Endophytic fungus bsas17 and application thereof

Abstract

The application discloses an endophytic fungus Bsas17 and application thereof, and belongs to the technical field of microorganisms. Bifusisporella The endophytic fungus Bsas17 is classified as Bifusisporella Endoxylaria sp. Bsas17 and is preserved in the China Center for Type Culture Collection with a preservation number of CCTCC NO: M 20252568. According to a comprehensive multi-gene joint phylogenetic analysis and morphological characteristics, the strain Bsas17 is identified as a new species of the genus Endoxylaria. The endophytic fungus can colonize plant root tissues and significantly improve the growth performance of plants by interacting with the plants. Specifically, the endophytic fungus Bsas17 is co-cultured with rice, tomatoes or coriander, and has a growth promoting effect on the plants. Therefore, the endophytic fungus Bsas17 has application values in guaranteeing the health of plants and promoting the growth of plants.

Description

Technical Field

[0001] This invention relates to the field of microbial technology, specifically to an endophytic fungus Bsas17 and its application in promoting plant growth. Background Technology

[0002] In traditional agricultural production models, the overuse of chemical fertilizers and pesticides has led to a series of serious problems, such as: damage to soil structure and decline in fertility, intensified eutrophication of water bodies, continuous reduction in biodiversity, continuous upgrading of pesticide resistance in pests and diseases, excessive pesticide residues in agricultural products, and the spread of heavy metal pollution. These problems are seriously threatening the balance of the ecosystem and human health and safety.

[0003] Plant endophytic fungi are a unique biological resource, specifically referring to fungal groups that colonize the interior of healthy plant tissues at some or all stages of their life cycle without causing obvious disease symptoms in the host. Studies have shown that endophytic fungi possess multiple ecological functions: firstly, they significantly enhance plant nutrient absorption efficiency through physiological activities such as nitrogen fixation, phosphorus solubilization, and potassium solubilization; secondly, they can synthesize plant growth regulators such as indoleacetic acid and gibberellins, directly promoting the growth and development of the host plant; thirdly, they effectively control various pests and diseases by inducing systemic resistance in plants or secreting secondary metabolites such as antibiotics and alkaloids; and fourthly, through molecular mechanisms such as regulating cell osmotic pressure and enhancing the activity of antioxidant enzyme systems, they significantly improve the plant's adaptability to abiotic stresses such as drought, salinity, heavy metal pollution, and extreme temperatures. These combined effects provide an effective way to reduce the application of chemical fertilizers and pesticides.

[0004] In recent years, the application value of endophytic fungi in promoting plant growth has received increasing attention. For example, patent document CN101486970A discloses a strain of the family Megalocerosidae (…). Magnaporthaceae ), genus *Pyrtomyces* Phialophora Endophytic fungal strain R5-6-1, when symbiotic with rice, can significantly promote rice growth and increase biomass. Patent document CN113388526A discloses a strain of the genus *Phyllostachys*. Phialophora Endophytic fungus FOR20, when colonized in the root tissue of rice, can significantly improve the quality of rice seedlings and increase rice yield.

[0005] It is evident that utilizing the growth-promoting properties of endophytic fungi to develop bio-fertilizers for application in various crops has profound theoretical value and practical significance for promoting green agricultural development and ensuring food security. Therefore, it is necessary to screen for more strains with superior performance to enrich microbial strain resources.

[0006] Bifusisporella The genus is Rejane MF Silva, which was included in the family Megalocerosidae in 2019. MagnaporthaceaeEstablished within, its model is Bifusisporella sorghi It is an endophytic fungus isolated from healthy leaves of Brazilian sorghum (Rejane MF Silva, et al.). Bifusisporella sorghi gen. et sp.nov. ( Magnaporthaceae (to accommodate an endophytic fungus from Brazil. Mycological Progress, 2019, 18: 847-854.). Currently, research on the function of this genus is still very limited. Summary of the Invention

[0007] The purpose of this invention is to provide a new strain of endophytic fungi for developing bio-fertilizers, which promote crop growth and improve crop yield and quality through symbiotic interaction with crop plants.

