Endophytic fungi for improving cold resistance of fraxinus mandshurica, composition and application

Abstract

The invention discloses an endophytic fungus for improving cold resistance of fraxinus mandshurica, a composition and application, and belongs to the technical field of microorganisms, the endophytic fungus is named as Didymellaapomorum fungus FMS1 and preserved in the China General Microbiological Culture Collection Center (CGMCC), and the preservation number is CGMCC NO.42058. The endophytic fungus has the advantages that the endophytic fungus is named as Didymellaapomorum fungus FMS1, and the preservation number is CGMCC NO.42058; according to the method, fraxinus mandshurica is used as a material, root irrigation or spraying treatment is conducted on the fraxinus mandshurica through endophytic fungi FMS1 of the fraxinus mandshurica, the cold resistance of the fraxinus mandshurica can be improved, and the plant height, stem diameter and other characters of the fraxinus mandshurica under low-temperature stress are remarkably improved; according to the technical scheme, the problems that fraxinus mandshurica is not cold-resistant and low in survival rate in the seedling stage can be effectively solved, the ability of the fraxinus mandshurica to deal with abnormal low temperature is improved, the low-temperature area planting area of the fraxinus mandshurica can be widened, and the method is suitable for large-area popularization and has important application prospects.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of microbial technology, and in particular to an endophytic fungus for improving cold resistance of Fraxinus mandshurica, a composition and applications. BACKGROUND

[0002] Fraxinus mandshurica Rupr. is a deciduous tree of Oleaceae and Fraxinus chinensis, and is a vulnerable species in the Chinese Plant Red List and a protected tree species. As a main component of natural forests in the northeast region of China, Fraxinus mandshurica is also a main afforestation tree species of artificial forests, and can be used for wood, landscaping and decoration purposes, and plays an important role in the protection and regulation of the environment in the northeast region of China, such as water conservation, soil conservation and prevention of environmental deterioration. However, the seed of Fraxinus mandshurica has a long dormancy period, the propagation speed is slow, the seedlings are easily affected by adverse factors such as drought, cold damage, insect pests and saline-alkali, and it is difficult to form a forest.

[0003] Temperature is one of the basic environmental factors affecting the survival and growth of plants, and is an important factor that seriously limits the distribution area of plants and affects the yield of organisms. Under low temperature conditions, the structure of cell membrane changes, leading to the leakage of electrolytes in cells, imbalance of oxidative metabolism in cells, accumulation of reactive oxygen species (ROS), and initiation of peroxidation of cell membrane lipids, causing damage to the cell membrane system; the synthesis of chlorophyll in plant cells is reduced, and photosynthesis is inhibited, thereby affecting the germination, growth, development and reproduction of plants. Research has found that early spring low temperature (mainly frost damage) still becomes an important obstacle for cultivating artificial forests of Fraxinus mandshurica, causing the top bud of seedlings to be damaged by frost, developing into a plant with no top bud in clusters, and significantly reducing the survival rate and preservation rate. Therefore, how to reduce the loss caused by low temperature stress and improve the resistance of trees has been a problem that needs to be solved for the cultivation of Fraxinus mandshurica. SUMMARY

[0004] The present application aims to provide an endophytic fungus for improving cold resistance of Fraxinus mandshurica to solve the problems in the background.

[0005] To achieve the above-mentioned purpose, the present application provides the following technical scheme: an endophytic fungus for improving cold resistance of Fraxinus mandshurica, named Didymella pomorum fungus FMS1, which is preserved in the China General Microbiological Culture Collection Center, with a preservation number of CGMCC NO.42058, a preservation address of No.3, Yitianyilu, Beichen, Chaoyang District, Beijing, and a preservation time of July 14, 2025.

[0006] Another object of the present application is to provide a composition for improving cold resistance of Fraxinus mandshurica, comprising the endophytic fungus and gamma-aminobutyric acid.

[0007] Another object of the present application is to provide an application of the endophytic fungus or the fermentation liquor thereof, or the composition in improving cold resistance of Fraxinus mandshurica.

[0008] Preferably, the endophytic fungus or the fermentation liquor thereof is used for reducing the conductivity and the content of malondialdehyde of Fraxinus mandshurica.

[0009] Preferably, the method for improving cold resistance of Fraxinus mandshurica comprises: using the endophytic fungus or the fermentation liquor thereof for root irrigation treatment of Fraxinus mandshurica, and using gamma-aminobutyric acid for foliar spraying treatment of Fraxinus mandshurica.

