A strain of Streptomyces micaceus DNW2 and its application

Abstract

The present invention relates to the field of microbial control technology, and specifically to a strain of Streptomyces mycarofaciens DNW2 and an application thereof. The Streptomyces of the present invention is named Streptomyces mycarofaciens DNW2 and was deposited in the General Microbiology Center of the China Microorganism Culture Collection Administration Committee on September 2, 2022, with a deposit number of CGMCC No. 25639. The Streptomyces mycarofaciens DNW2 of the present invention is isolated from the rhizosphere soil of the virgin forest in Shunchang County, Nanping City, Fujian Province. It has strong growth adaptability, high acid-base tolerance, and can be harmoniously compatible with the soil ecology. The biocontrol agent prepared by the strain can effectively prevent and control various plant diseases such as honey pomelo black spot disease, honey pomelo anthracnose, honey pomelo black spot disease, pepper wilt, tomato wilt, banana wilt, banyan anthracnose, succulent black spot disease, tomato leaf mold, orchid stem base rot, corn stalk rot, strawberry root rot, etc., especially for honey pomelo black spot disease. The control effect is significant, and it has the advantages of being pollution-free and promoting growth.

Description

Technical Field

[0001] The present invention relates to the technical field of microbial control, and in particular to a Streptomyces micaceus DNW2 and an application thereof. Background Art

[0002] Honey pomelo black spot disease, also known as sand bark disease, resin disease, and brown stem rot, is caused by Diaporthe fungi and primarily infects new shoots, young leaves, flowers, and young fruits. It causes tree weakness, flower drop, rough fruit surfaces, and even fruit rot, severely impacting both yield and quality. It has become a major disease of honey pomelo. Current methods for controlling honey pomelo black spot disease primarily rely on chemical pesticides. While these can alleviate symptoms to a certain extent, their effectiveness is unsatisfactory due to widespread overuse. The long-term and excessive use of chemical pesticides has led to increased resistance in the honey pomelo black spot pathogen, making control increasingly difficult. Furthermore, excessive pesticide use has resulted in significant pesticide residues, exacerbating environmental pollution and posing a serious threat to human health. Therefore, the development of effective and environmentally friendly control measures is imperative.

[0003] Biological microbial agents have the advantages of being non-toxic, residue-free, environmentally friendly, and pathogens are not prone to developing drug resistance. The use of antagonistic microorganisms to control plant diseases is an effective way to solve the above problems. It is a new resource for controlling plant diseases and an effective green and sustainable prevention and control measure. Streptomyces is the most widely used group in the Actinomycetes. It has strong environmental adaptability and is one of the most common microorganisms in the rhizosphere of plants. It has been reported to be used to control a variety of plant diseases. Streptomyces can produce secondary metabolites and extracellular hydrolases to inhibit the growth of a variety of pathogens. In addition, it can produce antibiotics and plant growth hormones to promote plant growth and improve the plant's resistance to adversity. Therefore, the application of Streptomyces agents to control Guanxi pomelo black spot disease is of great significance. Summary of the Invention

[0004] In response to the deficiencies in the prior art, the present invention provides a strain of Streptomyces mycarofaciens DNW2 and applications thereof. The Streptomyces mycarofaciens DNW2 is obtained from the soil of a primeval forest that has not been walked on for many years. The Streptomyces mycarofaciens DNW2 has a strong antagonistic inhibitory effect on a variety of plant pathogens, and has a particularly good preventive and therapeutic effect on honey pomelo black spot disease. In addition, because it is a microorganism existing in the biological world, it will not cause pathogens to develop drug resistance, and is friendly to the natural environment and pollution-free.

[0005] To achieve the above-mentioned purpose, the first aspect of the present invention provides a strain of Streptomyces, which is named Streptomyces mycarofaciens DNW2 and was deposited in the General Microbiology Center of the China Culture Collection of Microorganisms on September 2, 2022, with the deposit number CGMCC No. 25639. The deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences.

