Streptomyces griseoviridis AH001 isolated from aglaia odorata and application thereof

Abstract

This invention belongs to the field of microbial technology, specifically relating to a strain of *Streptomyces gracilis* AH001 isolated from *Anthurium stenoptera* and its applications. *Streptomyces gracilis* (… Streptomyces pratensis Streptomyces AH001 was deposited at the Guangdong Provincial Microbial Culture Collection Center on January 14, 2026, with accession number GDMCC No. 67655. Indoor experiments have shown that Streptomyces AH001 has a significant inhibitory effect on various pathogens originating from Anthurium andraeanum, and can be used to prepare highly effective microbial biocontrol agents to ensure biological and environmental safety while controlling Anthurium andraeanum diseases.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology, specifically relating to a strain of Streptomyces agrarianus AH001 isolated from Anthurium and its applications. Background Technology

[0002] Anthurium ( Anthurium andraeanum Anthurium (also known as Flamingo Flower or Anthurium 'Anthurium'), a member of the Araceae family, is an ornamental plant belonging to the Anthurium genus. It is a well-known high-end cut flower with significant economic benefits. However, with the expansion of Anthurium cultivation and the increase in its age, many root, leaf, and flower diseases are becoming increasingly frequent, greatly affecting the yield and quality of Anthuriums. In the context of diversified pest control methods, beneficial microorganisms, green biological agents, and the screening of highly resistant varieties have advantages in terms of good control effects and low environmental harm, ensuring biological and environmental safety while controlling Anthurium pests and diseases.

[0003] Streptomyces, as the most advanced actinomycetes, are widely distributed in nature. Studies have shown that over 13,700 of the more than 33,000 known microbial secondary metabolites are produced by actinomycetes. Streptomyces have multiple functions; some can produce antibiotics and plant growth hormones, promoting plant growth and enhancing their resistance to biotic and abiotic stresses. Streptomycin, widely used today, originates from Streptomyces.

[0004] There are currently no records of the inhibitory effect of Streptomyces on the pathogen of Anthurium in cut flowers. Summary of the Invention

[0005] To address the shortcomings of the existing technology, the purpose of this invention is to provide a *Streptomyces* strain AH001 isolated from *Anthurium stenoptera* that exhibits biocontrol effects against multiple pathogenic fungi, and its applications. To achieve the above objective, this invention adopts the following technical solution:

[0006] In a first aspect, the present invention provides a strain of *Streptomyces gracilis* AH001 isolated from *Anthurium rosaceum*. *Streptomyces gracilis* ( Streptomyces pratensis AH001 was deposited at the Guangdong Provincial Center for Microbial Culture Collection on January 14, 2026, with accession number GDMCC No. 67655.

[0007] A second aspect of the invention provides a derivative of Streptomyces agronomica AH001, comprising one or more of its live bacteria, inactivated bacterial cells, fermentation broth, exosomes, or metabolites.

[0008] A third aspect of the invention provides a microbial agent containing Streptomyces agronomicum AH001 or a derivative thereof.

[0009] In some embodiments, the microbial agent contains Streptomyces AH001 or its fermentation broth. Further, the microbial agent is a sterile fermentation filtrate of Streptomyces AH001.

[0010] This invention demonstrates through experiments that the aseptic fermentation filtrate of Streptomyces AH001 has the most significant antibacterial effect, achieving a 100% inhibition rate against Anthracnose bonningensis and Cladosporium, and an inhibition rate of over 75% against Anthracnose spp., Colletotrichum gloeosporioides, Anthracnose sicca, Fusarium moniliforme, Fusarium oxysporum, and Alternaria alternata, which is significantly higher than that of Streptomyces AH001 bacterial solution.

[0011] In some embodiments, the microbial agent may also include a carrier or excipients.

[0012] A fourth aspect of the present invention provides a method for preparing a microbial agent, comprising the steps of culturing Streptomyces agronomicum AH001 and obtaining a culture.

[0013] Further, the steps include: inoculating *Streptomyces aureus* AH001 into Gao's No. 1 liquid culture medium, and culturing at 25-30℃ and 150-200 r / min for 2 days to obtain seed culture; taking 1% of the seed culture and placing it in Gao's No. 1 liquid culture medium, and culturing at 25-30℃ and 150-200 r / min for 5 days to obtain fermentation broth; centrifuging the fermentation broth at 4℃ and 6000-8000 r / min for 8-10 min, and filtering the supernatant through a 0.22 μm filter membrane to remove bacterial cells, obtaining *Streptomyces aureus* AH001 sterile fermentation filtrate, which is the final product.