[0008] To achieve the above objectives, the present invention adopts the following technical solution: This invention isolated a plant belonging to the species from the root system of moso bamboo. Bifusisporella A novel endophytic fungal strain, Bsas17, was discovered. Its main biological characteristics are as follows: after 7 days of growth on PDA plates at 25°C, the colony diameter was 6 cm; the colonies were round, white, with a raised center and finely hair-like edges; aerial hyphae were underdeveloped, creeping along the culture medium surface; the hyphae were transparent or white; under a scanning electron microscope, the hyphae appeared tubular, clearly septate, and relatively smooth, with distinctive constrictions or swellings visible at some branching points. The ITS sequence of this endophytic fungus is shown in SEQ ID NO.1, the LSU sequence in SEQ ID NO.2, the RPB1 sequence in SEQ ID NO.3, and the SSU sequence in SEQ ID NO.4.

[0009] Based on a comprehensive multi-gene phylogenetic analysis and morphological characteristics, strain Bsas17 was identified as... Bifusisporella A new species of the genus, belonging to the kingdom Fungi ( Fungi Ascomycota ( Ascomycota ), Discotyledon ( Pezizomycotina ), class of fecal scabies ( Sordariomycetes ), Giant Crustacea ( Magnaporthales ), Giant Seashell Family ( Magnaporthaceae ), genus *Clostridium* ( Bifusisporella ).

[0010] Therefore, strain Bsas17 was classified and named Bifusisporellasp. Bsas17. On November 17, 2025, strain Bsas17 was deposited at the China Center for Type Culture Collection (CCTCC), Wuhan University, Wuhan, China, with accession number CCTCC NO: M 20252568.

[0011] Furthermore, the culture conditions for the endophytic fungus strain Bsas17 were as follows: the endophytic fungus Bsas17 was inoculated onto potato dextrose agar medium and cultured in the dark at 22-25°C.

[0012] This invention provides a formulation of the endophytic fungus Bsas17, which is prepared into a liquid microbial fertilizer. The preparation method of the liquid microbial fertilizer includes: inoculating the endophytic fungus Bsas17 into PDB liquid culture medium, culturing until the culture medium is filled with mycelia, and obtaining the liquid microbial fertilizer.

[0013] The composition of the PDB liquid culture medium is: 5 g potato extract powder, 10 g peptone, and 15 g glucose per 1000 mL.

[0014] The liquid microbial fertilizer can be applied as follows: the liquid microbial fertilizer is mixed into the nutrient soil, and the germinated crop seeds are sown in the nutrient soil containing the endophytic fungus Bsas17. During the seedling process, the endophytic fungus Bsas17 colonizes the roots of the seedlings.

[0015] This invention has found that colonizing the endophytic fungus Bsas17 in the root tissue of crops significantly promotes crop growth. The interaction between the endophytic fungus Bsas17 and crops can significantly enhance crop growth performance.

[0016] As a specific embodiment of the present invention, the present invention provides the application of the endophytic fungus Bsas17 in promoting rice growth, the application including: colonizing the endophytic fungus Bsas17 in the root tissue of rice.

[0017] This invention demonstrates that colonizing the endophytic fungus Bsas17 in rice root tissue can significantly promote rice plant growth. Specifically, the indicators of rice growth include plant fresh weight and stem width.

[0018] Furthermore, the application includes: inoculating the endophytic fungus Bsas17 into PDB liquid culture medium, culturing until the culture medium is covered with mycelia to obtain liquid microbial fertilizer; then mixing the liquid microbial fertilizer into a seedling substrate; and then sowing the germinated rice seeds into the seedling substrate for co-cultivation. Co-culturing the Bsas17 strain with rice in the form of microbial fertilizer allows the Bsas17 strain to colonize the roots of rice seedlings, which can enhance the quality of rice seedlings.

[0019] Furthermore, the co-culture conditions are: 22-25℃, 16 h light / 8 h dark culture.

[0020] As another specific embodiment of the present invention, the present invention provides the application of the endophytic fungus Bsas17 in promoting tomato growth, the application including: co-culturing the endophytic fungus Bsas17 with tomato plants.

[0021] This invention has discovered that co-culturing the endophytic fungus Bsas17 with tomatoes can significantly improve the growth performance of tomatoes through interaction. Specifically, the indicators of tomato growth include plant height, leaf width, and the number of flowers.

[0022] Furthermore, the application includes: after tomato seeds germinate, they are sown in a seedling substrate containing the endophytic fungus Bsas17 for co-culture. The Bsas17 strain is co-cultured with tomatoes to colonize the roots of tomato seedlings, thereby promoting tomato plant growth.

