Synthetic biocontrol flora for preventing and controlling wheat stem rot and application of synthetic biocontrol flora

Abstract

The invention discloses a synthetic biocontrol flora for preventing and controlling wheat stem rot and application of the synthetic biocontrol flora, and relates to the technical field of microorganisms. The synthetic biocontrol flora comprises streptomyces brown L69 and pseudomonas koreensis S25, and the synthetic biocontrol flora comprises streptomyces brown L69, pseudomonas koreensis S25, pseudomonas koreensis S25, pseudomonas koreensis S25, pseudomonas koreensis S25, pseudomonas koreensis S25, pseudomonas koreensis S25, pseudomonas koreensis S25, pseudomonas koreensis S25 and pseudomonas koreensis S25. The preservation number of the streptomyces brown L69 is GDMCC No: 67852, and the preservation number of the pseudomonas koreensis S25 is GDMCC No: 67851. The streptomyces brown L69 and the preservation number of the pseudomonas koreensis S25 are respectively GDMCC No: 67852. After the synthetic biocontrol flora provided by the invention is applied to plants, the colonization level of fusarium pseudograminearum in rhizosphere and root systems of wheat can be effectively reduced, and the accumulation of pathogenic bacteria of the wheat stem rot in plants is reduced, so that the prevention and control of the wheat stem rot are realized. The invention provides a new technical path for prevention and control of wheat basal stem rot soil-borne diseases, and has good popularization and application prospects.

Description

Technical Field

[0001] This invention relates to the field of microbial technology, and in particular to a synthetic biocontrol group for controlling wheat stem base rot and its application. Background Technology

[0002] In recent years, Fusarium pseudograss ( Fusarium pseudograminearum Wheat stem rot caused by [a pathogen] has been occurring and worsening in several major wheat-producing areas, becoming one of the important soil-borne diseases threatening stable wheat yields. This pathogen mainly infects the vascular tissue at the base of the plant stem, leading to premature aging, wilting, and whiteheads, severely affecting grain formation and final yield.

[0003] Current disease control measures mainly rely on the breeding of disease-resistant varieties, agricultural cultivation regulation, and the limited application of chemical agents. However, factors such as the scarcity of resistant resources, insufficient resistance stability, and the complex and variable field ecological environment limit the long-term control effectiveness of these measures. Furthermore, chemical control not only has limited efficacy but may also lead to ecological risks such as pesticide residues, increased pathogen resistance, and soil microecological imbalance. Therefore, there is an urgent need to explore greener, safer, and more sustainable new approaches to disease control.

[0004] The rhizosphere microbiome, as a crucial functional community in the interaction between plants and the soil environment, is considered the "second genome" for crop health regulation, playing a key barrier role in inhibiting soil-borne pathogen infection, maintaining rhizosphere ecological balance, and promoting crop growth. Utilizing beneficial microorganisms to regulate the rhizosphere microecological environment and enhance plant stress resistance is considered a green and sustainable disease control strategy. However, existing biocontrol agents are mostly applied in the form of single strains, lacking sufficient adaptability and stability in the field environment, and failing to simultaneously address the synergistic needs of disease suppression and improved crop growth and quality. Therefore, developing a synthetic biocontrol microbial community that can stably colonize in the field, possessing both biocontrol and growth-promoting functions, and suitable for controlling wheat stem rot is of great significance for improving wheat health production levels and promoting green and sustainable agricultural development. Summary of the Invention

[0005] The purpose of this invention is to provide a synthetic biocontrol bacterial group for controlling wheat stem base rot and its application, thereby solving the problems existing in the prior art. When applied to plants, this synthetic biocontrol bacterial group can effectively reduce the colonization level of *Fusarium graminearum* in the rhizosphere and root system of wheat, and reduce the accumulation of wheat stem base rot pathogens within the plant, thus achieving control of wheat stem base rot.

[0006] To achieve the above objectives, the present invention provides the following solution: This invention provides a synthetic biocontrol bacterial group for controlling wheat stem base rot, including *Streptomyces brownii* (… Streptomyces aquilus L69 and Pseudomonas aeruginosa ( Pseudomonas koreensis S25; The preservation numbers of the brown Streptomyces L69 and the Korean Pseudomonas S25 are GDMCC No: 67852 and GDMCC No: 67851, respectively.

[0007] The present invention also provides a brown Streptomyces L69 for controlling wheat stem base rot, wherein the preservation number of brown Streptomyces L69 is GDMCC No: 67852.

[0008] The present invention also provides a Korean Pseudomonas S25 for controlling wheat stem base rot, wherein the Korean Pseudomonas S25 has the accession number GDMCC No: 67851.

