Bacillus amyloliquefaciens GX0003936 and application thereof

Abstract

The invention discloses bacillus amyloliquefaciens GX0003936 and application thereof, and belongs to the technical field of microorganism application. The invention discloses bacillus amyloliquefaciens GX0003936 with a broad-spectrum bacteriostatic effect, which is preserved in the China General Microbiological Culture Collection Center on March 26, 2025, and the preservation number of the bacillus amyloliquefaciens GX0003936 is CGMCC (China General Microbiological Culture Collection Center) No.33987. The bacterial strain has broad-spectrum antibacterial activity, can be used for remarkably inhibiting the growth of mycelia of colletotrichum musae and colletotrichum frutescens, and can be used for remarkably reducing the disease incidence rate of tropical fruits such as bananas, mangoes and the like in the postharvest storage period. Meanwhile, the strain shows a good inhibition effect on various plant pathogenic fungi, and has a potential broad-spectrum biocontrol effect.

Description

Technical Field

[0001] The present invention relates to the field of microbial applications, in particular to a Bacillus amyloliquefaciens GX0003936 and applications thereof. Background Art

[0002] Mangoes and bananas are both climacteric fruits. Their respiration rate increases rapidly during the ripening process, causing the fruit to soften quickly and the storage period to be significantly shortened.

[0003] In addition to physiological changes, post-harvest diseases are also important factors affecting fruit quality and shelf life. Among them, anthracnose is the most common and serious. This disease not only affects the appearance of the fruit, but also often causes the flesh to rot, significantly reducing the commercial value. Fruit anthracnose (Colletotrichum fructicola) is the dominant species of mango anthracnose, while banana anthracnose is mostly caused by plantain anthracnose (Colletotrichum musae). These pathogens often exist in a latent form on the surface of the fruit or inside the tissue. Once they encounter suitable environmental conditions (such as high humidity, high temperature, and fruit ripening), they can rapidly infect and spread, leading to an outbreak of the disease. In the early stages of the disease, it often manifests as small brown spots on the fruit surface, which then expand and become sunken, and eventually the flesh rots and deteriorates.

[0004] Chemical preservatives have traditionally been used to extend the shelf life of fruit and reduce postharvest losses. However, long-term use of these chemicals poses risks to human health and the environment and can also induce pathogen resistance, reducing control effectiveness. Therefore, the development of green, safe, and environmentally friendly preservation technologies has become a research hotspot and an industry priority.

[0005] In recent years, microbial volatile organic compounds (VOCs) have garnered widespread attention as a novel preservation strategy. VOCs are a class of low-molecular-weight, volatile, and characteristically odorous organic compounds produced by microorganisms during their metabolism. They possess a variety of bioactive properties, including antimicrobial, antioxidant, resistance-inducing, and plant growth-promoting properties. Studies have shown that microorganisms such as Bacillus can synthesize a variety of VOCs with antibacterial activity, significantly inhibiting a variety of plant pathogens without inducing drug resistance, demonstrating excellent safety and sustainability.

[0006] Therefore, screening microbial strains that efficiently produce antimicrobial VOCs and applying them to the postharvest preservation of mangoes and bananas is a key research direction in the field of microbial preservation. Exploring the types of VOCs produced by these microorganisms and their mechanisms of action will not only facilitate the development of novel biopreservatives, but also provide theoretical support and technical pathways for promoting the green development of the tropical fruit industry. Summary of the Invention

[0007] The present invention aims to provide a strain of Bacillus amyloliquefaciens GX0003936 and its application to solve the problems existing in the above-mentioned prior art. The Bacillus amyloliquefaciens GX0003936 disclosed in the present invention has broad-spectrum antibacterial activity, can significantly inhibit the mycelial growth of Colletotrichum musae and Colletotrichum fructicola, and significantly reduce the incidence of post-harvest diseases of bananas and mangoes. The strain shows good inhibitory effects on various plant pathogenic fungi and has a significant effect on the post-harvest preservation of fruits. The development of this strain provides a new microbial resource for the green prevention and control of post-harvest diseases of tropical fruits, lays a foundation for the research and application of fruit biological preservation and biological control technology, and has good application prospects and promotion value.