[0006] Streptomyces benthamiana DNW2 was isolated from soil in a pristine forest that had been untouched for many years. It thrives on Gao's No. 1, PDA, ISP2, ISP3, ISP4, ISP7, and tyrosine media, and grows well on ISP5 and Czapek's medium. It can utilize glucose, α-galactose, maltose, D-fructose, rhamnose, D-arabinose, inositol, D-mannitol, and sucrose as carbon sources, and L-tryptophan, glycine, L-tyrosine, L-histidine, arginine, ammonium chloride, and potassium nitrate as nitrogen sources. It can coagulate and thaw milk and reduce nitrates, but cannot liquefy gelatin, nor produce H2S or melanin. It exhibits strong temperature tolerance, growing between 15 and 40°C, with an optimal growth temperature of 28°C. It also exhibits strong pH tolerance, tolerating a pH range of 4 to 13, with an optimal pH of 7.2.

[0007] The inventors discovered that the Streptomyces micaceus DNW2 strain thrives on Gao's medium No. 1, with lush, off-white aerial hyphae and orange-yellow basal hyphae. After three days of cultivation at 28°C, white spore threads begin to emerge from the center of the colony, gradually turning off-white after 10 days, and no soluble pigment is produced. Scanning electron microscopy revealed that the basal hyphae of the Streptomyces micaceus DNW2 strain were straight and highly branched, with open, primitive spirals or loose spirals, often irregular and clumping. Upon maturation, the spore threads form well-developed, beaded spore chains, which break into nearly spherical spores with spikes on the spore surface.

[0008] In a second aspect, the present invention provides a biological control agent comprising the aforementioned Streptomyces micaceus DNW2, and one or more of its fermentation broth product, fermentation supernatant, and a dilution of the fermentation supernatant.

[0009] In a third aspect, the present invention provides a method for preparing the above-mentioned biological control agent, wherein the Streptomyces micaceus DNW2 is activated and inoculated into a fermentation medium, and a fermentation broth product of the Streptomyces micaceus DNW2 is obtained after fermentation.

[0010] Furthermore, the fermentation broth of Streptomyces micaceus DNW2 was centrifuged to obtain a fermentation supernatant.

[0011] Furthermore, the fermentation supernatant is diluted 1 to 10 times to obtain a diluted solution.

[0012] Furthermore, the fermentation medium includes the following raw material components in percentage: 1-6% corn flour, 0.5-2% soybean flour, 0.1-0.5% NaCl, and 0.1-0.5% CaCO3. The pH of the fermentation medium is 7.0-7.2. The fermentation culture conditions are: fermentation temperature 26-28°C, fermentation time 5-7 days, liquid volume 50-100mL / 250mL, inoculation size 3-6%, and rotation speed 150-180r / min.

[0013] Specifically, the fermentation medium preferably comprises the following raw material components in percentage: 3.2% corn flour, 1% soybean flour, 0.25% NaCl, and 0.15% CaCO3.

[0014] In a fourth aspect of the present invention, the present invention provides the use of the above-mentioned Streptomyces DNW2 or the above-mentioned biological control agent in the preparation of a plant pathogen disease control product, wherein the plant pathogens are Diaporthe citri, Colletotrichum gloeosporioides, Phyllosticta citriasiana, Fusarium oxysporium, Fusarium oxysporium f.sp.lycopersici, Fusarium oxysporium f.sp.cubense, Colletotrichum gloeosporioides, Alternaria alternata, Fulvia fulva, Fusarium sp., Fusarium oxysporium f.sp.cubense, and Alternaria alternata. verticillioides), strawberry root rot fungus (Neopestalotiopsis clavispora) or one or more thereof.

[0015] Furthermore, the plant pathogen is preferably pomelo black spot pathogen.

[0016] Furthermore, the plant is preferably Guanxi Pomelo.

[0017] In a plate standoff test, the inventors found that live Streptomyces DNW2 strains had an inhibition band of 13.8±0.4mm against the black spot pathogen of pomelo, with an inhibition rate of 74.13%. Through fermentation optimization, the DNW2 fermentation broth had an inhibition band of 16.2mm against the black spot pathogen, a 14.8% increase compared to the pre-optimization level. Artificial inoculation control experiments showed that the active substances in the optimized fermentation broth of Streptomyces DNW2 had a significant control effect against pomelo black spot, with protective and therapeutic effects reaching 84.88% and 72.65%, respectively. This control effect surpassed the chemical agents currently used to control Guanxi pomelo black spot disease, and it also possessed the advantages of being drug-resistant, low-toxic, residue-free, and environmentally friendly.