[0023] Furthermore, the co-culture conditions are: 22-25℃, 16 h light / 8 h dark culture.

[0024] As another specific embodiment of the present invention, the present invention provides the application of the endophytic fungus Bsas17 in promoting the growth of coriander, the application including: co-culturing the endophytic fungus Bsas17 with coriander plants.

[0025] This invention has discovered that co-culturing the endophytic fungus Bsas17 with coriander plants can significantly improve the growth performance of coriander through interaction. Specifically, the indicators of coriander growth include: plant fresh weight and plant height.

[0026] Furthermore, the application includes: after cracking open the protective shell of coriander seeds, they are sown in a seedling substrate containing the endophytic fungus Bsas17 for co-cultivation. The Bsas17 strain is co-cultured with coriander to colonize the roots of coriander seedlings, thereby promoting plant growth.

[0027] Furthermore, the co-culture conditions are: 22-25℃, 16 h light / 8 h dark culture.

[0028] The beneficial effects of this invention are as follows: This invention provides a new Bifusisporella Based on a comprehensive analysis of multiple genes, phylogenetic characteristics, and morphological features, strain Bsas17 was identified as an endophytic fungus. BifusisporellaThis is a new species. This endophytic fungus can colonize plant root tissues and significantly improve plant growth performance through interaction. Specifically, co-culturing the endophytic fungus Bsas17 with rice, tomato, or coriander promotes plant growth. Therefore, the endophytic fungus Bsas17 has application value in ensuring plant health and promoting plant growth. Attached Figure Description

[0029] Figure 1 Morphological characteristics of strain Bsas17. A represents colony morphology; B represents hyphae morphology under a scanning microscope.

[0030] Figure 2 This is a multi-gene phylogenetic tree of strain Bsas17.

[0031] Figure 3 This shows the colonization of strain Bsas17 on rice roots. The top row is the control group without inoculation with strain Bsas17; the bottom row is the experimental group inoculated with strain Bsas17.

[0032] Figure 4 The effects of strain Bsas17 on the growth of potted rice are shown in the figure above, which compares the overall potted rice plants, while the figure below compares individual rice plants. CK represents the control group without inoculation with strain Bsas17, and Bsas17 represents the experimental group inoculated with strain Bsas17.

[0033] Figure 5 for Figure 4 Statistical results of the effects of strain Bsas17 on the growth indicators of potted rice, corresponding to stem width and fresh weight. CK represents the control group without strain Bsas17, and Bsas17 represents the experimental group inoculated with strain Bsas17. Bar charts represent mean ± standard deviation, n=9. Significant differences were determined using Tukey's method with multiple comparisons in one-way ANOVA; **** indicates p < 0.0001 compared to the control group.

[0034] Figure 6 The effect of strain Bsas17 on the growth of potted tomato seedlings was investigated. CK represents the control group without inoculation with strain Bsas17, and Bsas17 represents the experimental group inoculated with strain Bsas17.

[0035] Figure 7 for Figure 6The statistical results of the effects of strain Bsas17 on the growth indicators of potted tomatoes are shown, corresponding to plant height, leaf width, and number of flowers. CK represents the control group without strain Bsas17, and Bsas17 represents the experimental group inoculated with strain Bsas17. The bar chart represents the mean ± standard deviation, n=9. Significant differences were determined using Tukey's method with one-way ANOVA for multiple comparisons. * indicates p < 0.05 compared to the control group, and ** indicates p < 0.01 compared to the control group.

[0036] Figure 8 The effect of strain Bsas17 on the growth of coriander is shown in the figure below. The top figure compares the overall potted rice plants, and the bottom figure compares the individual coriander plants. CK represents the control group without inoculation with strain Bsas17, and Bsas17 represents the experimental group inoculated with strain Bsas17.

[0037] Figure 9 for Figure 8 The statistical results of the effects of strain Bsas17 on the growth indicators of potted coriander are shown, corresponding to single plant fresh weight, total weight, and plant height. CK represents the control group without strain Bsas17, and Bsas17 represents the experimental group inoculated with strain Bsas17. The bar chart represents the mean ± standard deviation, n=9. Significant differences were determined using Tukey's method with one-way ANOVA for multiple comparisons. ** indicates p < 0.01 compared to the control group, and **** indicates p < 0.0001 compared to the control group. Detailed Implementation

[0038] The present invention will be further described below with reference to specific embodiments. These embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Any modifications or substitutions made to the methods, steps, or conditions of the present invention without departing from the spirit and essence of the invention are within the scope of the invention.