[0008] To achieve the above object, the present invention provides the following solutions:

[0009] The present invention provides a Bacillus amyloliquefaciens GX0003936, which is deposited in the General Microbiology Center of the China Culture Collection Administration, the deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, the deposit date is March 26, 2025, and the deposit number is CGMCC No. 33987.

[0010] The present invention also provides the use of the Bacillus amyloliquefaciens GX0003936 in preventing and controlling plant pathogens.

[0011] The present invention also provides the use of the Bacillus amyloliquefaciens GX0003936 in preparing a biocontrol agent for plant pathogens.

[0012] Preferably, the plant pathogens include: Acanthopanax cucurbitae, Neochromis fascicularis, Botrytis cinerea, Fusarium oxysporum, Trichoderma tritici, Colletotrichum eschinoides, Fusarium chlamydosporium, Colletotrichum siamensis, Chlamydosporium syringae, Alternaria alternata, Polytrichomonas oleraceus and Discosporum spp.

[0013] The present invention also provides a biocontrol agent for plant pathogens, wherein the active ingredient of the biocontrol agent comprises the Bacillus amyloliquefaciens GX0003936 or its metabolites or volatile gases.

[0014] Preferably, the plant pathogens include: Acanthopanax cucurbitae, Neochromis fascicularis, Botrytis cinerea, Fusarium oxysporum, Trichoderma tritici, Colletotrichum eschinoides, Fusarium chlamydosporium, Colletotrichum siamensis, Chlamydosporium syringae, Alternaria alternata, Polytrichomonas oleraceus and Discosporum spp.

[0015] The present invention also provides use of the Bacillus amyloliquefaciens GX0003936 in preparing a fruit preservative.

[0016] The present invention also provides a fruit preservative, wherein the effective component of the fruit preservative comprises the Bacillus amyloliquefaciens GX0003936 or its metabolites or volatile gas.

[0017] Preferably, the fruit comprises mango or banana.

[0018] The present invention also provides a method for preventing and controlling fruit anthracnose, comprising the following steps: spraying fruit with a fruit preservative prepared from the Bacillus amyloliquefaciens GX0003936; or inoculating the Bacillus amyloliquefaciens GX0003936 into a culture medium, and allowing the volatile gas of the Bacillus amyloliquefaciens GX0003936 to act on the fruit through a closed fumigation method; the pathogenic bacteria of the fruit anthracnose include Colletotrichum musa or Colletotrichum frutescens.

[0019] The present invention also provides a method for extending the post-harvest shelf life of fruits, comprising the following steps: inoculating the Bacillus amyloliquefaciens GX0003936 into a culture medium, and allowing the volatile gas of the Bacillus amyloliquefaciens GX0003936 to act on the harvested fruits through a closed fumigation method.

[0020] Preferably, the fruit comprises banana or mango.

[0021] The present invention discloses the following technical effects:

[0022] The present invention provides a strain of Bacillus amyloliquefaciens GX0003936 with broad-spectrum antibacterial activity. The strain can significantly inhibit the growth and reproduction of various plant pathogenic fungi, including Aconitum cucurbitae, Neochromis cylindrica, Botrytis cinerea, Fusarium oxysporum, Helminthosporium tritici, Colletotrichum eschinoides, Fusarium chlamydosporum, Colletotrichum siamensis, chestnut blight pathogen, Alternaria alternata, Polytrichomoniasis truncatula of tea, and Discospora spp.

[0023] In addition, this strain and its metabolites and volatile organic compounds (VOCs) can effectively inhibit the infection of anthracnose pathogens on fruits such as bananas and mangoes, significantly reduce the incidence of post-harvest diseases, and effectively extend the shelf life of the fruit.

[0024] Therefore, Bacillus amyloliquefaciens GX0003936 provides a valuable bacterial resource for the research and development of new biological preservatives and growth-promoting agents, and has broad application potential and promotion value. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0026] Figure 1 The morphological observation results of Bacillus amyloliquefaciens GX0003936 are shown in Figure 1. A: colony morphology, B: bacterial morphology under Gram staining optical microscope (100× oil lens);

[0027] Figure 2 This is the phylogenetic tree of Bacillus amyloliquefaciens GX0003936 based on the 16S rDNA gene sequence;

[0028] Figure 3 This is the phylogenetic tree of Bacillus amyloliquefaciens GX0003936 based on the gyrB gene sequence;