[0009] The present invention also provides the application of the above-mentioned synthetic biocontrol bacteria in the preparation of microbial agents for the control of wheat stem base rot.

[0010] The present invention also provides a compound microbial preparation for controlling wheat stem base rot, comprising the above-mentioned synthetic biocontrol bacteria.

[0011] The present invention also provides the use of the above-mentioned Streptomyces brownii L69 or Pseudomonas kansui S25 in the preparation of microbial agents for the prevention and control of wheat stem base rot.

[0012] The present invention also provides a microbial preparation for controlling wheat stem base rot, comprising the above-mentioned Streptomyces brownii L69 or Pseudomonas koreanum S25.

[0013] The present invention also provides the application of the above-mentioned synthetic biocontrol bacteria, Streptomyces brownii L69 or Pseudomonas koreanum S25 in the prevention and control of wheat stem base rot.

[0014] The present invention also provides the application of the above-mentioned compound microbial preparation or microbial preparation in the prevention and control of wheat stem base rot.

[0015] The present invention also provides a method for preventing and controlling wheat stem base rot, comprising the step of applying the above-mentioned synthetic biocontrol bacteria, Streptomyces brownii L69 or Pseudomonas koreanum S25 to wheat plants.

[0016] The present invention discloses the following technical effects: Based on field surveys and bioinformatics analysis, this invention found that *Streptomyces brownii* L69 and *Pseudomonas kwangsiensis* S25 can significantly inhibit the growth of pathogens causing wheat stem rot, and no obvious antagonism was observed between the strains. Based on this, a synthetic biocontrol flora was constructed, including *Streptomyces brownii* L69 and *Pseudomonas kwangsiensis* S25, and its application was verified through root irrigation in pots and in the field. The results showed that the synthetic biocontrol flora provided by this invention can significantly reduce the wheat disease index and rhizosphere pathogen abundance, improve the root growth environment, and simultaneously significantly promote wheat growth, increasing traits such as single-plant fresh weight, plant height, and stem diameter.

[0017] This invention falls under the category of biological control technology. It does not rely on chemical pesticides and is characterized by safety, greenness, and environmental friendliness. It provides a stable and sustainable green control solution for soil-borne diseases of wheat. Furthermore, the synthetic biocontrol microbial community described in this invention, as a biocontrol microbial resource targeting soil-borne diseases of wheat, enriches the existing biocontrol microbial resource library, possesses good potential for field application and promotion value, and has broad application prospects and market value. Attached Figure Description

[0018] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0019] Figure 1 The results of screening for potential microorganisms related to resistance to wheat stem rot are shown in the figure. Among them, (a) is a volcano plot of bacterial community difference analysis between healthy and diseased groups; (b) is a validation volcano plot of differential ASV screening results; (c) is a comparison of the relative abundance of Bac_ASV279 in healthy and diseased wheat in different regions (Xinzheng, Xuchang, Shangqiu); (d) is a comparison of the relative abundance of Bac_ASV1448 in healthy and diseased wheat in different regions; (e) is a phylogenetic tree of Bac_ASV279 constructed based on 16S rRNA gene sequence; and (f) is a phylogenetic tree of Bac_ASV1448 constructed based on 16S rRNA gene sequence. Figure 2 The images show the morphological characteristics of two isolated microbial strains used to control wheat stem rot; (a) is a plate colony morphology image; (b) is a scanning electron microscope image. Figure 3 Phylogenetic tree diagram of strain L69; Figure 4 Phylogenetic tree diagram of strain S25; Figure 5The diagram shows the antagonistic effects of strains L69 and S25 on *Fusarium graminearum* and the compatibility analysis between the strains; (a) shows the results of plate confrontation culture and fermentation filtrate antibacterial test; (b) shows the turbidity (OD) of the bacterial filtrate in the control group (CK) and the L69 treatment group. 600 (c) Comparison chart; (d) Turbidity (OD) of bacterial culture in the control group (CK) and the S25 treatment group. 600 (d) Comparison diagram; (e) Diagram of plate confrontation culture of strains L69 and S25; (d) Diagram of chemotactic index determination results of strains L69 and S25. Figure 6 Figures showing the experimental results of the effects of different microbial inoculation treatments on wheat growth traits are shown below. (a) is a schematic diagram of each experimental treatment; (b) is a comparison diagram of plants in different treatment groups; (c) is a statistical graph of the disease index of each treatment group; (d) is a statistical graph of the abundance of pathogens (dry soil) of each treatment group; (e) is a statistical graph of the abundance of pathogens (roots) of each treatment group; (f) is a statistical graph of the abundance of pathogens (stems) of each treatment group; (g) is a statistical graph of the fresh weight of a single plant in each treatment group; (h) is a statistical graph of the plant height of each treatment group; (i) is a statistical graph of the leaf width of each treatment group; and (j) is a statistical graph of the stem diameter of each treatment group. Detailed Implementation