[0039] Unless otherwise specified, the experimental methods used in the following examples are conventional methods; the materials and reagents used are commercially available unless otherwise specified.

[0040] Example 1: Isolation, purification and identification of endophytic fungus strain Bsas17 I. Strains Isolation and Purification Strain Bsas17 was isolated from the roots of wild moso bamboo collected in Hangzhou. The specific method was as follows: the roots of wild moso bamboo were continuously rinsed with tap water to carefully remove soil particles and appendages. Healthy root tissue was selected and surface disinfected, first with 1% sodium hypochlorite for 20 min, then rinsed four times with sterile deionized water. The root tissue was then cut into 0.5 cm segments and placed in 2% malt extract agar (MEA, malt extract agar, OXOID; 50 mg / L chloramphenicol was added to inhibit the growth of endophytic bacteria) and incubated in the dark at 25°C. Hyphae were observed every 24 h. Endophytic fungal hyphae growing from the edges of the tissue cuts were carefully picked out with a toothpick, transferred to fresh PDA medium for purification, and the strain was recorded as Bsas17.

[0041] PDA medium: Each liter contains 20 g glucose, 200 g potato, and 15 g agar. Weigh the required amount of potato according to the volume of the medium to be prepared, boil it in water, mash and dissolve it, filter it, add glucose and agar, and autoclave at 121°C for 20 min.

[0042] II. Strain Identification 1. Morphological identification After isolation and purification, strain Bsas17 was inoculated onto PDA medium and cultured at 25°C for 7 days. A small amount of bacterial cells was picked up with a needle, prepared onto a glass slide, and observed and measured under a microscope.

[0043] like Figure 1 As shown, its morphological characteristics are as follows: strain Bsas17 grows rapidly on PDA plates, with a colony diameter of approximately 6 cm after 7 days of growth at 25°C; this strain forms white, round colonies with a raised center and finely fluffy edges on PDA medium. Aerial hyphae are underdeveloped, creeping along the surface of the medium, and the colonies are white, with transparent or white hyphae. Under a scanning electron microscope, the hyphae appear tubular, with distinct septa and a relatively smooth surface, and special constricted or swollen structures are visible at some branching points.

[0044] 2. Molecular identification (1) DNA extraction ① After culturing Bsas17 strain on PDA plates at 25℃ for 7 days, scrape the mycelium from the plates with a toothpick and place it into a sterilized centrifuge tube containing 300 μL of extraction buffer (1 M KCl, 100 mM Tris HCl, 10 mM EDTA, pH=8.0). ② Grind with an electric grinder and shake vigorously for 2 minutes; ③ Centrifuge at 10,000 rpm for 10 min; ④ Aspirate the supernatant and transfer it to another new centrifuge tube, discarding the precipitate; ⑤ Add an equal volume of isopropanol (analytical grade) to the supernatant, gently invert and mix several times, then centrifuge at 12000 rpm for 10 min to precipitate nucleic acids; ⑥ Gently pour off the supernatant and invert the centrifuge tube containing the precipitate onto absorbent paper to drain the water; ⑦ Add 300 μL of 70% ethanol, gently invert and mix several times, then centrifuge at 12000 rpm for 2 min; ⑧ Gently pour off the supernatant and repeat step ⑦ once; ⑨ Invert the centrifuge tubes onto absorbent paper to drain excess water, and place them at 37°C for 15 minutes to allow the ethanol to evaporate completely; ⑩ The precipitate was resuspended in 50 μL ddH2O to obtain Bsas17 genomic DNA at a concentration of 30 ng / μL.

[0045] (2) PCR amplification of conserved fungal sequences PCR amplification was performed in a 20 μL reaction system containing: 1 μL each of forward and reverse primers, 10 μL of Green TaqMix enzyme, 7 μL of ddH2O, and 1 μL of template DNA.