[0010] Preferably, the spraying concentration of the gamma-aminobutyric acid is 20-30 mmol / L.

[0011] Another object of the present application is to provide a root irrigation agent for improving cold resistance of Fraxinus mandshurica, comprising the endophytic fungus or the fermentation liquor thereof.

[0012] The present application uses Fraxinus mandshurica as a material, and uses the endophytic fungus FMS1 thereof for root irrigation treatment, so as to improve the cold resistance of Fraxinus mandshurica, and significantly improve the plant height, stem diameter and other properties of Fraxinus mandshurica under low temperature stress. The technical scheme provided by the present application can effectively solve the problems of low survival rate and poor cold resistance of Fraxinus mandshurica seedlings, improve the ability of Fraxinus mandshurica to cope with abnormal low temperature, widen the planting area of Fraxinus mandshurica in low temperature area, and is suitable for large-area popularization, and has important application prospect. BRIEF DESCRIPTION OF DRAWINGS

[0013] Figure 1 A morphological diagram of the endophytic fungus FMS1 separated for the embodiments of the present application;

[0014] Figure 2 A phenotype comparison diagram of Fraxinus mandshurica seedling after low temperature stress, and after treatment of the endophytic fungus FMS1 and combination of the endophytic fungus FMS1 and different exogenous signal substances;

[0015] Figure 3 A phenotype comparison diagram of Fraxinus mandshurica seedling after low temperature stress, and after treatment of the endophytic fungus FMS1 and combination of the endophytic fungus FMS1 and different exogenous signal substances;

[0016] Figure 4 A result diagram of the influence of the endophytic fungus FMS1 and combination of the endophytic fungus FMS1 and different exogenous signal substances on the plant height of Fraxinus mandshurica seedling;

[0017] Figure 5 A result diagram of the influence of the endophytic fungus FMS1 and combination of the endophytic fungus FMS1 and different exogenous signal substances on the stem diameter of Fraxinus mandshurica seedling;

[0018] A result diagram of the influence of the endophytic fungus FMS1 and combination of the endophytic fungus FMS1 and different exogenous signal substances on the stem diameter of Fraxinus mandshurica seedling;Figure 6 Figure for the effect of endophytic fungus FMS1 and different exogenous signal substances on the fresh weight of the aboveground part of Fraxinus mandshurica seedlings;

[0019] Figure 7 Figure for the effect of endophytic fungus FMS1 and different exogenous signal substances on the photosynthetic pigment of Fraxinus mandshurica seedlings;

[0020] Figure 8 Figure for the effect of endophytic fungus FMS1 and different exogenous signal substances on the relative electrical conductivity of Fraxinus mandshurica seedlings;

[0021] Figure 9 Figure for the effect of endophytic fungus FMS1 and different exogenous signal substances on the total activity of SOD of Fraxinus mandshurica seedlings;

[0022] Figure 10 Figure for the effect of endophytic fungus FMS1 and different exogenous signal substances on the chlorophyll content of Fraxinus mandshurica seedlings after low temperature stress;

[0023] Figure 11 Figure for the effect of endophytic fungus FMS1 and different exogenous signal substances on the nitrogen content of Fraxinus mandshurica seedlings after low temperature stress;

[0024] Figure 12 Figure for the effect of endophytic fungus FMS1 and different exogenous signal substances on the relative electrical conductivity of Fraxinus mandshurica seedlings after low temperature stress;

[0025] Figure 13 Figure for the effect of endophytic fungus FMS1 and different exogenous signal substances on the MDA content of Fraxinus mandshurica seedlings after low temperature stress;

[0026] Figure 14 Figure for the effect of endophytic fungus FMS1 and different exogenous signal substances on the POD activity of Fraxinus mandshurica seedlings after low temperature stress;

[0027] Figure 15 Figure for the effect of endophytic fungus FMS1 and different exogenous signal substances on the CAT enzyme activity of Fraxinus mandshurica seedlings after low temperature stress;

[0028] Figure 16 Figure for the effect of endophytic fungus FMS1 and different exogenous signal substances on the total activity of SOD of Fraxinus mandshurica seedlings after low temperature stress. DETAILED DESCRIPTION

[0029] The technical solutions in the embodiments of the present application will be clearly and completely described below with reference to the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, rather than all the embodiments of the present application. Based on the embodiments of the present application, all other embodiments obtained by those skilled in the art without creative efforts belong to the scope of protection of the present application.