[0029] Figure 4 The growth status of Bacillus amyloliquefaciens GX0003936 in different culture media; A: PKO inorganic phosphorus medium; B: Montgena organic phosphorus medium; C: CAS detection medium; D: amylase detection medium; E: nitrogen fixation medium;

[0030] Figure 5 The antifungal activity of Bacillus amyloliquefaciens GX0003936 against different plant pathogenic fungi, including: A: Acanthosporium cucurbitae; B: Neochromis cylindrica; C: Botrytis cinerea; D: Fusarium oxysporum; E: Polytrichomoniasis oleracea; F: Discosporium spp.; G: Helminthosporium truncatum; H: Discosporium eschinomeni; I: Fusarium chlamydosporium; J: Colletotrichum siamensis; K: Chrysophora parasitica; L: Alternaria alternata.

[0031] Figure 6 The diagram shows the inhibitory effect of Bacillus amyloliquefaciens GX0003936 on Colletotrichum musa; A: control group; B: Bacillus amyloliquefaciens GX0003936-treated group;

[0032] Figure 7 The diagram shows the inhibitory effect of Bacillus amyloliquefaciens GX0003936 on Colletotrichum oleraceus; A: control group; B: Bacillus amyloliquefaciens GX0003936-treated group;

[0033] Figure 8 The figure shows the inhibitory effect of volatile gases of Bacillus amyloliquefaciens GX0003936 on Colletotrichum musa; A: control group; B: Bacillus amyloliquefaciens GX0003936 treatment group;

[0034] Figure 9 The figure shows the inhibitory effect of volatile gases of Bacillus amyloliquefaciens GX0003936 on Colletotrichum fuscae; A: control group; B: Bacillus amyloliquefaciens GX0003936 treated group;

[0035] Figure 10 The figure shows the preservation effect of Bacillus amyloliquefaciens GX0003936 on post-harvest bananas; A: control group; B: preservation effect after GX0003936 volatile gas treatment;

[0036] Figure 11 This is a diagram showing the preservation effect of Bacillus amyloliquefaciens GX0003936 on post-harvest mangoes; A: control group; B: preservation effect after GX0003936 volatile gas treatment. DETAILED DESCRIPTION

[0037] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0038] It should be understood that the terms described herein are intended only to describe particular embodiments and are not intended to limit the present invention. In addition, for numerical ranges herein, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. The intermediate value within any stated value or stated range, and each smaller range between any other stated value or intermediate value within the stated range, is also encompassed within the present invention. The upper and lower limits of these smaller ranges may be independently included or excluded within the scope.

[0039] Unless otherwise indicated, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein may also be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of any conflict with any incorporated document, the contents of this specification shall prevail.

[0040] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments described herein without departing from the scope or spirit of the invention. Other embodiments will be apparent to those skilled in the art from the description of the invention. The description and examples are intended to be illustrative only.

[0041] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0042] The experimental materials used in the examples of the present invention come from the following sources:

[0043] Montgena organic phosphorus medium, PKO inorganic phosphorus medium, and CAS detection medium were purchased from Qingdao Haibo Biological Co., Ltd.;

[0044] Amylase assay medium (g / L): tryptone 10.0 g / L, yeast extract 5.0 g / L, NaCl 10.0 g / L, soluble starch 2 g / L, distilled water as solvent;

[0045] Nitrogen-fixing medium (g / L): mannitol 10 g / L, KH2PO4 0.2 g / L, MgSO4·7H2O 0.2 g / L, NaCl 0.2 g / L, CaSO4 0.1 g / L, CaCO3 5 g / L, agar 18 g / L, pH 7.2, distilled water as solvent;

[0046] Choanephora cucurbitarum, Bipolarissorokiniana, Alternaria alternata, Colletorichummusae, Fusarium oxysporum, Botryosphaeria dothidea, Neoscytalidium dimidiatum, Cryphonectria parasitic, Fusarium chlamydosporum, Colletotrichumaeschynomenes, Colletotrichum siamense, Coletotrichum fructicola, Pestalotiopsis theae, and Diaporthediscoidispora are from the Key Laboratory of Microbial Resources and Utilization, College of Ocean and Biotechnology, Guangxi University for Nationalities.

[0047] Unless otherwise specified, materials or reagents whose sources are not specified in the examples of the present invention can be purchased through conventional channels.