[0020] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0021] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Any stated value or intermediate value within a stated range, as well as each smaller range between any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0022] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0023] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0024] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0025] Biological Preservation: Brown Streptomyces ( Streptomyces aquilus L69 was deposited on February 10, 2026 at the Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC No: 67852. Korean Pseudomonas ( Pseudomonas koreensis S25 was deposited on February 10, 2026, at the Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCCNo: 67851.

[0026] Based on field surveys and bioinformatics analysis, this invention found that strains L69 and S25 can significantly inhibit the growth of wheat stem rot pathogens, and no obvious antagonism was observed among the strains. Based on this, a synthetic biocontrol flora was constructed, and its application in pots and fields was verified through root irrigation. The results show that the synthetic biocontrol flora provided by this invention can significantly reduce the wheat disease index and rhizosphere pathogen abundance, improving the root growth environment; at the same time, it significantly promotes wheat growth, increasing traits such as single-plant fresh weight, plant height, and stem diameter, as detailed below: Example 1 Screening for potential microorganisms associated with resistance to wheat stem rot: During the disease outbreak period of wheat stem rot, samples were collected from healthy or diseased wheat plants with consistent growth using a five-point sampling method, and experiments were conducted simultaneously at multiple test sites. High-throughput sequencing was used to analyze the bacterial and fungal community composition in the samples, and the relative abundance differences of wheat plant microorganisms under different disease states were compared at the genus level.

[0027] The results showed that the microbial community structure of healthy and diseased wheat plants differed significantly between different experimental sites. Screening for differentially expressed microorganisms at the amplicon sequence variant (ASV) level revealed several ASVs with significantly different abundances in the two disease states. Among them, Bac_ASV279 and Bac_ASV1448 were consistently and significantly enriched in healthy plants, showing a consistent trend across multiple experimental sites. Figure 1The relative abundance comparison results showed that the abundance of the above-mentioned ASVs in healthy plants was significantly or extremely significantly higher than that in diseased plants, exhibiting good stability and reproducibility. This indicates that Bac_ASV279 and Bac_ASV1448 can serve as potential core beneficial microbial resources significantly associated with resistance to wheat stem rot.

[0028] Example 2 Isolation of potentially resistant microorganisms: Using the ASV screened in Example 1 as the target, microorganisms were isolated from wheat rhizosphere samples using conventional isolation and culture methods. After purification, culture, and molecular biological identification, representative microbial strains corresponding to the ASV screened in Example 1 were successfully obtained. Among them, strain L69, corresponding to Bac_ASV279, was identified as *Streptomyces brownii*. Streptomyces aquilus Bac_ASV1448 corresponds to strain S25, which was identified as *Pseudomonas koraiensis*. Pseudomonas koreensis Colony morphology and scanning electron micrographs of strains L69 and S25 are shown in [reference needed]. Figure 2 Phylogenetic tree Figures 3-4 .

[0029] The aforementioned strains were identified as core rhizosphere microbial resources associated with wheat stem base rot and were used for the subsequent construction of synthetic biocontrol flora and the application verification of their biocontrol and growth-promoting effects.

[0030] Example 3 Verification of growth compatibility and antibacterial properties among strains: The compatibility between strains L69 and S25 obtained in Example 2 and their antagonistic effect against *Fusarium graminearum* were verified using a combination of plate confrontation culture and fermentation filtrate antibacterial test. The results are shown in [Figure 1]. Figure 5 .

[0031] like Figure 5 As shown in (a)-(c), both L69 and S25 effectively inhibited the mycelial growth of *Fusarium graminearum* during confrontation culture. Simultaneously, the metabolites of both strains also showed significant inhibitory effects on *Fusarium graminearum*, with L69 exhibiting a more pronounced inhibitory effect. Quantitative analysis of the fermentation filtrate showed that both L69 and S25 significantly reduced the biomass (OD) of *Fusarium graminearum*. 600 Furthermore, L69 exhibits stronger inhibitory activity than S25.

[0032] In addition, such as Figure 5 As shown in (d)-(e), no obvious inhibition zone was formed between strains L69 and S25 in the plate confrontation culture, and the colonies could grow normally in contact, indicating that there was no significant antagonistic effect between the two and that they had good growth compatibility.