[0018] In a fifth aspect, the present invention provides a method for preventing and controlling plant pathogenic bacteria diseases, wherein the above-mentioned biological control agent or the biological control agent prepared by the above-mentioned preparation method is applied to at least one of the fruits, leaves, flowers and branches of Guanxi pomelo.

[0019] Beneficial effects of the present invention:

[0020] This study, conducted by conducting microbial isolation and pathogen confrontation tests on rhizosphere soil from forests that had been uninhabited for many years, isolated and purified a biocontrol strain—Streptomyces micaceus DNW2—that is safe for humans and animals, exhibits a broad antibacterial spectrum, and has promising application prospects. The fermentation broth of this DNW2 strain exhibited strong inhibitory effects on the mycelial growth of various pathogens tested, achieving an inhibition band of 13.8 mm against the pomelo black spot pathogen, and after optimization, the inhibition band reached 16.2 mm.

[0021] The biocontrol agent provided by the present invention, which contains Streptomyces micaceus DNW2 and / or fermentation metabolites of the Streptomyces micaceus DNW2, is a biological preparation and is used to prevent and control black spot disease of Guanxi honey pomelo. It has good prevention and control effect and solves the problems of pathogen resistance, pesticide residues in fruits, and serious pollution of the natural environment on which humans depend for survival caused by chemical control in the prior art, and is conducive to the pollution-free production of Guanxi honey pomelo. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 3. The morphology of aerial hyphae and spore hyphae of Streptomyces micaceus DNW2 strain of the present invention;

[0023] Figure 2 spore chains and spore morphology of the Streptomyces micaceus DNW2 strain of the present invention;

[0024] Figure 3 The phylogenetic tree constructed from the 16S rDNA sequence of the Streptomyces micaceus DNW2 strain of the present invention;

[0025] Figure 4This is a diagram showing the antagonistic effect of the Streptomyces micaceus DNW2 strain of the present invention on pomelo black spot disease.

[0026] The present invention will be further described below with reference to the accompanying drawings and embodiments. DETAILED DESCRIPTION

[0027] In order to better illustrate the technical scheme of the present invention, the scheme of the present invention will be explained below in conjunction with embodiment.It will be understood by those skilled in the art that the following examples are only used to illustrate the present invention and should not be regarded as limiting the scope of the present invention.In the embodiment, if specific technology or conditions are not indicated, it is carried out according to the technology described in the literature or reference books in this area or according to the product instructions.The reagents used or the instruments and equipment that do not indicate the manufacturer are conventional products, and the reagents used are prepared by conventional methods unless otherwise specified.

[0028] The above-mentioned technical features of the present invention and the technical features specifically described below (such as implementation cases) can be combined with each other to form new or preferred technical solutions.

[0029] Example 1: Acquisition of Streptomyces micaceus DNW2 strain

[0030] In rhizosphere soil from a forest in Shunchang County, Nanping City, Fujian Province, where no one has walked for many years, the topsoil layer was removed, and soil samples were collected from the 10-15 cm depth. These samples were then brought back to the laboratory and allowed to dry naturally in the shade. After drying, the soil samples were diluted with sterile water to obtain dilutions of 10⁻¹, 10⁻², 10⁻³, 10⁻4, 10⁻5, and 10⁻6. These dilutions were then cultured on Gao's medium No. 1 at 28°C and purified using conventional plate separation. The purified Streptomyces natica DNW2 strain was transferred to Gao's liquid medium No. 1 and cultured at 28°C and 180 rpm to obtain fermentation broth of Streptomyces natica DNW2. The Streptomyces mycarofaciens DNW2 was deposited in the General Microbiology Center of the China Culture Collection Administration on September 2, 2022, with the deposit number CGMCC No. 25639. The deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, with the postal code 100101. Its classification name is Streptomyces mycarofaciens.