[0030] Plant endophytic fungi are a kind of microbial resources with wide application value, mainly referring to those fungi living in various tissues and organs of healthy plants at a certain stage or all stages of their life history. In the long-term co-evolution with plants, endophytic fungi can enhance the resistance function of the plant body through various ways, mainly in the absorption of nutrients, the protection system of the plant, the regulation of hormones, water metabolism, plant defense signal pathways, etc., to promote plant growth and enhance the adversity adaptability of the host. It is a very promising strategy to use endophytic fungi to overcome the damage caused by abiotic stress to plants.

[0031] The present application uses Fraxinus mandshurica as the material, uses a Fraxinus mandshurica endophytic fungus FMS1 and a combination of the endophytic fungus FMS1 and gamma-aminobutyric acid (GABA) to improve the cold resistance of Fraxinus mandshurica, provides new microbial resources for mining new strains of biological energy function, at the same time, provides a new path for reducing the loss caused by low temperature stress of forest trees and improving the cold resistance of forest trees, provides a scientific basis for Fraxinus mandshurica excellent variety breeding and establishes an efficient stress resistance cultivation technology system of Fraxinus mandshurica.

[0032] Specifically, in an embodiment of the present application, an endophytic fungus for improving the cold resistance of Fraxinus mandshurica is provided, which is named Didymella pomorum fungus FMS1, and is simply referred to as endophytic fungus FMS1. The endophytic fungus FMS1 is preserved in the China General Microbiological Culture Collection Center, the preservation number is CGMCC NO.42058, the preservation address is No.3, Yitianli 1st Courtyard, Beichen West Road, Chaoyang District, Beijing, China, and the preservation time is July 14, 2025.

[0033] In another embodiment of the present application, a composition for improving the cold resistance of Fraxinus mandshurica is also provided, which comprises the above-mentioned endophytic fungus and gamma-aminobutyric acid.

[0034] In another embodiment of the present application, the application of the above-mentioned endophytic fungus or its fermentation liquor or the above-mentioned composition in improving the cold resistance of Fraxinus mandshurica is also provided.

[0035] Specifically, the endophytic fungus or its fermentation liquor is used to reduce the conductivity and the content of malondialdehyde of Fraxinus mandshurica.

[0036] In practical applications, the method for improving the cold resistance of Fraxinus mandshurica includes: using the endophytic fungus or the fermentation liquor thereof for root irrigation treatment of Fraxinus mandshurica, and using gamma-aminobutyric acid for foliar spraying treatment of Fraxinus mandshurica.

[0037] Specifically, the spraying concentration of gamma-aminobutyric acid is 20-30 mmol / L.

[0038] In the embodiment of the present application, the endophytic fungus FMS1 is used for root irrigation of Fraxinus mandshurica seedlings, which significantly improves the plant height, stem diameter of Fraxinus mandshurica seedlings under low temperature stress and the cold resistance of the seedlings after low temperature treatment. For Fraxinus mandshurica seedlings, the combination of endophytic fungus FMS1 root irrigation and gamma-aminobutyric acid spraying, gamma-aminobutyric acid spraying once a day for one week before low temperature treatment, and endophytic fungus FMS1 root irrigation 2-3 times a week, can significantly improve the cold resistance of Fraxinus mandshurica. The spraying and irrigation treatment time is before early spring low temperature frost.

[0039] In another embodiment of the present application, a root irrigation agent for improving the cold resistance of Fraxinus mandshurica and / or promoting the growth of Fraxinus mandshurica is also provided, which comprises the above-mentioned endophytic fungus or fermentation liquor thereof. It should be noted that the root irrigation agent can also comprise solvent, diluent and other components.

[0040] In another embodiment of the present application, the application of the above-mentioned endophytic fungus FMS1 in the preparation of seed coating agents, seed dressings and the use of fertilizers is also provided.

[0041] In another embodiment of the present application, the application of the above-mentioned endophytic fungus FMS1 in the preparation of growth stimulants or regulators and the use of other fertilizers is also provided.

[0042] In another embodiment of the present application, the application of the above-mentioned endophytic fungus FMS1 in the preparation of biological pesticides and biological agent compounding is also provided.

[0043] The following examples are embodiments of the technical scheme of the present application in practical application, but are not limited thereto, wherein the materials and reagents involved are commercially available unless otherwise specified; the experimental methods used are conventional methods unless otherwise specified.

