Trichoderma pholiotae strain WDNT01 with broad-spectrum biocontrol activity and its application

Abstract

The present invention discloses a Trichoderma pholiotae strain WDNT01 with broad-spectrum biocontrol effects and its applications, relating to the field of microbial technology. The Trichoderma pholiotae strain WDNT01, deposited with CCTCC NO: M20231270, is a broad-spectrum antibacterial Trichoderma strain. Its antibacterial rates against the pathogens of pepper white rot, pepper wilt, and kiwifruit soft rot reach 97.49%, 91.08%, and 83.48%, respectively. The strain exhibits excellent antibacterial effects and has broad prospects for development and application.

Description

Technical Field

[0001] The present invention relates to the field of microbial technology, and more particularly to a Trichoderma pholiotae strain WDNT01 with broad-spectrum biocontrol effects and applications thereof. Background Art

[0002] White rot of pepper is a disease that occurs on peppers and is caused by infection with Sclerotium sclerotiorum. It mainly harms the base of the stem and the roots. The lesions are water-soaked at first, then gradually expand to dark brown and slightly sunken. There are white silky mycelium on the lesions, which gather into bundles and extend radially to the surrounding and upper parts, gradually spreading to adjacent plants and causing infection. Pepper wilt is a disease that occurs on peppers and is caused by Fusarium oxysporum. It mainly occurs in the seedling stage or the flowering and fruiting stage, and harms the roots or root collars. After the disease occurs, the plants wilt, and when the diseased stems are cross-sectioned, the vascular bundles of the diseased part turn brown. When the humidity is high, white, red or blue-green mold appears on the surface of the base of the stem. Kiwifruit soft rot is a fruit disease caused by Botrytis cinerea or Phomopsis spp., which mainly harms fruits near maturity (fruit drop on the tree) and during storage. The pathogen attacks fruit during the flowering and young fruit stages, remaining dormant in the flesh for a long time. The fruit is usually asymptomatic when harvested, and symptoms do not appear until the fruit ripens. Initially, the lesions appear brown and slightly sunken, and the fruit quickly softens. The lesions are surrounded by a yellow-green hue, and a dark green, water-soaked halo appears at the junction of the lesion and the healthy part. Peeling the lesions reveals white, flocculent rotting flesh, and the fruit undergoes soft rot, losing its edible value.

[0003] Currently, the prevention and control of crop pests and diseases primarily relies on chemical control. While pesticides are highly effective, simple, and low-cost, they also present challenges, such as increased pesticide residues, reduced soil microbial diversity, soil hardening, and increased pathogen resistance. Biological control, on the other hand, offers a promising and environmentally friendly approach to plant disease control.

[0004] Therefore, it is an urgent problem for those skilled in the art to find efficient biocontrol strains for diseases such as pepper white rot, pepper wilt, and kiwifruit soft rot. Summary of the Invention

[0005] In view of this, the present invention provides a Trichoderma pholiotae strain WDNT01 having a broad-spectrum biocontrol effect and its application.

[0006] In order to achieve the above object, the present invention adopts the following technical solutions:

[0007] A Trichoderma pholiotae strain WDNT01 with broad-spectrum biocontrol activity and its application. The strain WDNT01 has a deposition number of CCTCC NO: M 20231270 and was deposited with the China Center for Type Culture Collection at Wuhan University, Wuhan, China on July 12, 2023. It is classified as Trichoderma pholiotae.

[0008] Another object of the present invention is to provide a microbial agent comprising the above-mentioned strain WDNT01.

[0009] Preferably, the bacterial agent is in the form of a bacterial suspension, powder, tablet or fermentation filtrate.

[0010] Another object of the present invention is to provide the use of the above-mentioned strain WDNT01 or the above-mentioned microbial agent.

[0011] Preferably, the application is to prevent and control pepper white rot, pepper wilt or kiwifruit soft rot.