[0031] The preparation of Gao's No. 1 medium and PDA medium was based on the book "Plant Disease Research Methods" by Fang Zhongda, published by China Agriculture Press in 1998.

[0032] Example 2: Physiological and biochemical characteristics and culture characteristics of Streptomyces micaceus DNW2 strain

[0033] Streptomyces niger DNW2 was inoculated onto nine different culture media. Colonies began to form after 36–48 hours of incubation at 28°C. After 72 hours, white conidia began to grow from the center of the colonies. The colonies were smooth, with wrinkled edges that made them easy to lift. The spore masses ranged from milky white to dark gray. The colonies on the different media varied in morphology, conidia color, aerial hyphae color, basal hyphae color, soluble pigment production, and growth. Observation and comparison were performed using the methods and color plates provided in the "Streptomyces Identification Manual." The results are shown in Table 1.

[0034] Table 1 Culture characteristics of strain DNW2 on various culture media

[0035]

[0036] Note: “—”: no soluble pigments are produced, “+”: more and more vigorous growth

[0037] From the results in Table 1 , it can be seen that the DNW2 strain of Streptomyces nicotianae grows vigorously on most culture media, does not produce soluble pigments, the aerial hyphae change from off-white to gray to dark gray, and the basal hyphae are yellow-white to green-yellow.

[0038] Streptomyces naecogeneticus DNW2 was inoculated onto different carbon and nitrogen sources, gelatin, milk, Chessner, and nitrate-reducing media. Colonies began to form after 48–72 hours of incubation at 28°C. Colony growth varied on the different media, as observed using the method described in the Streptomyces Identification Manual. The results are shown in Table 2.

[0039] Table 2 Phenotypic characteristics of strain DNW2

[0040]

[0041] Note: “+” indicates a positive reaction; “-” indicates a negative reaction

[0042] From the results in Table 2, it can be seen that the DNW2 strain of Streptomyces nicotianae can utilize carbon sources such as α-galactose, D-arabinose, sucrose, maltose, D-fructose, rhamnose, glucose, inositol, and D-mannitol, as well as nitrogen sources such as L-tryptophan, glycine, L-tyrosine, L-histidine, arginine, ammonium chloride, and potassium nitrate; it can coagulate and freeze milk and reduce nitrates, but cannot liquefy gelatin or produce H2S and melanin.

[0043] The DNW2 strain of Streptomyces nicotianae has a strong tolerance to temperature and can grow between 15 and 40°C, with 28°C being the optimal growth temperature. It also has a strong tolerance to pH and can tolerate a pH range of 4 to 13, with an optimal pH of 7.2.

[0044] The DNW2 strain of Streptomyces nicotianae grew vigorously on Gao's medium No. 1, with abundant aerial hyphae. After culturing at 28°C for 48 hours, the colony was smooth and no spores were formed. After 72 hours, milky white spore hyphae began to grow from the middle of the colony, and after 240 hours, they began to turn grayish white, and no soluble pigment was produced.

[0045] The DNW2 strain of Streptomyces micaceus was observed under a scanning electron microscope. The hyphae in the base were straight and highly branched. The spore threads were open-ringed, primitive spiral or loose spiral, often irregular and clustered. When the spore threads matured, they formed beaded spore chains. The spore chains were well developed and formed nearly spherical spores after the spore chains broke. There were spikes on the surface of the spores. The spore size was 0.57-0.77 μm × 0.91-1.14 μm. Figure 1 Shown are the morphologies of aerial hyphae and spore hyphae of Streptomyces nicotianae DNW2 strain. Figure 2 The spore chain and spore morphology of Streptomyces micaceus DNW2 strain.