[0044] Example 1: The present example provides a method for isolating and identifying the endophytic fungus FMS1, which specifically comprises the following steps:

[0045] S1, isolation of endophytic fungus FMS1: under sterile conditions, cut the stem of Fraxinus mandshurica into 0.5 cm small pieces and place them in PDA medium. Invert, cultivate at 26-28 DEG C, observe every day, pick out the grown mycelium and grow on a new culture dish. After the colony grows, pick out a small amount of edge mycelium and repeat the operation twice to purify the fungus, to obtain the endophytic fungus FMS1, the morphological photo of which is as follows:Figure 1 as shown.

[0046] S2, ITS sequence identification of endophytic fungus FMS1: select well-grown colonies, scrape the mycelium into a 2 mL centrifuge tube. Use the CTAB method to extract endophytic fungus DNA, dissolve the obtained DNA product in ddH2O, and store in a -20°C refrigerator for amplification. Use fungal universal ITS primers ITS1 and ITS4 to amplify the full-length sequence of ITS under the reaction conditions (pre-denaturation 94°C for 2 min; denaturation 94°C for 40 s; annealing 57°C for 50 s; extension 72°C for 1 min; 35 cycles; extension 72°C for 10 min; 4°C forever); take 3.0 μL of the amplification reaction product and mix with 5xLoading Buffer, load on an agarose gel containing 10 mg / mL Gold Red 3%, and perform electrophoresis in 1.0xTAE buffer (voltage 120 V). After electrophoresis, remove the gel and place it in a gel imaging system; according to the results of PCR electrophoresis, send the PCR product to Shengong Bioengineering (Shanghai) Co., Ltd. for one-way sequencing of the upstream and downstream primers, and obtain the sequencing results.

[0047] Example 2: This example provides a pot experiment of different signal processing Fraxinus mandshurica 2-year-old or 3-year-old Fraxinus mandshurica seedling: seven different signals, namely 150 μmol / L of melatonin (MT150), 400 mg / L of melatonin (CH400), 25 mmol / L of γ-aminobutyric acid (GABA25), endophytic fungus FMS1 (S1), and the combination of endophytic fungus FMS1 and the three exogenous substances (denoted as S1+MT, S1+GABA, S1+CH, respectively), are sprayed (exogenous substances are sprayed) and irrigated (endophytic fungus FMS1 is irrigated) on Fraxinus mandshurica seedlings within one week; after one week, the treated Fraxinus mandshurica seedlings are placed in a cold room at -2°C for low-temperature stress for 2 hours, for a total of 9 treatments, denoted as LT (water low-temperature treatment group), NT (water normal-temperature treatment group), MT (melatonin MT150 treatment group), S1+MT (endophytic fungus FMS1+melatonin MT150 treatment group), CH (melatonin CH400 treatment group), S1+CH (endophytic fungus FMS1+melatonin CH400 treatment group), GABA (γ-aminobutyric acid treatment group), S1+GABA (endophytic fungus FMS1+γ-aminobutyric acid treatment group), and S1 (endophytic fungus FMS1 treatment group). After low-temperature stress, the Fraxinus mandshurica seedlings are sampled and observed for growth and physiological state after 1 day of recovery, and the results are shown in FIGS. 2A-2D. Figure 2As shown, the best exogenous signal for promoting the resistance of Fraxinus mandshurica seedling to low temperature stress is S1+GABA. Under low temperature stress (2h at-2℃), various treatments can significantly improve the cold resistance of 2-year-old or 3-year-old Fraxinus mandshurica seedlings, and reduce the damage caused by low temperature to seedlings, among which the S1+GABA group is the best.

[0048] Example 3: This example provides a pot experiment of Fraxinus mandshurica seedlings after different signal treatments: In June 2024, a pot experiment was conducted using 9cmx8cmx9cm plastic pots, and the cultivation substrate was organic soil and vermiculite (mass ratio of 3:1). The soil in each pot was consistent. Three seedlings were planted in each pot. After 60 days, seedlings with consistent growth were selected for low temperature stress experiment. The specific treatment was the same as in Example 2, including: LT (water low temperature treatment group), NT (water normal temperature treatment group), MT150 (melatonin MT150 treatment group), S1+MT (endophytic fungus FMS1+melatonin MT150 treatment group), CH400 (melatonin CH400 treatment group), S1+CH (endophytic fungus FMS1+melatonin CH400 treatment group), GABA25 (γ-aminobutyric acid treatment group), S1+GABA (endophytic fungus FMS1+γ-aminobutyric acid treatment group), S1 (endophytic fungus FMS1 treatment group). Three exogenous substances were sprayed continuously every day, and endophytic fungus FMS1 was treated by root irrigation twice a week. Each treatment had 24 seedlings. After one week of signal treatment, the NT treatment was placed in a low temperature incubator at 0℃ for one week. After recovery for 1d, the growth state of Fraxinus mandshurica seedlings was observed, as shown in Figure 3 As shown, under low temperature stress (7d at 0℃), each treatment can significantly enhance the cold resistance of Fraxinus mandshurica seedlings. In addition, physiological indicators were measured to determine the effect of different treatments on the growth and cold resistance of Fraxinus mandshurica seedlings.