[0046] The upstream primer ITS-F sequence is: 5'-TCCGTAGGTGAACCTGCGG-3'. The downstream primer ITS-R sequence is: 5'-TCCTCCGCTTATTGATATGC-3'.

[0047] The upstream primer LSU-F sequence is: 5'-GTACCCGCTGAACTTAAGC-3'. The downstream primer LSU-R sequence is: 5'-TCCTGAGGGAAACTTCG-3'.

[0048] The upstream primer RPB1-F sequence is: 5'-GARTGYCCDGGDCAYTTYGG-3'. The downstream primer RPB1-R sequence is: 5'-CCNGCDATNTCRTTRTCCATRTA-3'.

[0049] The upstream primer SSU-F sequence is: 5'-GTAGTCATATGCTTGTCTC-3'. The downstream primer SSU-R sequence is: 5'-CTTCCGTCAATTCCTTTAAG-3'.

[0050] PCR amplification was performed on a Langqi MG96G PCR instrument. Reaction conditions: 94℃ pre-denaturation for 2 min; then 35 cycles including: 94℃ denaturation for 30 sec, 55℃ annealing for 40 sec, 72℃ extension for 1 min; and a final extension at 72℃ for 10 min.

[0051] (3) Gene sequencing and sequence analysis The purified and recovered target DNA fragments, after electrophoresis detection, were sent to the Hangzhou Sequencing Department of Youkang Biotechnology Co., Ltd. for sequencing. After rigorous verification, the sequencing results showed that the ITS sequence was a 503 bp DNA fragment sequence as shown in SEQ ID NO.1; the LSU sequence was a 906 bp DNA fragment sequence as shown in SEQ ID NO.2; the RPB1 sequence was an 811 bp DNA fragment sequence as shown in SEQ ID NO.3; and the SSU sequence was a 1037 bp DNA fragment sequence as shown in SEQ ID NO.4.

[0052] On the NCBI website, the ITS sequence of strain Bsas17 was analyzed and compared with the GenBank database using the BLAST algorithm. The BLAST comparison results showed that the ITS sequence of this strain was similar to that of accession number NR_200407.1. Bifusisporella graminicola The coverage rate of the CGMCC 3.24891 type strain was 85%; compared with the accession number MK606156.1 Bifusisporella sorghi The similarity to the URM 7864 type strain was 97.56%; it was similar to the order Megaloceros (…). Magnaporthales ) some unnamed strains (such as Magnaporthales The similarity to sp. 1 MJ-L-2014 was as high as 98.32%. Preliminary sequence alignment confirmed that strain Bsas17 belongs to the genus level. Bifusisporella .

[0053] To further clarify the taxonomic position of strain Bsas17, the family Megalocerosidae (Giant's Shell Family) was searched in the NCBI database. Magnaporthaceae )middle Bifusisporella , Gaeumannomyces , Magnaporthiopsis , Omnidemptus Multiple gene sequence searches were performed on several closely related genera to construct a multigene phylogenetic tree (ML tree) containing Bsas17 and its closely related genera, such as... Figure 2 As shown in the phylogenetic tree, strain Bsas17 is related to... Bifusisporella graminicola , Bifusisporella sorghi They cluster together into a branch with high support (Bootstrap > 90%), clearly categorized as... Bifusisporella .

[0054] In a multigene phylogenetic tree, Bsas17 and Bifusisporella graminicola The fact that this strain has formed parallel and independent branches with known species indicates a significant evolutionary difference at the genomic level. Combined multi-gene phylogenetic analysis and morphological characteristics support the identification of strain Bsas17 as… Bifusisporella A new species ( Bifusisporella zhejiangensis sp. nov., belongs to the kingdom Fungi ( Fungi Ascomycota ( Ascomycota ), class of fecal scabies ( Sordariomycetes ), Giant Crustacea ( Magnaporthales ), Giant Seashell Family ( Magnaporthaceae ), spp. of Clostridium ( Bifusisporella Therefore, strain Bsas17 was classified and named... Bifusisporella sp. Bsas17.

[0055] On November 17, 2025, strain Bsas17 was deposited at the China Center for Type Culture Collection (CCTCC), Wuhan University, Wuhan, China, with accession number CCTCC NO: M 20252568. The collection was confirmed to be viable on November 24, 2025.