[0046] Example 3: 16S rDNA sequence analysis

[0047] The Streptomyces DNW2 strain of Streptomyces spp. was inoculated on Gao's solid medium No. 1, cultured at 28°C for 7 to 8 days, and sent to Sangon Biotech (Shanghai) Co., Ltd. for strain identification. The genomic DNA of strain DNW2 was extracted using a bacterial genome kit, and 16S rDNA was amplified by PCR using universal primers 7F (SEQ ID NO.1): 5'-CAGAGTTTGATCCTGGCT-3' and 1540R (SEQ ID NO.2): 5'-AGGAGGTGATCCAGCCGCA-3'. The target DNA fragment was purified and recovered using a gel purification kit (AXYGEN), and after ligation, transformation, and identification, sequencing was completed to obtain a full-length sequence of 1468 bp, as shown in SEQ ID NO.3. The sequencing results were compared by BLAST sequence alignment on the NCBI website, and the 16S rDNA phylogenetic tree was constructed using the neighbor-joining method using the software MEGA 7.0, as shown in Figure 3 Strain DNW2 belongs to the same branch as Streptomyces mycarofaciens (gene accession number EF063483.1), with a similarity of 99.93%. Combining morphological and cultural characteristics, strain DNW2 was identified as Streptomyces mycarofaciens.

[0048] Example 4: Antibacterial test of Streptomyces micaceus DNW2

[0049] The plate standoff assay was used to determine the antagonistic activity of Streptomyces niger DNW2 against various crop pathogens. S. niger DNW2 was inoculated in Gao's No. 1 liquid culture medium and cultured at 28°C with shaking at 180 rpm for 7 days to obtain the fermentation broth. The test pathogens were activated by inoculating them onto polydimethylsiloxane (PDA) plates to form a bacterial cake (d = 4 mm). The cake, with the mycelial side facing downward, was placed in the center of the PDA plate. Two circular filter paper discs (d = 4 mm) soaked in the fermentation broth were placed 25 mm apart on either side of the cake. Each treatment was replicated three times. The plates were cultured at 28°C for 5-7 days, and the inhibition distance between the antagonistic bacterial colony and the pathogen colony edge was measured. Strain DNW2 exhibited varying degrees of inhibitory activity against various crop pathogens. The antagonistic effects are shown in Table 3.

[0050] Table 3 Antagonistic effect of Streptomyces micaceus DNW2 strain against 14 different crop pathogens

[0051]

[0052] Note: Data in the table are mean ± standard deviation. Different uppercase and lowercase letters in the same column indicate significant differences at P < 0.01 and P < 0.05 levels as tested by Duncan's new multiple range method.

[0053] The results in Table 3 show that Streptomyces DNW2 strain has a certain degree of antagonism against all tested pathogens. It has a strong antagonistic effect against pomelo black spot pathogen, pomelo black spot pathogen, pomelo anthracnose pathogen, fig anthracnose pathogen, tomato leaf mold pathogen, and strawberry root rot pathogen (Neocladactylodiscus hirsutus), with inhibition zones greater than 10 mm. Figure 4 These are photos of a plate confrontation test of Streptomyces DNW2 strain against pomelo black spot pathogen, indicating that DNW2 strain has a strong antagonistic effect against pomelo black spot pathogen.

[0054] Example 5: Preparation of biocontrol agent A-Streptomyces micaceus DNW2 fermentation broth

[0055] A fermentation medium was prepared using distilled water according to the following formula (w / v): 3.2% corn starch, 1% soybean flour, 0.25% NaCl, 0.15% CaCO₃, pH 7.2. 75 mL was dispensed into 250 mL Erlenmeyer shake flasks and sterilized by high-pressure steam at 121°C for 20 minutes. A 5% inoculum of Streptomyces mikae DNW2 was aseptically inoculated into the sterilized fermentation medium, sealed with eight layers of gauze, and incubated at 28°C at a constant temperature of 160 rpm for 5 days to obtain a fermentation broth, which is biocontrol agent A.

[0056] Example 6: Preparation of fermentation supernatant of biocontrol agent B / C-Streptomyces micaceus DNW2

[0057] A fermentation medium was prepared with distilled water according to the following formula (w / v): 4.5% corn starch, 0.5% soybean flour, 0.5% NaCl, 0.3% CaCO₃, pH 7.1. 75 mL of the medium was dispensed into 250 mL Erlenmeyer shake flasks and sterilized by high-pressure steam at 121°C for 20 minutes. A 5% inoculum of Streptomyces micaceus DNW2 was aseptically inoculated into the sterilized fermentation medium. The medium was sealed with eight layers of gauze and incubated at 28°C for 6 days with shaking at 180 rpm to obtain a fermentation broth. The fermentation broth was centrifuged at 6000 rpm for 20 minutes, and the mycelium was removed using a sterilizing filter to obtain a fermentation supernatant, which was biocontrol agent B. This fermentation supernatant was diluted 10-fold to obtain biocontrol agent C.