[0048] Example 1 Identification of Bacillus amyloliquefaciens GX0003936

[0049] 1. Isolation and morphological observation of strains

[0050] Strain GX0003936 was isolated from the roots of tea plants and is an endophytic fungus. The target strain was isolated and purified using the dilution spread plating method. The strain was streaked onto LB solid medium and incubated at 30°C for 24 hours. The morphological characteristics of individual colonies, such as color, size, and transparency, were observed. Subsequently, individual colonies were selected for Gram staining and observed under an optical microscope.

[0051] The morphological observation results are as follows Figure 1 As shown: Figure 1 As shown in A, the strain forms white, wrinkled, transparent and viscous colonies on LB solid medium; Figure 1 Figure B is a microscopic image after Gram staining. The bacteria are rod-shaped and the Gram staining result is blue-purple, indicating that it is a Gram-positive bacillus.

[0052] 2. Physiological and biochemical characteristics

[0053] Referring to the methods in the Manual of Identification of Common Bacteria Systems and the Bergey's Manual of Bacteria Identification, the GX0003936 strain was tested for multiple physiological and biochemical indices, including lecithinase activity, starch hydrolysis, gelatin liquefaction, and methyl red test.

[0054] The results of physiological and biochemical characterization are shown in Table 1. As shown in Table 1, the strain was motile and negative for citrate utilization, methyl red, and lecithinase tests. However, it was positive for the Voges-Proskauer (VP) test, starch hydrolysis, gelatin liquefaction, and catalase tests. These characteristics provided important evidence for subsequent identification and classification of the strain.

[0055] Table 1 Results of physiological and biochemical characteristics analysis

[0056]

[0057] 3. Molecular biology identification

[0058] After activating strain GX0003936, total DNA was extracted and used as a template for PCR amplification of the 16S rRNA gene and gyrB gene of strain GX0003936 using universal bacterial primers 27F and 1492R and primers UP-1 and UP-2r for the housekeeping gene gyrB, respectively. The nucleotide sequences of primers 27F, 1492R and primers UP-1 and UP-2r are as follows:

[0059] 27F: 5'-AGAGTTTGATCCTGGCTCAG-3' (SEQ ID NO. 1);

[0060] 1492R: 5'-TACGGCTACCTTGTTACGAGTT-3' (SEQ ID NO. 2);

[0061] UP-1: 5'-ATTTGGCGCTGGCGGTTAT-3' (SEQ ID NO. 3);

[0062] UP-2r: 5'-GGTTTCGGCTGGGCTGGTA-3' (SEQ ID NO. 4).

[0063] PCR amplification system (50 μL): 1 μL template, 22 μL ddH2O, 25 μL 2× Taq PCR Mix, 1 μL each of upstream and downstream primers.

[0064] PCR amplification program: pre-denaturation at 95°C for 5 min; denaturation at 95°C for 1 min, annealing at 55°C for 1 min, extension at 72°C for 2 min, 32 cycles; extension at 72°C for 10 min; storage at 4°C.

[0065] The amplified products were sent to Aoke Biotechnology (Wuhan) Co., Ltd. for sequencing. The sequences were compared with those in the NCBI database for homology, and a phylogenetic tree was constructed using MEGA 12.0 software.

[0066] The molecular biological identification results are as follows Figure 2 and Figure 3 As shown, the 16S rRNA gene sequence of strain GX0003936 was homologously analyzed with existing data in the NCBI database and a phylogenetic tree was constructed. It was found that the 16S rRNA sequence of strain GX0003936 had an 88% similarity with Bacillus amyloliquefaciens. The gyrB sequence alignment results showed that it had the highest similarity with Bacillus amyloliquefaciens, and the phylogenetic tree clustered in the same branch.

[0067] Based on the strain's morphological characteristics, physiological and biochemical properties, and molecular biological identification results, the strain was identified as Bacillus amyloliquefaciens. Bacillus amyloliquefaciens GX0003936 was deposited on March 26, 2025, at the General Microbiology Center of the China Culture Collection Administration (CMC), Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No. 33987.

[0068] Example 2 Determination of Growth Promotion and Enzyme Production Ability of Bacillus amyloliquefaciens GX0003936

[0069] Preparation of Bacillus amyloliquefaciens GX0003936 culture medium: inoculate Bacillus amyloliquefaciens GX0003936 into 50 mL LB liquid medium and culture at 30°C and 180 rpm until OD 600 ≈1.0.