[0012] Preferably, the application is applying the fermentation filtrate to the roots of peppers or kiwifruit.

[0013] Preferably, in the application, xylose is used as the carbon source and peptone is used as the nitrogen source for preparing the fermentation filtrate.

[0014] Beneficial effects: The Trichoderma pholiotae strain WDNT01 provided by the present invention is a broad-spectrum antibacterial Trichoderma strain, and its antibacterial rates against the pathogens of pepper white rot, pepper wilt, and kiwifruit soft rot reach 97.49%, 91.08%, and 83.48%, respectively. It has good antibacterial effect and broad development and application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] 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 or the description of the prior art. Obviously, the drawings described below are merely embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on the provided drawings without paying any creative work.

[0016] Figure 1 These are the effects of the initial screening strains against pathogens; the left side of each picture is the treatment, and the right side is the blank control (pepper white rot pathogen).

[0017] Figure 2 These are the effects of the initial screening strains against pathogens; the left side of each picture is the treatment, and the right side is the blank control (pepper wilt pathogen).

[0018] Figure 3 These are the effects of the initial screening strains against pathogens; the left side of each picture is the treatment, and the right side is the blank control (kiwifruit soft rot pathogen).

[0019] Figure 4 The morphological identification results of strain WDNT01 are shown, where a represents mycelium and spore-forming structures, and b represents conidia.

[0020] Figure 5 Phylogenetic tree of strain WDNT01 based on ITS sequences.

[0021] Figure 6 Phylogenetic tree of strain WDNT01 based on TEF sequences.

[0022] Figure 7 Figure 3 is the effect of different carbon sources on the antibacterial effect of the fermentation broth of strain WDNT01, where a is against pepper white rot; b is against pepper wilt; c is against kiwifruit soft rot; CK is the blank control of each pathogen.

[0023] Figure 8 Figure 3 shows the effect of different nitrogen sources on the antibacterial effect of WDNT01 fermentation broth, where a is against pepper white rot - from left to right and from top to bottom are urea, ammonium chloride, sodium nitrate, CK, ammonium nitrate, glycine, tryptophan, tyrosine, peptone, and ammonium acetate; b is against pepper wilt - from left to right and from top to bottom are ammonium acetate, ammonium nitrate, urea, CK, tyrosine, ammonium chloride, glycine, sodium nitrate, tryptophan, and peptone; c is against kiwifruit soft rot - from left to right and from top to bottom are tyrosine, peptone, glycine, tryptophan, sodium nitrate, ammonium nitrate, ammonium chloride, ammonium acetate, urea, and CK. DETAILED DESCRIPTION

[0024] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0025] Example 1 Isolation, screening and identification of strain WDNT01

[0026] 1. Separation

[0027] Using the five-point sampling method, soil samples were collected from the pepper rhizosphere 10-15 cm from the soil surface in the pepper planting area. Sterile water and the collected soil sample were diluted in a ratio of 9:1, and the soil suspension was prepared by shaking at a speed of 150 r / min for 20 minutes. The concentrations of 10-1 , 10 -2 , 10 -3 and 10 -4 Prepare a soil suspension of varying concentrations. Under sterile conditions, aspirate 100 μL of the soil suspension and evenly spread it onto the surface of a Trichoderma selective culture medium. Repeat three times for each concentration. Finally, place the culture dish in a 28°C incubator in the dark for 3-5 days. Observe and record the number of fungal colonies. Select intact, single colonies and isolate and purify the Trichoderma using PDA medium for preservation.

[0028] Trichoderma selective medium: MgSO4 0.2 g, KH2PO4 0.9 g, KCl 0.15 g, NH4NO3 1.0 g, glucose 3.0 g, pentachloronitrobenzene 0.3 g, agar 18 g, rose bengal 0.15 g, chloramphenicol 0.25 g, streptomycin 0.05 g and 1000 mL of distilled water, sterilized at 121 ° C for 20 min, natural pH.