[0058] Example 7: Biocontrol Agents for the Control of Guanxi Pomelo Black Spot Disease

[0059] Infection of Guanxi Pomelo Black Spot Pathogen: Guanxi Pomelo Black Spot Pathogen was inoculated with alfalfa decoction + Czapek medium (formula: 100 g alfalfa stems and leaves, 2 g sodium nitrate, 0.5 g potassium chloride, 0.01 g ferrous sulfate, 1 g dimethyl phosphate, 0.5 g magnesium sulfate, 30 g sucrose, 15 g agar powder, 1000 mL water), and cultured at 26°C for 25-30 days. Spore horns and spore-bearing bodies were then picked to prepare 1×10 5 mL -1 Healthy, fresh young pomelo fruits were selected, surface disinfected with 75% alcohol, rinsed three times with sterile water, and air-dried. Using the puncture inoculation method, the fruit surface was punctured (approximately 1 mm in depth) with a No. 5 insect needle (d = 0.7 mm). Then, 20 μL of black spot pathogen spore suspension was inoculated into the wound and the surrounding area of ​​the young fruit. After inoculation, the fruit was placed in a plastic box (relative humidity approximately 90%) and placed in a 28°C light incubator. Each treatment included 15 fruits, and was repeated three times.

[0060] Comparison of antibacterial effects of fermentation broth: 24 h before inoculation and 24 h after inoculation, DNW2 fermentation broth (biocontrol agent A), DNW2 fermentation supernatant (biocontrol agent B), 10-fold dilution of DNW2 fermentation supernatant (biocontrol agent C), 500-fold dilution of 80% mancozeb, 500-fold dilution of 10% difenoconazole, and 5 mL of sterile water were evenly applied to the wounds of young fruits and the surrounding fruit surfaces. After inoculation, the disease condition of the fruits was observed every day, and the incidence rate, disease index, and control effect were calculated.

[0061] Incidence rate = (total number of diseased plants in the treatment / total number of plants in the treatment) / 100%;

[0062] Disease index = [∑(number of diseased plants × representative value) / (total number of plants × representative value of the most severe disease level)] × 100;

[0063] Control effect (%): (control disease index - treatment disease index) / control disease index × 100;

[0064] The disease index grading standard refers to the fruit disease grading standard of Chen Guoqing et al.

[0065] Level 0: No lesions on the fruit;

[0066] Level 1: The area of ​​lesions on the fruit accounts for less than 5% of the fruit area;

[0067] Level 3: The area of ​​lesions on the fruit accounts for 6% to 10% of the fruit area;

[0068] Level 5: The area of ​​lesions on the fruit accounts for 11% to 25% of the fruit area;

[0069] Level 7: The area of ​​lesions on the fruit accounts for 26% to 50% of the fruit area;

[0070] Level 9: The area of ​​lesions on the fruit accounts for more than 50% of the fruit area.

[0071] Table 4 Control effect of Streptomyces micaceus DNW2 on black spot disease of Guanxi pomelo

[0072]

[0073] Note: Data in the table are mean ± standard deviation. Different uppercase and lowercase letters in the same column indicate significant differences at P < 0.01 and P < 0.05 levels as tested by Duncan's new multiple range method.