[0070] Preparation of single colonies of Bacillus amyloliquefaciens GX0003936: Bacillus amyloliquefaciens GX0003936 was inoculated on LB solid medium and cultured in three zones for 1 day.

[0071] 5 μL of Bacillus amyloliquefaciens GX0003936 culture solution (OD 600 ≈1.0) in the center of Montgena organic phosphorus medium, PKO inorganic phosphorus medium, CAS detection medium, and amylase detection medium, and cultured at 30°C for 5 days. The amylase detection medium was developed with iodine solution.

[0072] A single colony of GX0003936 was picked and inoculated into nitrogen-fixing medium in three stripes, and cultured at 30°C for 3 days to observe whether it could grow on the nitrogen-fixing medium.

[0073] The qualitative test results of the growth-promoting function and enzyme production characteristics of Bacillus amyloliquefaciens GX0003936 are as follows Figure 4 As shown, Bacillus amyloliquefaciens GX0003936 produced clear zones on PKO inorganic phosphate medium, Montgena organic phosphate medium, and amylase assay medium; it produced an orange-yellow halo on CAS assay medium; and it grew normally on nitrogen-fixing medium. These results demonstrate that Bacillus amyloliquefaciens GX0003936 has the ability to degrade inorganic and organic phosphates, produce siderophores, fix nitrogen, and produce amylase.

[0074] Example 3 Detection of antibacterial ability of Bacillus amyloliquefaciens GX0003936

[0075] The plate standoff assay was used to determine the inhibitory activity of Bacillus amyloliquefaciens GX0003936 against 12 common plant pathogens: Amyloliquefaciens, Neofaunal Arthrospora, Botrytis cinerea, Fusarium oxysporum, Helminthosporium tritici, Colletotrichum eschinophyllum, Fusarium chlamydosporum, Colletotrichum siamensis, Psoralea corylifolia, Alternaria alternata, Polytrichomonas truncatula, and Discosporium spp. The experimental procedure involved inoculating a 6 mm diameter pathogen cake in the center of a PDA plate. Then, 5 μL of the Bacillus amyloliquefaciens GX0003936 culture (prepared in Example 2) was inoculated 21 mm from the center of the plate. PDA medium inoculated with the pathogen alone served as a control. The plates were incubated at 28°C for 5 days. The antibacterial activity of the strain was observed, and the inhibition rate was calculated.

[0076] Inhibition rate (%) = (colony radius of the control group - colony radius of the treated group) / colony radius of the control group × 100%.

[0077] The antibacterial effect test results of Bacillus amyloliquefaciens GX0003936 are as follows Figure 5 As shown in the figure, the upper layer shows the colony morphology of the control group, and the lower layer shows the colony morphology of the treatment group. The results showed that Bacillus amyloliquefaciens GX0003936 had an inhibitory effect on 12 common plant pathogens, including Trichoderma cucurbitae, Neochromis cylindrica, Botrytis cinerea, Fusarium oxysporum, Trichoderma tritici, Colletotrichum eschinophyllum, Fusarium chlamydophorum, Colletotrichum siamensis, Psoralea corylifolia, Alternaria alternata, Polytrichomoniasis truncatula, and Discosporium spp. The inhibition rates are shown in Table 2, showing that the inhibition rate of Bacillus amyloliquefaciens GX0003936 against 10 pathogenic fungi was greater than 62%.

[0078] Table 2 Antibacterial spectrum determination results of Bacillus amyloliquefaciens GX0003936

[0079]

[0080]

[0081] Example 4 Detection of the antibacterial activity of Bacillus amyloliquefaciens GX0003936 against Colletotrichum musa and Colletotrichum oleraceus

[0082] 1. Inhibitory effect of Bacillus amyloliquefaciens GX0003936 on Colletotrichum musa

[0083] The plate standoff method was used to test the inhibitory effect of Bacillus amyloliquefaciens GX0003936 on Colletotrichum musa. The experimental process was the same as that in Example 3.

[0084] The inhibitory effect of Bacillus amyloliquefaciens GX0003936 is as follows Figure 6 As shown, the results showed that Bacillus amyloliquefaciens GX0003936 had a significant inhibitory effect on Colletotrichum musa, with an inhibition rate (%) of 89.73±1.33%.