[0033] The above results indicate that there is good compatibility between strain L69 and S25, and both can inhibit Fusarium pseudograminearum through their own growth and metabolites, showing the potential for synergistic application.

[0034] Example 4 Verification of the control effect and growth promotion effect of the synthetic microbial community on wheat basal stalk rot: Using the pot inoculation test method, the obtained strains L69 and S25 in Example 2 were used alone or in combination (mixed at a viable cell number ratio of 1:1) to inoculate wheat plants, so as to verify the control effect and growth promotion effect of L69 and S25 on wheat basal stalk rot. Five treatment groups were set up in the experiment: CK (sterile water control), F (only inoculated with Fusarium pseudograminearum), P+F (L69 + Fusarium pseudograminearum), S+F (S25 + Fusarium pseudograminearum), and SC+F (synthetic microbial community + Fusarium pseudograminearum). As Figure 6 shown in (a) and Table 1, first, the bacterial suspensions (10 8 cfu / mL) of strains L69 and S25 alone or in combination were used for root irrigation treatment. Three days later, Fusarium pseudograminearum (10 6 spores / mL) was inoculated (by root irrigation), and the disease occurrence and plant growth indexes were investigated during the disease onset period.

[0035] Table 1 Experimental grouping As Figure 6 shown in (c), compared with the treatment of only inoculating the pathogen (Group F), the P+F, S+F, and SC+F groups all significantly reduced the wheat disease index. Among them, the control effect of the combined treatment (SC+F) was the most stable, and the incidence rate decreased the most.

[0036] The detection results of the pathogen content in the soil showed that each treatment with bacteria could reduce the abundance of Fusarium pseudograminearum in the rhizosphere to varying degrees. The inhibitory effect of the synthetic microbial community was more obvious, as shown in Figure 6 (d)-(f).

[0037] In terms of plant growth indexes, as Figure 6 shown in (g)-(j), compared with the F treatment, the application of strains significantly increased the fresh weight per plant, plant height, and stem diameter of wheat. Among them, the growth promotion effect of the SC+F group was more prominent. As Figure 6 shown in (b), the overall growth of the plants in the P+F, S+F, and SC+F groups was significantly better than that of the treatment with only inoculating the pathogen, showing more developed roots and stronger above-ground growth.

[0038] The above results indicate that the separate or combined application of strains L69 and S25 can effectively inhibit the occurrence of wheat basal stalk rot and promote wheat growth. Among them, the synthetic microbial community shows a synergistic advantage in stable disease control and growth promotion, and has good application potential.

[0039] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A synthetic biocontrol bacterial group for controlling wheat stem base rot, characterized in that, Including Streptomyces brownii ( Streptomyces aquilus L69 and Pseudomonas aeruginosa ( Pseudomonas koreensis S25; The preservation numbers of the brown Streptomyces L69 and the Korean Pseudomonas S25 are GDMCC No: 67852 and GDMCC No: 67851, respectively.

2. A brown Streptomyces L69 strain for controlling wheat stem rot, characterized in that, The preservation number of the brown Streptomyces L69 is GDMCC No: 67852.

3. A type of *Pseudomonas aeruginosa* S25 for controlling wheat stem base rot, characterized in that... The preservation number of the Korean Pseudomonas S25 is GDMCC No: 67851.

4. The application of the synthetic biocontrol bacteria as described in claim 1 in the preparation of microbial agents for controlling wheat stem base rot.

5. A compound microbial preparation for controlling wheat stem base rot, characterized in that, Includes the synthetic biocontrol bacteria as described in claim 1.

6. The use of *Streptomyces brownii* L69 as described in claim 2 or *Pseudomonas koreana* S25 as described in claim 3 in the preparation of a microbial agent for controlling wheat stem rot.

7. A microbial preparation for controlling wheat stem base rot, characterized in that, Includes *Streptomyces brownii* L69 as described in claim 2 or *Pseudomonas koreanum* S25 as described in claim 3.

8. The application of the synthetic biocontrol bacteria group as described in claim 1, the brown Streptomyces L69 as described in claim 2, or the Korean Pseudomonas S25 as described in claim 3 in the control of wheat stem base rot.

9. The application of a compound microbial preparation as described in claim 5 or a microbial preparation as described in claim 7 in the prevention and control of wheat stem base rot.

10. A method for controlling wheat stem base rot, characterized in that, The method includes the step of applying the synthetic biocontrol bacteria of claim 1, the brown Streptomyces L69 of claim 2, or the Korean Pseudomonas S25 of claim 3 to wheat plants.

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