[0029] Using this method, a total of 37 Trichoderma strains were isolated and purified from the collected soil samples, as shown in Table 1.

[0030] Table 1 Trichoderma isolated from rhizosphere soil of different types of peppers

[0031]

[0032] 2. Screening

[0033] Sclerotium rolfsii, Fusarium oxysporum, and Phomopsis lithocarpus (provided by the Biological Control Laboratory, Institute of Plant Protection, Guizhou Academy of Agricultural Sciences) were used as target bacteria, and Trichoderma strains isolated from soil samples were used as test bacteria. Using the plate-to-plate method, 5-mm-diameter cakes of the pathogen and the test bacteria were inoculated on the right and left sides of a PDA culture medium, respectively. The inoculated plates were incubated in the dark at 28°C for 5 days. The target bacterial colony diameters were measured, and the growth of the target bacteria was recorded. The inhibitory rates were calculated and compared to screen and preserve broad-spectrum antagonistic strains.

[0034] Growth inhibition rate (%) = (control colony radius - treated colony radius / control colony radius) × 100

[0035] In this example, 9 strains were screened from 37 candidate strains, showing good antibacterial effects on the three pathogens. WDNT01 was the best, as shown in Table 2 and the attached Figure 1-3 .

[0036] Table 2 Antibacterial effect of antagonistic strains on three pathogens

[0037]

[0038] Note: Lowercase letters indicate significant differences (p<0.05).

[0039] Antagonistic screening results showed that all nine strains listed in Table 2 exhibited broad-spectrum antifungal activity against the three pathogens. The inhibition rates of the nine strains against pepper bacterial rot ranged from 29.19% to 97.49%. The order of inhibition, from highest to lowest, was WDNT01 > WDNT-M03 > THT-M01 > BCT-M02 > DST-M02 > WDNT-M04 > THT-M04 > THT-M03 > DST-M03. Three strains, THT-M01, WDNT-M03, and WDNT01, exhibited inhibitory effects exceeding 90% against pepper bacterial rot, with WDNT01 showing the strongest inhibitory activity, achieving an inhibition rate of 97.49%, significantly higher than the other treatments. The nine strains showed inhibition rates against pepper Fusarium wilt ranging from 79.31% to 93.36%. The order of inhibition, from highest to lowest, was WDNT-M04 > THT-M01 > WDNT01 > DST-M02 > WDNT-M03 > BCT-M02 > THT-M04 > THT-M03 > DST-M03. Four strains (WDNT-M04, THT-M01, WDNT01, and DST-M02) exhibited inhibitory effects exceeding 90% against pepper Fusarium wilt, reaching 93.36%, 91.30%, 91.08%, and 90.22%, respectively. The nine strains demonstrated inhibition rates of 73.56% to 83.48% against the kiwifruit soft rot pathogen. The order of inhibition, from highest to lowest, was WDNT01 > DST-M02 > WDNT-M03 > THT-M01 > THT-M04 > BCT-M02 > WDNT-M04 > DST-M03 > THT-M03. WDNT01 showed the highest inhibitory effect, achieving an inhibition rate of 83.48%. Therefore, based on the combined antibacterial activity against the three pathogens, WDNT01 demonstrated the broadest and most effective antibacterial activity.

[0040] 3. Identification

[0041] The strain WDNT01, identified as having the best antibacterial activity, was inoculated onto PDA medium and cultured in the dark at 28°C for 5 days. The colony morphology was recorded and images were collected. Mycelium and spores were gently picked from the edge of the WDNT01 colony using an inoculating needle. Slides were prepared with distilled water and placed under a microscope for observation and recording. Morphological characteristics, including mycelium, spores, and spore-producing structures, were measured and images were collected to preliminarily determine its taxonomic status.