[0085] 2. Inhibitory effect of Bacillus amyloliquefaciens GX0003936 on Colletotrichum oleraceus

[0086] The plate standoff method was used to test the inhibitory effect of Bacillus amyloliquefaciens GX0003936 on Colletotrichum citrinum. The experimental process was the same as that in Example 3.

[0087] The inhibitory effect of Bacillus amyloliquefaciens GX0003936 is as follows Figure 7 As shown, the results showed that Bacillus amyloliquefaciens GX0003936 had a significant inhibitory effect on Colletotrichum oleraceus, with an inhibition rate (%) of 86.40±1.21%.

[0088] 3. Antibacterial activity of volatile gases from Bacillus amyloliquefaciens GX0003936 against Colletotrichum musa

[0089] The inhibitory effect of volatile gases produced by Bacillus amyloliquefaciens GX0003936 on Colletotrichum musa was determined using the double-plate method. Bacillus amyloliquefaciens GX0003936 was inoculated at 2% in 50 mL of LB liquid medium and incubated at 30°C with shaking at 180 rpm for 24 hours. 100 μL of the fermentation broth was evenly spread on a solid LB plate. A 6 mm diameter colony of Colletotrichum musa was placed in the center of a PDA plate. The two plates were sealed and incubated at 28°C for 5 days. A control plate containing a blank medium was used instead of the Bacillus amyloliquefaciens GX0003936 plate.

[0090] Antibacterial activity of volatile gases from Bacillus amyloliquefaciens GX0003936 against Colletotrichum musa Figure 8 As shown, the results showed that the volatile gas produced by Bacillus amyloliquefaciens GX0003936 can also significantly inhibit the growth and reproduction of banana anthracnose.

[0091] 4. Antibacterial activity of volatile gases from Bacillus amyloliquefaciens GX0003936 against Colletotrichum oleraceus

[0092] The inhibitory effect of volatile gases produced by Bacillus amyloliquefaciens GX0003936 on Colletotrichum citrinum was determined using the double-plate method. Bacillus amyloliquefaciens GX0003936 was inoculated at 2% in 50 mL of LB liquid medium and incubated at 30°C with shaking at 180 rpm for 24 hours. 100 μL of the fermentation broth was evenly spread on a solid LB plate. A 6 mm diameter Colletotrichum citrinum cell was placed in the center of a PDA plate. The two plates were sealed and incubated at 28°C for 5 days. A control plate containing a blank medium was used instead of the Bacillus amyloliquefaciens GX0003936 plate.

[0093] Antibacterial activity of volatile gases from Bacillus amyloliquefaciens GX0003936 against Colletotrichum syringae Figure 9 As shown, the results showed that the volatile gas produced by Bacillus amyloliquefaciens GX0003936 can also significantly inhibit the growth and reproduction of fruit anthracnose.

[0094] Example 5: Effect of Bacillus amyloliquefaciens GX0003936 on postharvest banana anthracnose and mango anthracnose

[0095] 1. Wipe the surface of 80% ripe bananas with 2% sodium hypochlorite solution, rinse repeatedly with sterile water, and dry them for later use. Use the volatile gas of Bacillus amyloliquefaciens GX0003936 to preserve the freshness of post-harvest bananas:

[0096] Volatile gas treatment: Take 100 μL of Bacillus amyloliquefaciens GX0003936 culture medium and evenly spread it on the LB solid plate. Place it in a transparent box together with the bananas after the above disinfection treatment. Store it in a sealed box at a constant temperature of 25°C and observe the bananas periodically.

[0097] The experiment used disinfected bananas as the control group.

[0098] The results are as follows Figure 10 As shown in the results, the control group began to turn yellow on the 7th day, lesions began to appear on the 14th day, and the lesion area was greater than 50% on the 22nd day. The volatile gas treatment group began to turn yellow on the 22nd day, the fruit lesion area was less than 10%, and the fruit was hard overall and still had a banana aroma. This shows that the volatile gas of GX0003936 has a good preservation effect on post-harvest bananas and can delay the time of banana ripening and decay.