[0014] A fifth aspect of the present invention provides the application of *Streptomyces cepacia* AH001 or its derivatives or inoculants in antagonizing plant pathogens that cause disease in plants, wherein the plant pathogen is *Anthracnose boningensis* (…). Colletotrichum boninense ), fruit anthracnose ( Colletotrichum fructicola ), Colloidal anthrax bacteria ( Colletotrichum gloeosporioides ), Siamese anthrax bacteria ( Colletotrichum siamense Fusarium moniliforme ( ) Fusarium proliferatum Fusarium oxysporum ( Fusarium oxysporum ), Alternaria ( Alternaria alternata ) or apical spores ( Acremonium sp.).

[0015] A sixth aspect of the present invention provides a product comprising *Streptomyces cepacia* AH001 or the above-mentioned fungal agent; the product has the effect of antagonizing plant pathogens on plant pathogens, wherein the plant pathogen is *Anthracnose boningensis* (…). Colletotrichum boninense ), fruit anthracnose ( Colletotrichum fructicola ), Colloidal anthrax bacteria ( Colletotrichum gloeosporioides ), Siamese anthrax bacteria ( Colletotrichum siamense Fusarium moniliforme ( ) Fusarium proliferatum Fusarium oxysporum ( Fusarium oxysporum ), Alternaria (Alternaria alternata ) or apical spores ( Acremonium sp.).

[0016] A seventh aspect of the present invention provides a method for preventing and controlling plant diseases, the method comprising applying Streptomyces agrophytes AH001 or the above-mentioned fungal agent or the above-mentioned product to the plant.

[0017] Compared with the prior art, the technical solution of the present invention has the following beneficial effects: This invention isolated a strain of Streptomyces gracilis AH001, Streptomyces gracilis ( Streptomyces pratensis The strain AH001 was deposited at the Guangdong Provincial Microbial Culture Collection Center on January 14, 2026, with accession number GDMCC No. 67655. Indoor experiments showed that *Streptomyces simonii* AH001 exhibited significant inhibitory effects on various pathogens isolated from *Anthurium stenoptera*. This strain can be used to prepare highly effective microbial biocontrol agents, effectively controlling diseases in *Anthurium stenoptera* while ensuring biosafety and environmental safety, demonstrating promising application prospects. Attached Figure Description

[0018] Figure 1 These are colony morphology photographs of strain AH001 in this embodiment of the invention. (A) shows the front view of a single colony of strain AH001; (B) shows the back view of a single colony of strain AH001; (C) shows the hyphae morphology of strain AH001 in the substrate; and (DG) show the colony morphology of AH001 at 3d, 5d, 7d, and 30d, respectively.

[0019] Figure 2 The optical microscope images show the morphological characteristics of strain AH001 in this embodiment of the invention; where (A) is the hyphal morphology of AH001; (B) is the sporulation structure of AH001; and (C) is the conidial chain of AH001.

[0020] Figure 3 The electron microscope morphological characteristics of strain AH001 in the embodiments of the present invention are shown.

[0021] Figure 4 The above are the bacterial culture and sterile fermentation filtrate of Streptomyces agronomica AH001 in the embodiments of the present invention; wherein (A) is the bacterial culture of Streptomyces agronomica AH001 2d; and (B) is the sterile fermentation filtrate of Streptomyces agronomica AH001.

[0022] Figure 5This figure shows the evaluation of the antibacterial effect of Streptomyces agronomicum AH001 on pathogens in an embodiment of the present invention. In the figure, CK1 represents the colony morphology of pathogens after 7 days of incubation with sterile water, A represents the antibacterial effect of experimental group A on pathogens, B represents the antibacterial effect of experimental group B on pathogens, CK2 represents the colony morphology of pathogens after 7 days of incubation, and C represents the antibacterial effect of experimental group C on pathogens. Detailed Implementation

[0023] It should be noted that the following detailed descriptions are exemplary and intended to provide further illustration of the invention. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0024] It should be noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the exemplary embodiments of the present invention. As used herein, the singular form is intended to include the plural form as well, unless the context clearly indicates otherwise. Furthermore, it should be understood that when the terms "comprising" and / or "including" are used in this specification, they indicate the presence of features, steps, operations, and / or combinations thereof.

