms, Penicillium, as a common fungus, has long been closely watched by scientists due to its unique biological activity and extensive ecological distribution. Among them, P. chrysogenum, as a kind of Penicillium, has attracted much attention because of its potential agricultural application value. However, despite the potential of Penicillium and its species such as P. chrysogenum in agricultural production, especially in the attempt to improve saline-alkali soil, the current technology is not mature and the improvement effect is not ideal. Penicillium is a kind of fungus widely distributed in soil, air and plant surface, which participates in the decomposition of organic matter and nutrient cycling in soil by producing various enzymes and organic acids. These characteristics make Penicillium have potential application value in agricultural production. P. chrysogenum, as a kind of Penicillium, has stronger environmental adaptability and biological activity. It can produce various secondary metabolites, including antibiotics, antibacterial peptides, etc., which have significant inhibitory effect on pathogenic microorganisms in soil, thereby helping to reduce the occurrence of plant diseases. In agricultural production, the application of Penicillium and its species such as P. chrysogenum mainly reflects in two aspects of soil improvement and plant disease prevention and control. By inoculating Penicillium strains, the soil microecological environment can be improved, the decomposition of soil organic matter and the release of nutrients can be promoted, thereby improving soil fertility and crop yield. At the same time, the secondary metabolites produced by Penicillium can also inhibit pathogenic bacteria in soil, reduce the occurrence of plant diseases, and improve the stress resistance of crops. However, the application of Penicillium and its species such as P. chrysogenum in saline-alkali soil improvement is a challenging task. Saline-alkali soil refers to the soil type that contains too much soluble salt, leading to soil salinization, decreased fertility, and hindered crop growth. The improvement of saline-alkali soil needs to consider the improvement of soil physical and chemical properties, the adjustment of soil microbial community, and the improvement of crop salt tolerance, etc. Although Penicillium and its species such as P. chrysogenum have certain potential in soil improvement, the current technology is not mature and the improvement effect is not ideal. The reasons are as follows: first, the high salt content of saline-alkali soil greatly limits the growth and reproduction of Penicillium. High salt environment can damage the cell structure of Penicillium, inhibit its metabolic activity, and thus reduce its biological activity and improvement effect. Second, the imbalance of pH in saline-alkali soil makes it difficult for Penicillium to colonize and reproduce in soil. Saline-alkali soil is usually alkaline or strongly alkaline, while Penicillium grows better in slightly acidic or neutral soil. The mismatch of pH will lead to the decrease of survival rate of Penicillium in saline-alkali soil, and the improvement effect is naturally not ideal. Third, the complex microbial community structure in saline-alkali soil makes it difficult for Penicillium to form a dominant group. The saline-alkali soil contains a variety of microorganisms, and there is a complex interaction between them. Penicillium needs to compete with other microorganisms for nutrients and space in saline-alkali soil to form a dominant group and play an improvement role. However, due to the special environment of saline-alkali soil, Penicillium is often difficult to survive, and the improvement effect is also difficult to show.Despite the numerous challenges faced by Penicillium and its species, such as *Penicillium clarithrum*, in the improvement of saline-alkali soils, scientists have not given up their exploration and research in this field. By optimizing inoculation methods, improving salt tolerance, and adjusting the soil microecological environment, scientists are continuously striving to enhance the effectiveness of Penicillium in saline-alkali soil improvement. At the same time, they look forward to the emergence of more new technologies and methods in the future, providing new ideas and solutions for the improvement of saline-alkali soils and the sustainable development of agriculture. In summary, Penicillium and its species, such as *Penicillium clarithrum*, have potential application value in agricultural production, but they face many challenges in the improvement of saline-alkali soils. Current technologies are not yet mature, and the improvement effects are not ideal. Summary of the Invention
[0003] To solve the above problems, the present invention provides the following specific solution:
[0004] An application of Penicillium clarithromyces in improving saline-alkali soil: a compound microbial agent containing Penicillium clarithromyces is prepared for improving saline-alkali soil. The compound microbial agent includes Penicillium clarithromyces, Sphingomonas, and Artemisia annua extract.
[0005] The strain of Penicillium kurine is named Penicillium kurine 23301-20, with accession number CGMCCNo.41049, accession date January 22, 2024, and deposited at the China General Microbiological Culture Collection Center, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing.