[0099] 2. Wipe the surface of 50% ripe mangoes with 2% sodium hypochlorite solution, rinse repeatedly with sterile water, and dry for later use. Use the volatile gas of Bacillus amyloliquefaciens GX0003936 to preserve the post-harvest mangoes:

[0100] Volatile gas treatment: Take 100 μL of Bacillus amyloliquefaciens GX0003936 culture medium and evenly spread it on an LB solid plate. Place it in a transparent box together with the mangoes after the above disinfection treatment. Store it in a sealed box at a constant temperature of 25°C and observe the condition of the mangoes periodically.

[0101] The mangoes that had been disinfected were used as the control group.

[0102] The results are as follows Figure 11 As shown in the results, the control group began to develop lesions on the 6th day, the lesion area was greater than 70% on the 9th day, and the mangoes had become soft. On the 12th day, the lesion area was greater than 90%, while the volatile gas treatment group developed lesions on the 9th day, and the mangoes were harder on the 12th day. This shows that the volatile gas of GX0003936 has a good preservation effect on post-harvest mangoes and can delay the time of mango ripening and decay.

[0103] The embodiments described above are merely descriptions of preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Without departing from the spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by persons skilled in the art should fall within the scope of protection defined by the claims of the present invention.

Claims

1. A strain of Bacillus amyloliquefaciens GX0003936, characterized in that: It is deposited in the China General Microorganism Collection Center, the deposit address is No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, the deposit date is March 26, 2025, and the deposit number is CGMCC No.33987.

2. The use of Bacillus amyloliquefaciens GX0003936 in preventing and controlling plant pathogens according to claim 1, characterized in that: The plant pathogens include: Choanephora cucurbitarum, Neoscytalidium dimidiatum, Botryosphaeria dothidea, Fusarium oxysporum, Pestalotiopsis theae, Diaporthediscoidispora, Bipolaris sorokiniana, Colletotrichum aeschynomenes, Fusarium chlamydosporum, Colletotrichum siamense, Cryphonectria parasitic, and Alternaria alternata.

3. The use of Bacillus amyloliquefaciens GX0003936 according to claim 1 in preparing a biocontrol agent for plant pathogens, characterized in that: The plant pathogens include: Choanephora cucurbitarum, Neoscytalidium dimidiatum, Botryosphaeria dothidea, Fusarium oxysporum, Pestalotiopsis theae, Diaporthe discoidispora, Bipolaris sorokiniana, Colletotrichum aeschynomenes, Fusarium chlamydosporum, Colletotrichum siamense, Cryphonectria parasitic, and Alternaria alternata.

4. A plant pathogen biocontrol agent, characterized in that: The active ingredient of the biocontrol agent comprises the Bacillus amyloliquefaciens GX0003936 or its metabolites or volatile gases according to claim 1.

5. The plant pathogen biocontrol agent according to claim 4, wherein The plant pathogens include: Choanephora cucurbitarum, Neoscytalidium dimidiatum, Botryosphaeria dothidea, Fusarium oxysporum, Pestalotiopsis theae, Diaporthe discoidispora, Bipolaris sorokiniana, Colletotrichum aeschynomenes, Fusarium chlamydosporum, Colletotrichum siamense, Cryphonectria parasitic, and Alternaria alternata.

6. Use of the Bacillus amyloliquefaciens GX0003936 according to claim 1 in preparing a fruit preservative.

7. A fruit preservative, characterized in that: The effective ingredient of the fruit preservative comprises the Bacillus amyloliquefaciens GX0003936 or its metabolites or volatile gases according to claim 1.

8. A fruit preservative according to claim 7, characterized in that: The fruit includes mango or banana.

9. A method for preventing and treating fruit anthracnose, characterized in that: The method comprises the following steps: spraying fruits with the fruit preservative prepared by the Bacillus amyloliquefaciens GX0003936 according to claim 1; or inoculating the Bacillus amyloliquefaciens GX0003936 into a culture medium, and allowing the volatile gas of the Bacillus amyloliquefaciens GX0003936 to act on the fruit through a closed fumigation method; The pathogens of fruit anthracnose include Colletotrichum musa or Colletotrichum oleraceus.

10. A method for extending the shelf life of fruits after harvest, characterized in that: The method comprises the following steps: inoculating the Bacillus amyloliquefaciens GX0003936 according to claim 1 into a culture medium, and allowing the volatile gas of the Bacillus amyloliquefaciens GX0003936 to act on the harvested fruit through a closed fumigation method.

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