[0025] To enable those skilled in the art to better understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0026] Example 1: Isolation and Identification of Strain AH001 (1) Isolation of strains: The strains were isolated by tissue isolation method. The collected healthy plant tissues were cut into small pieces of about 5 mm × 5 mm, surface disinfected with 75% alcohol for 30 s, then soaked in 2% sodium hypochlorite solution for 45 s, then rinsed 3 times with sterile water and drained. The disinfected tissues were placed in a sterile mortar, ground into a homogenate with a small amount of sterile water, and allowed to stand for 10 min. The supernatant was then placed in a 1.5 mL centrifuge tube, centrifuged at 12000 r / min for 3 min, and 2-3 μL was evenly spread on Gao's No. 1 solid culture medium plates and incubated upside down in a constant temperature and dark incubator at 25℃ for 7-14 days.

[0027] (2) Morphological identification: The AH001 strain was streaked onto Gao's No. 1 solid medium and cultured at 25℃ for 7-30 days. The morphology of single colonies was then observed. Figure 1 As shown, a single colony was picked and inoculated into Gao's No. 1 medium (slightly thicker than ordinary medium). A sterile coverslip was inserted at a 45° angle at the streaked position. After 15 days, the bacterial cell structure, spore formation, and conidia morphology were observed under an optical microscope. Figure 2As shown, a single colony was picked and inoculated into Gao's No. 1 liquid medium and cultured for 2 days to obtain bacterial suspension. The culture with an OD value of 0.5-0.8 was aspirated, centrifuged, and the medium was discarded. The bacterial precipitate was collected at the bottom of the tube in the size of a soybean, washed 1-2 times with PBS, and the supernatant was discarded. Pre-cooled 2.5% glutaraldehyde fixative solution (4℃) was slowly added along the tube wall. The hyphae and micromorphology were observed under a scanning electron microscope as shown in the figure. Figure 3 As shown.

[0028] Morphological identification results showed that the colonies of strain AH001 were round or oval with a slight bulge in the middle. The color was initially orange-yellow and moist, turning white later. The texture was dry and powdery. Later, the colonies spread outward in a concentric pattern, gradually enlarging, and the culture medium gradually turned dark yellow.

[0029] (3) Molecular identification: The 16S rDNA sequence obtained from the sequencing of strain AH001 is shown below.

[0030]

[0031] The 16S rDNA gene sequence was homology-matched in the NCBI database and identified as *Streptomyces gracilis*. Streptomyces pratensis Streptomyces aureus AH001 was deposited at the Guangdong Provincial Microbial Culture Collection Center on January 14, 2026, with accession number GDMCC No. 67655, and classified as follows: Streptomyces pratensis This is consistent with the identification results.

[0032] Example 2 Evaluation of the antibacterial effect of Streptomyces agrophytes AH001 and its aseptic fermentation filtrate Plate inoculation: Streptomyces aureus AH001 was streaked onto Gao's No. 1 medium and cultured at 25-28℃.

[0033] Preparation of bacterial culture: Four bacterial discs of 5 mm in size were placed in 100 mL of Gao's No. 1 liquid culture medium and cultured at 28℃ and 180 r / min for 2 days. Figure 4 A).

[0034] Preparation of sterile fermentation filtrate: First, prepare the seed culture (same as the preparation of the bacterial culture). Then, take 1% (v / v) of the seed culture and place it in 100 mL of Gao's No. 1 liquid culture medium. Incubate at 28 ℃ and 180 r / min for 5 days to prepare the fermentation broth. Take an appropriate amount of the fermentation broth into a 50 mL centrifuge tube and centrifuge at 4 ℃ and 6000-8000 r / min for 8-10 min. Take the supernatant into a clean centrifuge tube and filter it through a 0.22 μm microporous membrane to remove bacterial cells, thus obtaining a sterile fermentation filtrate free of bacterial strain cells. Figure 4 B).

[0035] Experimental materials: The pathogen tested in this study was Streptomyces, which was isolated from the tissue of cut flower anthurium disease. The pathogen information is shown in Table 1.

[0036] Table 1 Information on tested pathogenic bacterial strains

[0037] The antibacterial effect of Streptomyces agrophytes AH001 on the pathogen of Anthurium erythrorhizon was evaluated through three sets of experimental designs.

[0038] Experimental Group A: A 5 mm pathogenic bacterial cake was inoculated in the center of a PDA plate, and a 5 mm sterile filter paper was placed on each side of the bacterial cake. 5 μL of Streptomyces aureus AH001 bacterial solution was added to each filter paper, and the plate was incubated at 25℃ for 7 days.

[0039] Experimental Group B: First, place 5 mm sterile filter paper discs on both sides of a PDA plate and add 5 μL of bacterial suspension. After pre-culturing for 2 days, inoculate a 5 mm pathogenic bacterial cake in the center of the plate and continue culturing at 25 ℃ for 7 days. The control group was the same as the treatment group A except that an equal amount of sterile water was added to the filter paper discs.