[0074] The results in Table 4 show that the disease incidence and disease index of the control fruits were 84.44% and 34.32% respectively; the disease incidence and disease index of the fruits sprayed with biocontrol agent A before inoculation with pathogens were 20.00% and 5.19% respectively, while the disease incidence and disease index of the fruits sprayed with biocontrol agent A after inoculation with pathogens were 31.11% and 9.38% respectively. The protective and therapeutic efficacy of the fruits treated with biocontrol agent B were 64.72% and 60.38% respectively; the protective and therapeutic efficacy of the fruits treated with biocontrol agent C were 58.31% and 46.01% respectively; the protective and therapeutic efficacy of the fruits treated with 500-fold diluted 80% mancozeb were 87.84% and 71.61% respectively; and the protective and therapeutic efficacy of the fruits treated with 500-fold diluted 10% difenoconazole were 54.70% and 43.12% respectively. The experiment found that lesions on control fruits expanded over the following weeks, and some fruits rotted. However, lesions on treated fruits barely expanded. Biocontrol agents A, B, and C all showed significant inhibitory effects against the black spot pathogen of honey pomelo, demonstrating a good control effect even on honey pomelo infected with black spot. The protective and therapeutic effects of biocontrol agent A were slightly lower than those of a 500-fold dilution of 80% mancozeb, but the difference was not significant. However, they were significantly higher than those of a 500-fold dilution of 10% difenoconazole.

[0075] Finally, it should be emphasized that the above is only a preferred embodiment of the present invention and is not intended to limit the present invention. For those skilled in the art, the present invention may have various changes and modifications. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A strain of Streptomyces, characterized in that The Streptomyces was named Streptomyces spp. Streptomyces mycarofaciens )DNW2, and was deposited in the General Microbiology Center of China Culture Collection Administration on September 2, 2022, with the deposit number CGMCC No.25639.

2. A biological control agent, characterized in that: The invention comprises the Streptomyces micaceus DNW2 or its fermentation broth product according to claim 1.

3. The method for preparing the biological control agent according to claim 2, wherein: The Streptomyces micaceus DNW2 is activated and inoculated into a fermentation medium, and a fermentation liquid product of the Streptomyces micaceus DNW2 is obtained after fermentation.

4. A biological control agent, characterized in that: The fermentation supernatant comprising the Streptomyces micaceus DNW2 of claim 1 or a dilution of the fermentation supernatant.

5. The method for preparing the biological control agent according to claim 4, characterized in that: The Streptomyces micaceus DNW2 is activated and inoculated into a fermentation medium, and a fermentation liquid of the Streptomyces micaceus DNW2 is obtained after fermentation. The fermentation liquid is centrifuged to obtain a fermentation supernatant.

6. The method for preparing the biological control agent according to claim 5, characterized in that: The fermentation supernatant is diluted 1 to 10 times to obtain a diluted fermentation supernatant.

7. The method for preparing the biological control agent according to claim 3 or 5, characterized in that: The fermentation medium includes the following raw material components in percentage: 1-6% corn flour, 0.5-2% soybean flour, 0.1-0.5% NaCl, and 0.1-0.5% CaCO3. The pH of the fermentation medium is 7.0-7.

2. The fermentation culture conditions are: fermentation temperature 26-28°C, fermentation time 5-7 days, liquid volume 50-100 mL / 250 mL, inoculation size 3-6%, and rotation speed 150-180 r / min.

8. Use of the Streptomyces according to claim 1 or the biological control agent according to claim 2 in the preparation of a plant pathogen disease control product, characterized in that: The plant pathogens are one or more of pomelo black spot pathogen, pomelo anthracnose pathogen, pomelo black spot pathogen, pepper wilt pathogen, tomato wilt pathogen, banana wilt pathogen, banyan anthracnose pathogen, succulent black spot pathogen, tomato leaf mold pathogen, orchid stem base rot pathogen, corn stem rot pathogen, and strawberry root rot pathogen; the strawberry root rot pathogen is Neorobustidae, Fusarium spp., or Fusarium solani.

9. The use according to claim 8, characterized in that The plant pathogen is pomelo black spot pathogen.

10. The use according to claim 8 or 9, characterized in that: The plant is Guanxi honey pomelo.

11. Use of the biological control agent according to claim 4 in preparing a product for controlling honey pomelo black spot pathogen.

12. A method for preventing and controlling plant pathogenic bacteria diseases, characterized in that: The biological control fungicide according to claim 2 or the biological control fungicide according to claim 4 is applied to at least one of the fruits, leaves, flowers and branches of Guanxi pomelo to prevent and control the black spot disease of pomelo.

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