[0042] The mycelium of this strain grows radially on PDA medium, and the colonies are orange-red. Under a microscope, the mycelium of Trichoderma WDNT01 is transparent and septate. The mycelium branches out to form conidiophores, which are colorless. 2-3 bottle-shaped conidiophores are produced at the top of the conidiophores. Spherical or ellipsoidal conidia are colorless and 3.57-5.80μm×3.20-4.28μm in size (see attached). Figure 4 The above morphological characteristics were consistent with those of Trichodermapholiotae reported previously, and strain WDNT01 was preliminarily identified as T. pholiotae.

[0043] Strain WDNT01 was inoculated onto PDA and activated in the dark at 28°C. Mycelia were scraped from the colony on the third day of growth and ground into a powder in liquid nitrogen. DNA was extracted using a fungal DNA extraction kit (Shanghai Bioengineering). Sequences were amplified using primer pairs ITS1 and ITS4, and TEF1-728F and TEF1-rev, respectively. A 25 μL reaction system consisted of 9.5 μL of sterile water, 1 μL of DNA template, 1 μL of each upstream and downstream primer, and 12.5 μL of 2× Es Taq Master Mix. PCR amplification conditions included pre-denaturation at 94°C for 5 min, 34 cycles of denaturation at 94°C for 30 s, annealing at 55°C for 30 s, and extension at 72°C for 19 min, followed by extension at 72°C for 10 min. Amplified products were analyzed by 1.5% agarose gel electrophoresis and sent to the company for sequencing. The sequencing results were searched for similarity against the NCBI database, and ITS and TEF sequences of closely related species of strain WDNT01 were downloaded from GeneBank. ClustalW software was used for systematic alignment and assembly. Using MEGA11 as the testing platform, a phylogenetic tree was constructed using the Maximum Likelihood method and tested using bootstrap techniques. This confirmed the taxonomic status of strain WDNT01.

[0044] The nucleotide sequence of strain WDNT01 was amplified using ITS and tef primers. The resulting ITS and tef product fragments were approximately 550-600 bp in length. A phylogenetic tree of Trichoderma was constructed based on the ITS and tef gene sequences. In the tree constructed with the ITS sequence, strain WDNT01 and Trichoderma pholiotae were located on the same branch with a support level of 89% (see Appendix). Figure 5 In the tree constructed based on tef sequences, strain WDNT01 and T. pholiotae were also located on the same branch with a support strength of 100% (see Appendix Figure 6 ).

[0045] Based on the above morphological observation and molecular identification, strain WDNT01 was identified as Trichoderma pholiotae.

[0046] 4. Preservation

[0047] Trichoderma pholiotae WDNT01 was deposited in the China Center for Type Culture Collection on July 12, 2023, at Wuhan University, Wuhan, China, with the deposit number: CCTCC NO: M 20231270, and was classified as Trichoderma pholiotae.

[0048] Example 2 Optimization of fermentation conditions of strain WDNT01

[0049] Activate the Trichoderma WDNT01 strain onto a PDA plate for later use. After the colonies have grown for 48 hours, use a 5 mm diameter punch to punch out bacterial cakes uniformly at the edge of the colonies.

[0050] 1. Effects of different carbon sources on the antibacterial activity of Trichoderma WDNT01 fermentation products

[0051] Sabouraud liquid medium (SD medium: put 40g glucose and 10g peptone into 1000mL pure water, stir until completely dissolved, natural pH value) was used as the basal medium for the fermentation process. At the same time, lactose, maltose, soluble starch, xylose, fructose, and glycerol were used to replace the carbon source - glucose in the basal medium. Finally, fermentation media with 7 different carbon sources were prepared. Each treatment was repeated 3 times, and the liquid volume of each bottle of culture medium was 60%. Then, 4 bacterial cakes were inoculated into each bottle of culture medium and placed in a shaker at 150 r / min and 25°C for fermentation for 5 days. After the fermentation, the mycelium was first filtered with filter paper, and the mycelium was collected with filter paper and placed in a 60°C drying oven for 4 hours before being weighed. The empty filter paper pieces dried simultaneously were used as controls, and each treatment was repeated 3 times. The filtrate after preliminary filtration was filtered again with a 0.45 μm filter to obtain a sterile fermentation filtrate. The filtrate was then mixed with melted and cooled PDA culture medium at a ratio of 1:10 and poured onto plates. The center of the plate was inoculated with pathogenic fungi of pepper white rot, pepper wilt and kiwifruit soft rot with a diameter of 5 mm. Only SD culture medium was added as a control. Both the treatment and the control were repeated 3 times. All plates were placed in a constant temperature incubator at 28°C for dark culture. The colony diameter was measured on the third day of growth, and the inhibition rate of the fermentation liquid under different treatments was calculated.