[0040] Experimental group C: Sterile fermentation filtrate of *Streptomyces simonii* AH001 was added to PDA medium at a volume ratio of 5%. After mixing, plates were prepared, and a 5 mm pathogenic bacterial cake was inoculated in the center of each plate. The plates were incubated at 25°C for 7 days. A control group without the addition of sterile fermentation filtrate was used. The inhibition rate was calculated, and the results are shown in Table 2 below.

[0041] Antibacterial rate (%)

[0042] Table 2. Antimicrobial effects of three different treatments on the biocontrol Streptomyces AH001 fermentation broth against pathogens.

[0043] Experimental results showed that *Streptomyces simonii* AH001 had a significant inhibitory effect on all eight pathogens of cut anthurium. In treatments A and B, *Streptomyces simonii* AH001 showed the most significant inhibitory effect against *Alternaria alternata* (HZ7), with inhibition rates reaching 62.05% and 72.14%, respectively. In treatment C, the inhibitory effect was the most significant, achieving 100% inhibition rates against *Anthracnose bonningensis* and *Cladosporium cladosporium*, and inhibition rates against the other six pathogens were all above 75%, significantly higher than groups A and B. Figure 5 (Table 2).

[0044] Finally, it should be noted that the above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of them. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A strain of *Streptomyces gracilis* AH001 isolated from *Anthurium stenoptera*, characterized in that, Streptomyces cepacia ( Streptomyces pratensis AH001 was deposited at the Guangdong Provincial Center for Microbial Culture Collection on January 14, 2026, with accession number GDMCC No. 67655.

2. The derivative of Streptomyces aureus AH001 according to claim 1, characterized in that, This includes one or more of its live bacteria, inactivated bacterial cells, fermentation broth, exosomes, or metabolites.

3. A microbial agent, characterized in that, It contains the Streptomyces agrarianis AH001 as described in claim 1 or the derivative as described in claim 2.

4. The microbial agent as described in claim 3, characterized in that, The microbial agent contains Streptomyces agrophytes AH001 or its metabolites; preferably, the microbial agent also contains a carrier or excipients.

5. The method for preparing the microbial agent according to claims 3-4, characterized in that, The method includes the step of culturing the Streptomyces aureus AH001 as described in claim 1 and obtaining a culture.

6. The application of *Streptomyces aureus* AH001 as described in claim 1, or the derivative as described in claim 2, or the fungal agent as described in claims 3-4, in antagonizing plant pathogens, wherein the plant pathogen is *Anthracnose boningensis* (…). Colletotrichum boninense ), fruit anthracnose ( Colletotrichum fructicola ), Colloidal anthrax bacteria ( Colletotrichum gloeosporioides ), Siamese anthrax bacteria ( Colletotrichum siamense Fusarium moniliforme ( ) Fusarium proliferatum Fusarium oxysporum ( Fusarium oxysporum ), Alternaria ( Alternaria alternata ) or apical spores ( Acremonium sp.).

7. The application as described in claim 6, characterized in that, The plant mentioned includes anthurium.

8. A product characterized in that, The product contains the Streptomyces aureus AH001 as described in claim 1 or the inoculum as described in claim 2; Preferably, the product has the effect of antagonizing plant pathogens that cause plant diseases, wherein the plant pathogen is *Anthracnose boningensis* (…). Colletotrichum boninense ), fruit anthracnose ( Colletotrichum fructicola ), Colloidal anthrax bacteria ( Colletotrichum gloeosporioides ), Siamese anthrax bacteria ( Colletotrichum siamense Fusarium moniliforme ( ) Fusarium proliferatum Fusarium oxysporum ( Fusarium oxysporum ), Alternaria ( Alternaria alternata ) or apical spores ( Acremonium sp.).

9. A method for preventing and controlling plant diseases, characterized in that, The method includes applying Streptomyces aureus AH001 as described in claim 1, or the fungal agent as described in claim 2, or the product as described in claim 8 to the plant.

10. The method as described in claim 9, characterized in that, The plant pathogen is *Anthracnose boningensis* (Boning anthracnose fungus). Colletotrichum boninense ), fruit anthracnose ( Colletotrichum fructicola ), Colloidal anthrax bacteria ( Colletotrichum gloeosporioides ), Siamese anthrax bacteria ( Colletotrichum siamense Fusarium moniliforme ( ) Fusarium proliferatum Fusarium oxysporum ( Fusarium oxysporum ), Alternaria ( Alternaria alternata ) or apical spores ( Acremonium sp.).

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