[0056] Example 2: Colonization of endophytic fungus Bsas17 in rice roots 1. Activation culture of bacterial strains The Bsas17 strain preserved on the slant was inoculated onto potato dextrose agar (PDA) solid medium for activation culture, and cultured in the dark at 25°C for 7 days for later use.

[0057] 2. Conversion and expression of GFP fluorescent protein Agrobacterium strains containing GFP fluorescent protein recombinant plasmids preserved in the laboratory were selected and inoculated into LB liquid medium containing kanamycin (Kana, 50 mg / L). The cultures were incubated overnight at 28°C and 190 rpm with shaking to activate the strain. The next day, the bacterial suspension was centrifuged to collect the cells, which were then resuspended in AIM induction medium and induced at 28°C and 180 rpm for 4-6 h. Spores or hyphae on the surface of endophytic fungi were scraped off with sterile water, and 100 μL of spore solution (hyphae solution) was added to a 2 mL centrifuge tube containing 100 μL of induced AIM medium. Subsequently, 200 μL of the mixture was spread onto AIM plates with an NC membrane, incubated at 22°C in the dark for 2 days, and then the NC membrane was removed and transferred to plates containing chlorimuron-methyl resistant plates for further incubation for 3-5 days. Hyphae from single colonies growing on the NC membrane were picked and observed for fluorescence under a microscope. Successful gene expression was confirmed by fluorescence phenotype. Colonies successfully transformed with GFP fluorescence were screened and plated on PDA plates. After purification for 6-8 days, the colonies were stored. The following co-culture steps used the Bsas17 strain, which successfully expressed GFP fluorescent protein.

[0058] AIM induction medium: per 1000 mL contains 1.0 g yeast extract, 2.0 g peptone, 2.0 g glucose, 0.5 g magnesium sulfate heptahydrate (MgSO4·7H2O), 0.5 g potassium dihydrogen phosphate (KH2PO4), 100 μmol acetylsuccinone (As), 50 mg kanamycin (Kana), pH 7.0, autoclaved at 121℃ for 20 min.

[0059] 3. Co-culture of Bsas17 strain with rice After removing the husks from rice seeds, shriveled, dry, and insect-infested grains were removed. Healthy rice seeds were disinfected with 75% alcohol for 5 minutes, followed by 1% NaClO for 20 minutes. The seeds were then rinsed five times with sterile water before use. The disinfected seeds were transferred to half MS medium using sterile forceps, sealed with sealing film, and incubated at 25°C for germination (16 h light / 8 h dark). After 3-4 days, the emerging seeds were inoculated into square dishes containing half MS + PDB medium, with 8-9 seeds per dish. The treatment group had 6-7 GFP-labeled Bsas17 bacterial discs (5 mm in diameter) inoculated below the seeds, while the control group had sterile bacterial discs in half MS + PDB medium. Each treatment was repeated in triplicate, incubated at 25°C for 16 h light / 8 h dark for 15-20 days until the three-leaf stage.

[0060] 1 / 2 MS + PDB medium: per 1000 mL contains 0.5 g MES, 2.2 g MS Salt, 5 g sucrose, 8 g agar powder, 17.5 g PDB, pH 5.7, autoclaved at 121℃ for 20 min.

[0061] 4. Observation of rice root colonization The rice roots of the control group and the treatment group were cleaned, dried, and then cross-sectioned and observed under a laser confocal microscope.

[0062] 5. Results Analysis like Figure 3 As shown, the GFP-labeled Bsas17 strain exhibited significant fluorescence signals in the epidermis, cortex, and stele cells of rice roots, indicating that the Bsas17 strain has successfully colonized these three tissue regions of rice.

[0063] Example 3: Endophytic fungus Bsas17 promotes rice growth 1. Activation culture of bacterial strains The Bsas17 strain preserved on the slant was inoculated onto potato dextrose agar (PDA) solid medium for activation culture, and cultured in the dark at 25°C for 7 days for later use.

[0064] 2. Preparation of Bsas17 liquid microbial fertilizer Endophytic fungal mycelial blocks (0.5 cm in diameter) of Bsas17, cultured for 7 days, were placed in PDB liquid fermentation medium (containing 5 g potato extract, 10 g peptone, and 15 g glucose per 1000 mL) for fermentation (25℃, 150 rpm, 7 days). Bsas17 liquid microbial fertilizer was obtained when the PDB liquid fermentation medium was fully colonized with mycelium.