[0052] Growth inhibition rate (%) = (control colony diameter - treated colony diameter) / control colony diameter × 100

[0053] The experimental results showed that the quality of mycelium produced by Trichoderma WDNT01 after fermentation in liquid culture medium with different carbon sources was different. Glycerol, lactose and fructose were conducive to mycelium production. The drying weight of mycelium after fermentation for 5 days was 1.03×10 -2 g, 0.94×10 -2 g, 0.90×10 -2 g, which was significantly different from the other treatments. The mycelium production during maltose fermentation was the smallest, only 0.21×10 -2 g (see Table 3). The antibacterial activity of the sterile fermentation filtrates of strain WDNT01 after fermentation with seven different carbon sources against three pathogens also varied. When acting on S. rolfsii, the antibacterial activity of the fermentation filtrates of xylose, glucose, and glycerol among the seven carbon sources tested was significantly higher than that of the other treatments. Among them, the antibacterial activity of the fermentation filtrate of xylose against S. rolfsii was the highest, with an inhibition rate of 76.70% (see Table 3 and Appendix). Figure 7 When xylose was used as the carbon source, the fermentation filtrate of WDNT01 had the highest antibacterial activity against pepper wilt pathogen and kiwifruit soft rot pathogen, with the inhibition rates of 46.45% and 70.93% respectively (see Table 3 and Appendix). Figure 7 Therefore, xylose is the optimal carbon source for fermentation of strain WDNT01.

[0054] Table 3 Effects of different carbon sources on the antibacterial activity of fermentation filtrate and mycelium quality

[0055]

[0056]

[0057] Note: Lowercase letters indicate significant differences (p<0.05).

[0058] 2. Effects of different nitrogen sources on the antibacterial activity of Trichoderma WDNT01 fermentation products

[0059] The fermentation basal medium for different nitrogen sources was Sabouraud broth (SD). The carbon source in Sabouraud broth was replaced with xylose. The nitrogen source, peptone, was then replaced with sodium nitrate, ammonium chloride, ammonium acetate, ammonium nitrate, glycine, tyrosine, tryptophan, and urea, respectively, to create nine fermentation media with different nitrogen sources. Each treatment was replicated three times, with a 60% liquid volume. Each flask was inoculated with four bacterial cakes and then shaken at 150 rpm and 25°C for 5 days. Subsequent experimental treatments were the same as those for the different carbon source groups.

[0060] The experimental results showed that the quality of mycelium produced by Trichoderma WDNT01 after fermentation in liquid culture medium with different nitrogen sources was significantly different. Among them, peptone and tyrosine were the most conducive to mycelium production. The drying weight of mycelium after 5 days of fermentation was 0.76×10 -2 g and 0.54×10 -2 g, which was significantly higher than that of other treatments. However, the mycelium production was the smallest when ammonium acetate was used as nitrogen source, which was only 0.14×10 -2 g (see Table 4). The antibacterial activity of the sterile fermentation filtrates of strain WDNT01 after fermentation with 9 different nitrogen sources against the three pathogens also varied. When acting on Sclerotium rolfsii of pepper, the antibacterial activity of the fermentation filtrates of ammonium acetate and peptone among the 9 nitrogen sources tested was significantly higher than that of the other treatments, with inhibition rates of 87.35% and 72.53%, respectively (see Table 4 and Appendix). Figure 8 When used as nitrogen source for pepper wilt pathogen, the antibacterial activity of WDNT01 fermentation filtrate was significantly higher than that of other treatments, with an inhibition rate of 51.04%, which was significantly higher than that of other treatments (see Table 4 and Appendix Figure 8 b); When used against kiwifruit soft rot pathogen, the fermentation filtrates of tyrosine, glycine, peptone and tryptophan among the nine nitrogen sources tested had significantly higher antibacterial activities than those of the other treatments, with inhibition rates of 72.54%, 64.69%, 62.42% and 61.16%, respectively (see Table 4 and Appendix Figure 8 (c)