[0006] The colony characteristics of Penicillium clarithii are that the colony is initially white, then turns dark green, and the conidiophores branch multiple times to form a typical broom-like structure, continuously producing conidia at the top, with the conidia being nearly spherical.
[0007] Preferably, the *Sphingomonas* strain is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 19766.
[0008] Preferably, the live bacteria ratio of Penicillium clarithromyces and Sphingosine monocytogenes is (3-5):(2-3);
[0009] Preferably, the total viable count of the compound microbial agent is 1.5 × 10⁻⁶. 8 -5.5×10 9 cfu / mL.
[0010] Preferably, the preparation method of the Artemisia annua extract includes:
[0011] S1. Take fresh and clean Artemisia argyi roots, crush them and sieve them to obtain crushed roots;
[0012] S2. The obtained pulverized roots were soaked in an ethanol-water solution, and then heated and refluxed for extraction.
[0013] S3. After cooling the mixture to room temperature, filter it and concentrate the filtrate to obtain Artemisia annua extract.
[0014] Preferably, the sieving in step S1 is through a 30-60 mesh sieve; the ethanol content in the ethanol aqueous solution in step S2 is 70-75 vol%; and the mass / volume ratio of the pulverized material to the ethanol aqueous solution in step S2 is 1 g:(10-20) mL.
[0015] Preferably, the soaking temperature in step S2 is 20-35℃, and the soaking time is 18-24h; the heating and reflux is carried out at a reflux temperature of 78.5℃ and maintained in reflux state for 2-3h.
[0016] Preferably, the concentration in step S3 involves rotary evaporating the extract until no more solvent evaporates, obtaining the extract, which is then stored at 4°C for later use.
[0017] Preferably, the content of the Artemisia annua extract is 0.03-0.05 g / mL.
[0018] Preferably, the compound microbial agent includes PDA solid culture medium.
[0019] Preferably, the compound microbial agent is used for plant cultivation, and the method of application is root irrigation, with a dosage of 0.5-1.0 L / m2.
[0020] Preferably, the compound microbial agent is used for plant cultivation, and the method of use is to add it to the hydroponic system, with the volume ratio of compound microbial agent to water being 1:(500-1000).
[0021] Preferably, the preparation method of the compound microbial agent is as follows:
[0022] (1) Sphingomonas and Penicillium clarkii were inoculated into PDA solid medium and diluted with liquid to a viable count of 1.8-3.0×10⁹ cfu / mL, respectively;
[0023] (2) The obtained bacterial solution was mixed with the viable bacteria ratio of Penicillium clarithromyces and Sphingomonas at (3-5):(1-2), and the total viable bacteria count was adjusted to 1.8-3.5×109cfu / mL. Artemisia annua extract was added to the mixture to obtain a compound bacterial agent.
[0024] Preferably, the amount of Artemisia annua extract added in step (2) is 0.03-0.05 g / mL.
[0025] The beneficial effects of this invention are as follows:
[0026] The Penicillium keloidea and its composition provided by this invention can be used to improve saline-alkali soil, with significant effects and long-lasting effects, and does not cause secondary pollution to the soil. Attached Figure Description
[0027] Figure 1 This is a scanning electron microscope image of a soil sample from the planting layer after treatment in Example 1 of the present invention;
[0028] Figure 2 This is a mass spectrum of the planting layer soil sample after treatment in Example 1 of the present invention;
[0029] Figure 3 This is a microscopic particle distribution diagram of the planting layer soil sample after treatment in Example 1 of the present invention;
[0030] Figure 4 This is a scanning electron microscope image of the planting layer soil sample after treatment in Comparative Example 3 of this invention;
[0031] Figure 5 This is a mass spectrum of the planting layer soil sample after treatment in Comparative Example 3 of this invention;
[0032] Figure 6 This is a microscopic particle distribution diagram of the planting layer soil sample after treatment in Comparative Example 3 of this invention.
[0033] Figure 7 This is a scanning electron microscope image of the planting layer soil sample after treatment in Comparative Example 6 of this invention;
[0034] Figure 8 This is a mass spectrum of the planting layer soil sample after treatment in Comparative Example 6 of this invention;
[0035] Figure 9 This is a microscopic particle distribution diagram of the planting layer soil sample after treatment in Comparative Example 6 of this invention. Detailed Implementation
[0036] The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.