[0065] 3. Co-culture of strain Bsas17 (1) Application of liquid microbial fertilizer: Mix the fermented liquid microbial fertilizer with the seedling substrate (liquid microbial fertilizer: substrate soil = 0.1L: 3L) and use it as new substrate soil for rice pot cultivation. Soak the seeds of Zhejing 99 rice and place them in a 30℃ dark constant temperature incubator for 2 days to germinate. When the seeds show white sprouts, sow them evenly.

[0066] (2) Rice seedling pot cultivation: Rice seeds were cultivated in seedling trays for 30 days (22-25℃, 16 h light / 8 h dark culture), and the growth was observed and recorded. Various growth indicators were statistically analyzed.

[0067] 4. Results Analysis like Figure 4 and Figure 5As shown, under pot culture conditions, plants inoculated with strain Bsas17 exhibited significantly higher stem width and fresh weight per plant than the control group. Specifically, the average stem width of the control group was 0.204 cm, while the average stem width of the Bsas17 inoculated group was 0.297 cm, an increase of approximately 45.5%. The average fresh weight per plant in the control group was 0.148 g / plant, while the average fresh weight per plant in the Bsas17 inoculated group was 0.217 g / plant, an increase of approximately 47.0%. Both overall plant performance and individual plant performance indicated that the plants inoculated with strain Bsas17 showed better growth than those in the uninoculated group.

[0068] Example 4: Endophytic fungus Bsas17 promotes tomato growth 1. Activation culture of bacterial strains The Bsas17 strain preserved on the slant was inoculated onto potato dextrose agar (PDA) solid medium for activation culture, and cultured in the dark at 25°C for 7 days for later use.

[0069] 2. Preparation of Bsas17 liquid microbial fertilizer Endophytic fungal mycelial blocks (0.5 cm in diameter) of Bsas17, cultured for 7 days, were placed in PDB liquid fermentation medium (containing 5 g potato extract, 10 g peptone, and 15 g glucose per 1000 mL) for fermentation (25℃, 150 rpm, 7 days). Bsas17 liquid microbial fertilizer was obtained when the PDB liquid fermentation medium was fully colonized with mycelium.

[0070] 3. Co-culture of strain Bsas17 with tomato (1) Application of liquid microbial fertilizer: Mix the fermented liquid microbial fertilizer with the seedling substrate (liquid microbial fertilizer: seedling substrate = 0.4 L: 40 L) and use it as new substrate soil for tomato pot cultivation. Soak the tomato seeds (Cooperation 903) and place them in a 37℃ dark constant temperature incubator to germinate for 2 days. When the seeds show white sprouts, sow them evenly in the seedling tray.

[0071] (2) Pot cultivation of tomato seedlings: Tomato seeds are cultivated in seedling trays for 30 days (22-25℃, 16 h light / 8 h dark).

[0072] (3) Pot cultivation of tomatoes during flowering period: Tomato seedlings that have been raised for 30 days are transplanted into planting pots with a diameter of 25 cm and a volume of 12 liters and cultivated for 3 months (22-25℃, 16 h light / 8 h dark culture). The growth is observed and recorded, and various growth indicators are statistically analyzed.

[0073] 4. Results Analysis like Figure 6 and Figure 7As shown, under pot culture conditions, plants inoculated with strain Bsas17 exhibited significantly higher plant height, leaf width, and number of flowers compared to the control group. Specifically, the average plant height of the control group was 45.3 cm, while the average plant height of the inoculated group was 74.1 cm, an increase of approximately 63.6%; the average leaf width of the control group was 2.85 cm, while the average leaf width of the inoculated group was 4.37 cm, an increase of approximately 53.1%; and the average number of flowers per plant was 1.0 in the control group and 4.7 in the inoculated group, an increase of approximately 3.7 flowers per plant. Overall, the plants inoculated with strain Bsas17 showed better growth than those in the uninoculated group.

[0074] The above results indicate that strain Bsas17 can effectively promote the vegetative and reproductive growth of the tested plants.

[0075] Example 5: Endophytic fungus Bsas17 promotes the growth of coriander 1. Activation culture of bacterial strains The Bsas17 strain preserved on the slant was inoculated onto potato dextrose agar (PDA) solid medium for activation culture, and cultured in the dark at 25°C for 7 days for later use.