[0049] Specifically, the determination method of each index is as follows:

[0050] (1) Growth index determination: After the seedlings treated with different signals were cultured for a certain time at normal temperature, the height, stem diameter, and root-shoot ratio (fresh weight of aboveground part / fresh weight of aboveground part) of the seedlings were measured, and the results are shown in Figure 4 , Figure 5 , Figure 6 as shown.

[0051] The whole seedling was dug out, the roots were washed clean with flowing water, the surface water was absorbed with absorbent paper, and then the roots were placed in the reaction solution at 37℃ for dark reaction for 2h. After that, the roots were taken out, the reaction solution was absorbed, and the roots were ground in ice bath. The content of stress resistance substances was detected.

[0052] Three plants of uniform growth were selected for each treatment. The entire plant, still covered in soil, was carefully removed from the pot and separated from the soil, preserving its integrity as much as possible to ensure experimental accuracy. Excess soil and impurities were then washed away from the roots with water, and the above-ground parts were trimmed. Roots were scanned using a root scanner (GXY-A, Top Cloud Agriculture), and root density, specific root length, and average diameter were calculated. (Root density = root dry weight / root volume; specific root length = root length / root dry weight). All measurements were repeated three times.

[0053] (2) Photosynthetic index determination: The photosynthetic characteristics of Manchurian ash seedlings under dry low temperature stress were detected after soaking with different signals and combinations. The detection time was from 9 to 11 am. The third leaf of each group was randomly selected and the chlorophyll and nitrogen content of the leaves were measured using a plant nutrient meter (LD-YD).

[0054] (3) Antioxidant enzyme activity assay: Superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) were measured by ammonia blue tetrazolium photoreduction method, guaiacol colorimetric method and ultraviolet spectrophotometric absorption method, respectively.

[0055] (4) Determination of the content of permeating substances: The content of malondialdehyde (MDA) was determined by the heating method of thiobarbituric acid.

[0056] (5) Conductivity measurement: Cell membrane permeability conductivity was measured using a Leici DDSJ-308A conductivity meter.

[0057] The measurement results of the above-mentioned relevant indicators are as follows:

[0058] Low temperatures reduced the root-to-rhizome ratio in *Fragranceia spp.*, but this damage was mitigated by the endophytic fungus FMS1 (S1) and the combination of 25 mmol / L γ-aminobutyric acid (GABA) with FMS1 (S1+GABA). The fresh weight of the underground parts of seedlings treated with S1 and S1+GABA increased by 46.91% and 43.77% respectively compared to the drought control group. Different signal treatments significantly increased photosynthetic and transpiration rates, and chlorophyll content compared to the low-temperature control. Figure 7 As shown, after 1 day of recovery, the chlorophyll a (Ca) content in the S1 treatment increased by 19.36% compared to the control, the chlorophyll b (Cb) content increased by 11% compared to the control, and the total chlorophyll (C... T The content of ) increased by 16.6% compared to the control, and in terms of relative conductivity (e.g. Figure 8 As shown), the S1 treatment reduced SOD activity by 24.7% compared to the low-temperature control, and in terms of SOD enzyme activity (e.g. Figure 9 As shown in the figure, the S1 treatment significantly reduced the damage caused by low temperature to the seedlings of Manchurian ash (S. spp.) by 8.84% compared to the low temperature control.