[0037] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those familiar to those skilled in the art. Furthermore, any methods and materials similar to or equivalent to those described herein may be applied to this invention. The preferred embodiments and materials described herein are for illustrative purposes only and do not limit the scope of this application.
[0038] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, and the experimental materials used in the following examples are all purchased from commercial channels.
[0039] Preparation Example 1: Preparation of Artemisia annua extract:
[0040] S1. Take fresh and clean Artemisia argyi roots, crush them and sieve them to obtain crushed roots;
[0041] S2. The obtained pulverized roots were soaked in an ethanol-water solution, and then heated and refluxed for extraction.
[0042] S3. After cooling the mixture to room temperature, filter it and concentrate the filtrate to obtain Artemisia annua extract.
[0043] The sieving in step S1 is through a 60-mesh sieve; the ethanol content in the ethanol-water solution in step S2 is 70 vol%; the mass / volume ratio of the pulverized material to the ethanol-water solution in step S2 is 1 g: 10 mL.
[0044] The soaking temperature in step S2 is 25°C and the soaking time is 24 hours; the heating and reflux is carried out by heating until the solvent boils and maintaining the reflux state for 2 hours.
[0045] The concentration step S3 involves rotary evaporating the extract until no more solvent evaporates, thus obtaining the extract.
[0046] Preparation Example 2: Preparation of Artemisia annua extract:
[0047] S1. Take fresh and clean Artemisia argyi roots, crush them and sieve them to obtain crushed roots;
[0048] S2. The obtained pulverized roots were soaked in an ethanol-water solution, and then heated and refluxed for extraction.
[0049] S3. After cooling the mixture to room temperature, filter it and concentrate the filtrate to obtain Artemisia annua extract.
[0050] The sieving in step S1 is through a 60-mesh sieve; the ethanol content in the ethanol-water solution in step S2 is 75 vol%; the mass / volume ratio of the pulverized material to the ethanol-water solution in step S2 is 1 g: 20 mL.
[0051] The soaking temperature in step S2 is 35°C and the soaking time is 18 hours; the heating and reflux process involves heating the solvent until it boils and maintaining the reflux state for 3 hours.
[0052] The concentration step S3 involves rotary evaporating the extract until no more solvent evaporates, thus obtaining the extract.
[0053] Example: Preparation of compound microbial agent:
[0054] (1) Sphingomonas and Penicillium clarkii were inoculated into R2A liquid medium and PDA solid medium, respectively, and diluted with purified water to a viable count of 1.0-9.0 × 10⁻⁶. 9 cfu / mL;
[0055] (2) The obtained bacterial suspension was mixed with Penicillium clarithromyces and Sphingomonas at a viable ratio of (3-5):(2-3), and the total viable count was adjusted to 1.5×10⁻⁶. 8 -5.5×10 9 Add cfu / mL of Artemisia annua extract to the resulting mixture to obtain a compound microbial agent.
[0056] The amount of Artemisia annua extract added in step (2) is 0.03-0.05 g / mL.
[0057] Example 1
[0058] The compound microbial agent was prepared according to the above method, with a viable ratio of Penicillium clarkii and Sphingomonas in the agent of 3:3; the total viable count of the compound microbial agent was 1.5 × 10⁻⁶. 8 The concentration of the Artemisia annua extract (prepared in Preparation Example 1) was 0.03 g / mL.
[0059] Example 2
[0060] The compound microbial agent was prepared according to the above method, with a viable ratio of Penicillium clarkii and Sphingomonas in the agent of 5:2; the total viable count of the compound microbial agent was 1.5 × 10⁻⁶. 8 The concentration of the Artemisia annua extract (prepared in Preparation Example 1) was 0.03 g / mL.
[0061] Example 3
[0062] The compound microbial agent was prepared according to the above method, with a viable ratio of Penicillium clarkii and Sphingomonas in the agent of 5:2; the total viable count of the compound microbial agent was 5.5 × 10⁻⁶. 9 The concentration of the Artemisia annua extract (prepared in Preparation Example 1) was 0.03 g / mL.
[0063] Example 4
[0064] The compound microbial agent was prepared according to the above method, with a viable ratio of Penicillium clarkii and Sphingomonas in the agent of 5:2; the total viable count of the compound microbial agent was 5.5 × 10⁻⁶. 9 The concentration of the *Artemisia annua* extract (prepared in Preparation Example 2) was 0.03 g / mL.