[0061] In summary, peptone was the most suitable nitrogen source for fermentation of WDNT01 mycelium production and had the best inhibitory effect on the three pathogens. Therefore, it was clear that peptone was the optimal nitrogen source for fermentation of strain WDNT01.

[0062] Table 4 Effects of different nitrogen sources on the antibacterial activity of fermentation filtrate and mycelium quality

[0063]

[0064] Note: Lowercase letters indicate significant differences (p<0.05).

[0065] Example 3 Field control efficacy of biocontrol Trichoderma WDNT01 against pepper white rot

[0066] 1. Preparation of biocontrol bacteria fermentation liquid

[0067] Based on the above research, the fermentation conditions of Trichoderma WDNT01 were clarified as follows: 60% liquid volume, 1% inoculum volume, 40 g / L xylose as the main fermentation nutrient, 10 g / L ammonium acetate, 1 L distilled water, pH 5-6, 150 r / min, 25 ° C fermentation for 5 days, and dilution preparation after fermentation to make the spore content 1×10 6 pcs / ml.

[0068] 2. Application of biocontrol bacteria

[0069] (1) Application of Trichoderma WDNT01 on peppers

[0070] Before transplanting each pepper plant, the planting hole was first filled with spores containing 1×10 6 Plants were transplanted and covered with soil after being treated with 50 mL of Trichoderma WDNT01 fermentation solution (containing 50 mL of Trichoderma WDNT01 per plant). A control plot was used. Before peppers bloomed, roots were irrigated with 50 mL of Trichoderma WDNT01 fermentation solution per plant. Another biocontrol agent was applied before the peppers set fruit. Fourteen days after the final application, the occurrence of pepper wilt and white rot in the experimental plots was investigated, and disease index and control efficacy were calculated.

[0071] Pepper wilt disease grading: Level 0 - the plant grows normally; Level 1 - the base of the plant stem turns brown and shrinks, the leaves become yellow and reticular, and a quarter of the leaves are withered, with little impact on fruiting; Level 2 - the base of the plant stem turns brown and shrinks, the leaves become yellow and reticular, and half of the leaves are withered, and the fruit grows slowly; Level 3 - three-quarters of the leaves on the plant wilt from bottom to top, or the side branches wilt; Level 4 - the whole plant wilts, and it produces very little fruit, or even no fruit or dies.

[0072] Grading of pepper white rot disease: Level 0 - normal plant growth; Level 1 - the base of the stem turns dark brown in the early stage of plant infection, and less than a quarter of the leaves on the plant wilt; Level 2 - white silky mycelium grows at the discolored part of the stem base, extending radially to the surrounding areas, and half of the leaves on the plant wilt; Level 3 - the mycelium in the diseased part and the surface mycelium produce brown cabbage seed-like sclerotia, and less than three-quarters of the leaves on the plant wilt; Level 4 - more than three-quarters of the leaves on the plant wilt and fall off, and the entire plant dies.