[0059] The nitrogen content of S1 and S1+GABA treatment groups increased by 20.44% and 18.44% respectively compared with the low temperature control group (as shown in Figure 11 As shown in Figure 12 Under low temperature stress, the relative conductivity of different treatments was lower than that of the low temperature control group, and the conductivity of S1 and S1+GABA treatment groups decreased by about 16%-29% compared with the low temperature control group. After low temperature stress, S1 and S1+GABA treatment could effectively reduce the content of seedling malondialdehyde (as shown in Figure 13 The above treatments significantly reduced the damage of Fraxinus mandshurica seedlings under low temperature. Compared with the low temperature control group, different signal treatments significantly improved the photosynthetic and transpiration rates, and the chlorophyll content. The photosynthetic rate increased by more than 90%, and the chlorophyll content increased by 21.08% and 19.44% (as shown in Figure 10 After low temperature stress, the SOD enzyme activity of S1 and S1+GABA treatment groups increased by 15% (as shown in Figure 16 The POD enzyme activity of S1 treatment group increased by 91.15% compared with the low temperature control group (as shown in Figure 14 The CAT enzyme activity of S1+GABA treatment group increased by 32.25% compared with the low temperature control group (as shown in Figure 15

[0060] The above results show that endophytic fungus FMS1 alone can enhance the photosynthetic pigment content and significantly promote the growth of Fraxinus mandshurica, thereby increasing the height and diameter of the new stem. It also reduces the relative conductivity, increases the antioxidant enzyme activity, and thus enhances the cold resistance of Fraxinus mandshurica seedling. The combination of endophytic fungus root irrigation treatment and 25mmol / L γ-aminobutyric acid (S1+GABA) treatment significantly enhances the cold resistance of Fraxinus mandshurica seedlings, significantly promotes the increase of stress resistance substances, enhances the chlorophyll and photosynthesis, and reduces the relative conductivity and conductivity. It also promotes the increase of root flavonoids, polyphenols content and leaf nitrogen content, thereby enhancing the photosynthetic performance and promoting growth. When γ-aminobutyric acid is used together with endophytic fungus S1, the cold resistance of Fraxinus mandshurica is stronger.

[0061] ​In summary, the endophytic fungus FMS1 can significantly improve the plant height, stem diameter and cold resistance of Fraxinus mandshurica seedling under low temperature stress, and provide excellent strain resources for microbial fertilizer to promote the growth of Fraxinus mandshurica. In addition, the combination treatment of endophytic fungus FMS1 and 25 mmol / L gamma-aminobutyric acid can significantly improve the photosynthetic and transpiration rates, chlorophyll and nitrogen contents of Fraxinus mandshurica seedlings, significantly reduce the conductivity and MDA content of the seedlings, significantly reduce the damage of low temperature to the Fraxinus mandshurica seedlings, effectively solve the problems of low cold resistance and low survival rate of Fraxinus mandshurica seedlings, improve the ability of Fraxinus mandshurica to cope with abnormal low temperature, and have wide application value. The technical scheme provided in the embodiments of the present application is pollution-free, low-cost, highly operable, can be used on a large scale, and has high efficiency, and provides a practical new technology for effectively solving the bottleneck problem of low cold resistance in the process of seedling raising, transplanting, planting and forestation of Fraxinus mandshurica.

[0062] Based on the above ideal embodiments according to the present application, through the above description, relevant personnel can make various changes and modifications without deviating from the technical idea of the present application. The technical scope of the present application is not limited to the contents in the specification.

Claims

1. An endophytic fungus that enhances the cold resistance of Ash trees, characterized in that, The fungus, named FMS1 of the genus Didymellapomorum, is deposited at the China General Microbiological Culture Collection Center with accession number CGMCC NO.42058.

2. A composition for improving the cold resistance of Manchurian ash, characterized in that, It includes the endophytic fungus as described in claim 1 and γ-aminobutyric acid.

3. The use of an endophytic fungus as described in claim 1 or its fermentation broth, or the composition as described in claim 2, in improving the cold resistance of Ash.

4. The application according to claim 3, characterized in that, The endophytic fungi or their fermentation broth are used to reduce the electrical conductivity and malondialdehyde content of Ash.

5. The application according to claim 3, characterized in that, Methods to improve the cold resistance of Manchurian ash include: applying the endophytic fungus or its fermentation liquid to the roots of Manchurian ash by irrigation or spraying, and applying γ-aminobutyric acid to the leaves of Manchurian ash in combination.

6. The application according to claim 5, characterized in that, The spray concentration of γ-aminobutyric acid is 20-30 mmol / L.

7. A root drenching agent for improving the cold resistance of Manchurian ash, characterized in that, Includes the endophytic fungus or its fermentation broth as described in claim 1.

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