[0065] Example 5
[0066] The compound microbial agent was prepared according to the above method, with a viable ratio of Penicillium clarkii and Sphingomonas in the agent of 5:2; the total viable count of the compound microbial agent was 5.5 × 10⁻⁶. 9 The concentration of the Artemisia annua extract (prepared in Preparation Example 2) was 0.05 g / mL.
[0067] The difference between Comparative Example 1 and Example 1 is that the compound bacterial agent does not include Sphingomonas, and the number of viable Penicillium clarithromyces is the same as the total number of viable bacteria in Example 1.
[0068] The difference between Comparative Example 2 and Example 1 is that the compound bacterial agent does not include Penicillium clarkii, and the number of viable Sphingomonas is the same as the total number of viable bacteria in Example 1.
[0069] The difference between Comparative Example 3 and Example 1 is that the compound microbial agent does not include Artemisia annua extract.
[0070] Comparative Example 4: A bacterial suspension of Sphingosine Monoclonal was prepared by eluting and diluting with purified water using R2A liquid medium until the viable count was the same as the total viable count in Example 1.
[0071] Comparative Example 5: A bacterial suspension of Penicillium clarithromyces was prepared by washing and diluting with purified water using PDA solid culture medium until the viable count was the same as the total viable count in Example 1.
[0072] Comparative Example 6: The Artemisia annua extract prepared in Preparation Example 1 was added to purified water at a concentration of 0.03 g / mL.
[0073] For the blank example, prepare purified water for later use.
[0074] Experimental example:
[0075] The following experiment was conducted at our planting base in Heilongjiang Province: Soil samples were tested before the experiment; the pH was 8.77-8.92, and the total salt content was 7.66-7.91 g / kg. Corn was planted 100 kg / mu of the product obtained in the above-mentioned examples, comparative examples, and blank examples after mixing with the soil. Other planting measures and parameters were completely identical. Soil samples were taken on days 15, 30, 60, and 90 after planting to test soil salinity and pH. Soil morphology was also periodically examined, and the soil structure was scanned using electron microscopy to detect the particle content and distribution. The results were compiled and analyzed. (Soil data were collected by dividing the soil into blocks, and the average value of three points within a defined range was taken. The test samples were from the planting layer soil sample taken 15 cm below the soil surface.)
[0076] Table 1. Results of soil total salt (soluble salt g / kg) and pH test.
[0077]
[0078]
[0079] The above test results are shown in Table 1 and the attached figures in the specification. As can be seen from the results, the microbial agent provided by the present invention enhances the improvement effect of the core microbial species Penicillium kuribii on saline-alkali soil through the synergistic effect of additives and auxiliary microbial species.
[0080] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.
[0081] The present invention and its embodiments have been described above. This description is not restrictive, and the accompanying drawings are only one embodiment of the present invention. The actual application is not limited to this. In conclusion, if those skilled in the art are inspired by this description and design similar methods and embodiments without departing from the spirit of the present invention, they should all fall within the protection scope of the present invention.
Claims
1. Use of Penicillium chrysogenum in the improvement of saline-alkali soil, characterized in that: A complex microbial agent including paenibacillus chondrinos is prepared for improving saline-alkali soil, and the complex microbial agent includes paenibacillus chondrinos, sphingomonas and ironweed extract; The strain name of the paenibacillus chondrinos is paenibacillus chondrinos 23301-20, the preservation number is CGMCC No. 41049, the preservation time is January 22, 2024, the preservation unit is China General Microbiological Culture Collection Center, and the preservation address is No. 1, Xibaixili, Chaoyang District, Beijing.
2. Use of P. chrysogenum according to claim 1 in the improvement of saline soils, characterized in that: The colony character of the paenibacillus chondrinos is initially white, then becomes dark green, the conidial phialide is branched multiple times to form a typical broom structure, and the conidium is continuously produced at the top.
3. Use of P. chrysogenum according to claim 1 in the improvement of saline soils, characterized by: The live bacteria ratio of the paenibacillus chondrinos and sphingomonas is (3-5) :(2-3).
4. The use of P. chrysogenum according to claim 1 in the improvement of saline soils, characterized by the fact that: The total viable bacteria number of the complex bacterial agent is 1.5×10 8 -5.5×10 9 cfu / mL.