[0076] 2. Preparation of Bsas17 liquid microbial fertilizer Endophytic fungal mycelial blocks (0.5 cm in diameter) of Bsas17, cultured for 7 days, were placed in PDB liquid fermentation medium (containing 5 g potato extract, 10 g peptone, and 15 g glucose per 1000 mL) for fermentation (25℃, 150 rpm, 7 days). Bsas17 liquid microbial fertilizer was obtained when the PDB liquid fermentation medium was fully colonized with mycelium.

[0077] 3. Co-culture of strain Bsas17 with coriander (1) Application of liquid microbial fertilizer: After crushing the protective shell of 0.7 g of Anselmi coriander seeds, evenly sow them on the surface of the substrate soil containing liquid microbial fertilizer (liquid microbial fertilizer: seedling substrate = 0.4 L: 40 L) for coriander pot planting.

[0078] (2) Cultivation of coriander in pots: Cultivate coriander for 60 days (22-25℃, 16 h light / 8 h dark), observe and record the growth, and statistically analyze various growth indicators.

[0079] 4. Results Analysis like Figure 8 and Figure 9As shown, under pot cultivation conditions, plants inoculated with strain Bsas17 exhibited significantly higher fresh weight per plant, total weight (total weight of coriander after 0.7 g of seeds matured), and plant height than the control group. Specifically, the average fresh weight per plant in the control group was 1.79 g / plant, while the inoculated group averaged 4.19 g / plant, an increase of approximately 133.6%; the average total fresh weight in the control group was 197.0 g, while the inoculated group averaged 287.0 g, an increase of approximately 45.7%; and the average plant height in the control group was 15.8 cm, while the inoculated group averaged 20.8 cm, an increase of approximately 31.6%. Both overall plant performance and individual plant performance indicated that the coriander plants inoculated with strain Bsas17 showed significantly better growth than the uninoculated group. Furthermore, both overall plant performance and individual plant performance demonstrated that the plants inoculated with strain Bsas17 exhibited superior growth compared to the uninoculated group.

[0080] The above results indicate that strain Bsas17 has a significant effect on promoting the growth and increasing the yield of coriander.

[0081] The above description is merely a specific embodiment of the present invention, intended to enable those skilled in the art to understand the content of the present invention and implement it accordingly, and should not be construed as limiting the scope of protection of the present invention. All equivalent modifications or substitutions made based on the essence of the present invention should be covered within the scope of protection of the present invention.

Claims

1. An endophytic fungus Bsas17, characterized in that, The endophytic fungus Bsas17 is classified as follows: Bifusisporella sp. Bsas17 is deposited at the China Center for Type Culture Collection, accession number CCTCC NO: M20252568.

2. A liquid microbial fertilizer containing endophytic fungi Bsas17, characterized in that, The method for preparing the liquid microbial fertilizer includes: inoculating the endophytic fungus Bsas17 as described in claim 1 into PDB liquid culture medium, culturing until the culture medium is filled with mycelia, and obtaining the liquid microbial fertilizer.

3. The application of the endophytic fungus Bsas17 as described in claim 1 in promoting rice growth, characterized in that, The application includes colonizing the endophytic fungus Bsas17 in rice root tissue.

4. The application as described in claim 3, characterized in that, The indicators for rice growth include plant fresh weight and stem width.

5. The application as described in claim 3, characterized in that, The application includes: inoculating the endophytic fungus Bsas17 into PDB liquid culture medium, culturing until the culture medium is covered with mycelium to obtain liquid microbial fertilizer; then mixing the liquid microbial fertilizer into the seedling substrate; and then sowing the germinated rice seeds into the seedling substrate for co-cultivation.

6. The application of the endophytic fungus Bsas17 as described in claim 1 in promoting tomato growth, characterized in that, The application includes co-culturing the endophytic fungus Bsas17 with tomato plants.

7. The application as described in claim 6, characterized in that, The indicators for tomato growth include plant height, leaf width, and number of flowers.

8. The application of the endophytic fungus Bsas17 as described in claim 1 in promoting the growth of coriander, characterized in that, The application includes co-culturing the endophytic fungus Bsas17 with coriander plants.

9. The application as described in claim 8, characterized in that, The indicators for coriander growth include: plant fresh weight and plant height.

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