[0073] (2) Application of Trichoderma WDNT01 on kiwifruit

[0074] Before closing the garden in winter and during the budding period, the spore concentration was 1×10 6 Root irrigation with Trichoderma WDNT01 fermentation solution (2L / mL) was applied to each kiwifruit plant. During the middle and late stages of fruiting, the sterile fermentation solution was sprayed on the fruit in the evening. This improved the soil microecology and enhanced tree vigor, thereby enhancing the plant's disease resistance. After harvesting, the kiwifruit was left at room temperature for 5 days before observing and investigating the disease.

[0075] Kiwifruit soft rot disease grading: Level 0 - normal fruit, no lesions; Level 1 - fruit lesion area accounts for <5% of the fruit surface area; Level 3 - fruit lesion area accounts for 6% to 10% of the fruit surface area; Level 5 - fruit lesion area accounts for 11% to 20% of the fruit surface area; Level 7 - fruit lesion area accounts for 21% to 50% of the fruit surface area; Level 7 - fruit lesion area accounts for >50% of the fruit surface area.

[0076]

[0077]

[0078] 3. Prevention effectiveness investigation

[0079] Investigation results: A field investigation on pepper white rot and wilt was conducted 14 days after the last application of Trichoderma WDNT01 optimized fermentation liquid. The investigation results showed that the diseased plant rate and disease index of both diseases decreased after treatment with the biocontrol Trichoderma WDNT01 fermentation liquid. The diseased plant rate and disease index of white rot in the biocontrol treatment area were 5.90% and 2.78, respectively, while the diseased plant rate and disease index of pepper white rot in the diseased control area were 13.89% and 11.89, respectively. The field control efficacy of Trichoderma WDNT01 against pepper white rot was 76.34%; the diseased plant rate and disease index of wilt in the Trichoderma WDNT01 treatment area were 8.33% and 3.47, respectively, while those of the diseased control area were 22.57% and 17.71. The field control efficacy of Trichoderma WDNT01 against pepper wilt was 80.39%. An investigation of kiwifruit found that the incidence of soft rot in kiwifruit was also reduced after treatment with the biocontrol Trichoderma WDNT01 fermentation liquid. The diseased plant rate and disease index of the biocontrol treatment were 14.00% and 4.43, respectively, while the diseased plant rate and disease index of the control were 20.50% and 11.63.

[0080] Table 6

[0081]

[0082] In summary, the survey results showed that the optimized fermentation broth of strain WDNT01 had good antibacterial effects against pathogens of pepper white rot, pepper wilt, and kiwifruit soft rot, with a broad antibacterial spectrum, and is a biocontrol fungus resource worthy of in-depth study.

[0083] The various embodiments in this specification are described in a progressive manner, and each embodiment focuses on the differences from other embodiments. The same or similar parts between the various embodiments can be referenced to each other.

[0084] The above description of the disclosed embodiments is intended to enable one skilled in the art to implement or use the present invention. Various modifications to these embodiments will be readily apparent to one skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention is not limited to the embodiments shown herein but is intended to conform to the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A strain with broad-spectrum biocontrol effects Trichoderma pholiotae strain WDNT01, characterized in that The strain WDNT01 was deposited with CCTCC NO: M 20231270 on July 12, 2023, at the China Center for Type Culture Collection, Wuhan University, Wuhan, China, and was classified as Trichoderma pholiotae .

2. A microbial agent, characterized in that: The invention comprises the strain WDNT01 according to claim 1.

3. The microbial agent according to claim 2, characterized in that The bacterial agent dosage form is bacterial suspension, powder, tablet or fermentation filtrate.

4. Use of the strain WDNT01 according to claim 1 or the microbial agent according to claim 2 or 3, characterized in that: The application is to prevent and control pepper white rot, pepper wilt or kiwifruit soft rot.

5. The use according to claim 4, characterized in that The application is to apply the fermentation filtrate to the roots of peppers or kiwifruit.

6. The use according to claim 4, characterized in that In the application, xylose is used as a carbon source and peptone is used as a nitrogen source for the preparation of the fermentation filtrate.

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