Trichoderma and burkholderia co-culture fermentation broth, microbial inoculum and application of trichoderma and burkholderia co-culture fermentation broth and microbial inoculum

Abstract

The invention relates to the technical field of biological control, in particular to co-culture fermentation liquor of trichoderma and burkholderia, a microbial inoculum and application of the co-culture fermentation liquor and the microbial inoculum. The co-culture fermentation liquor is obtained by co-culture and fermentation of trichoderma harzianum T11-W and Burkholderia Vietnamese P418; wherein the preservation number of the trichoderma harzianum T11-W is CGMCC (China General Microbiological Culture Collection Center) No.7938, and the preservation number of the Burkholderia Vietnamese P418 is CGMCC No.1212. Compared with a single biocontrol strain, bacteriostatic and growth-promoting secondary metabolites generated by co-culture fermentation are wide, resistance to the tomato root knot nematode disease is not easy to generate, and the effect is remarkable; compared with a composite microbial agent obtained by mixing after single-bacterium culture, the two strains of the co-cultured fermentation liquor / microbial agent have no antagonism, and the effects of the two strains of the co-cultured fermentation liquor / microbial agent are mutually superposed and enhanced.

Description

Technical Field

[0001] The invention relates to the technical field of biological control, and in particular to a co-culture fermentation liquid of Trichoderma and Burkholderia, a bacterial agent and application thereof. Background Art

[0002] Root-knot nematodes ( Meloidogyne spp.) is a highly specialized omnivorous plant pathogenic nematode with a fast reproduction rate, strong ecological adaptability, and a wide host range, which seriously threatens the sustainable development of facility agriculture. At present, the prevention and control of root-knot nematodes mainly relies on chemical control. Due to the problems of high toxicity, high residue, and easy environmental pollution, the use of biological control to replace chemical control has received more and more attention. A lot of research has been done on the biological control of root-knot nematodes, but because the control effect of single biological control bacteria in the field is unstable, most of them remain in the experimental stage. The research and development of high-efficiency composite bacterial agents has become a key scientific issue that needs to be solved in the biological control of nematodes.

[0003] As important biocontrol strains, Trichoderma and Burkholderia have good control effects on plant diseases caused by root-knot nematodes. CN 105660256 A discloses a composite microbial agent for antagonizing tomato root-knot nematodes, which can be prepared by using spore fermentation liquid of Vietnamese Burkholderia P418, spore powder of Trichoderma harzianum T11-W and spore fermentation liquid of Trichoderma viridis L-10 in a mass ratio of 1-6:1-8:1-3. The composite microbial agent can prevent and control the infection and occurrence of tomato root-knot nematode disease, but has limited growth-promoting effect on tomatoes when used alone, and cannot significantly increase tomato yield.

[0004] Based on this, it is necessary to provide a microbial preparation that can not only prevent and control tomato root-knot nematode diseases, but also provide a variety of plant hormones and antifungal compounds for tomato growth. Summary of the invention

[0005] In view of the current technical problem of lack of microbial preparations that can both prevent and control tomato root-knot nematode diseases and provide plant hormones and antifungal compounds, the present invention provides a co-culture fermentation liquid of Trichoderma and Burkholderia, a bacterial agent and its application. Compared with a single biocontrol strain, the antibacterial and growth-promoting secondary metabolites produced by co-culture fermentation are extensive, and it is not easy to produce drug resistance in the prevention and control of tomato root-knot nematode diseases, and the effect is significant; compared with the traditional "composite microbial agent" product obtained by mixing after single bacterial culture, there is no antagonism between the two strains of the co-culture fermentation liquid / microbial agent of the present invention, and the efficacy of the two is superimposed and enhanced, the preparation process is simpler, and it has a high application value.

[0006] The technical solution of the present invention is as follows: In a first aspect, the present invention provides a co-culture fermentation broth of Trichoderma and Burkholderia, comprising Trichoderma harzianum (Trichoderma harzianum )T11-W and Burkholderia vietnamese ( Burkholderia vietnamiensis ) P418 was obtained by co-culturing and fermenting at 28-30°C and 180-200 rpm for 5-7 days; Among them, Trichoderma harzianum T11-W was deposited in the General Microbiological Center of China Microbiological Culture Collection Administration on July 19, 2013, with the deposit address at No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No. 7938, and the classification name is Trichoderma harzianum Trichoderma harzianum ; Vietnamese Burkholderia P418 was deposited in the General Microbiological Center of China Microbiological Culture Collection Administration on August 30, 2004. The deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing. The deposit number is CGMCC No. 1212 and the classification name is Vietnamese Burkholderia Burkholderia vietnamiensis .

[0007] Furthermore, Trichoderma harzianum T11-W and Burkholderia vietnamensis P418 were inoculated into MKB fermentation medium at a live bacterial count of 1:1 for co-culture and fermentation.

[0008] Furthermore, the concentration was 1.0×10 8 cfu / mL of Trichoderma harzianum T11-W seed solution and a concentration of 1.0×10 8 The seed liquid of Burkholderia vietnamese P418 with a volume percentage of 10% was inoculated into MKB medium for co-cultivation.

[0009] Furthermore, the co-culture fermentation broth includes fermentation cells, fermentation supernatant and metabolites produced by co-culture, and the metabolites include the following main secondary metabolites: Cyclic (L-pro-L-tyro) dipeptide, cyclo (L-pro-L-valine) dipeptide, cyclo (D-leu-L-pro) dipeptide, indoleacetic acid, indole-2-carboxylic acid, indole-3-acrylic acid, gibberellin A4, natamycin, puromycin, ofloxacin and tanspiramycin.

[0010] In a second aspect, the present invention provides a co-culture agent of Trichoderma and Burkholderia, the preparation method of which is: The co-culture fermentation liquid of the Trichoderma and Burkholderia is adsorbed by a carrier to form a solid bacterial agent, and the solid bacterial agent is mixed with a binder to prepare a co-culture bacterial agent.

[0011] Furthermore, the carrier is selected from diatomaceous earth, medical stone or peat soil, and the mass ratio of the fermentation liquid to the carrier is 5:1.

[0012] Furthermore, the binder is selected from sodium hydroxymethyl cellulose and / or sodium alginate, and the mass ratio of the solid bacterial agent to the binder is 100:6.

[0013] In a third aspect, the present invention also provides an application of the co-culture fermentation liquid and / or co-culture bacterial agent of the above-mentioned Trichoderma and Burkholderia, which is used to prevent and control tomato root-knot nematode diseases and promote tomato growth, thereby increasing tomato yield.

[0014] The beneficial effects of the present invention are: Compared with most "composite microbial agent" products obtained by culturing and then mixing single bacteria, the Trichoderma harzianum T11-W and Burkholderia vietnam P418 in the co-culture fermentation broth and co-culture agent of the present invention can coexist symbiotically and their effects are superimposed on each other. There will be no antagonism and inhibition between the two, which will affect their respective growth.

[0015] Compared with single bacteria culture, the co-culture fermentation of Trichoderma harzianum T11-W and Burkholderia vietnamensis P418 of the present invention increases the yield of secondary metabolites that inhibit pathogenic fungi and promote plant growth. Among them, natamycin, puromycin, ofloxacin and tanspiramycin and cyclic dipeptides such as cyclo (L-pro-L-valine) dipeptide, cyclo (L-pro-L-tyro) dipeptide and cyclo (D-leu-L-pro) dipeptide show strong inhibitory and killing effects on southern root-knot nematodes, thereby achieving the effect of preventing and controlling tomato root-knot nematode diseases.

[0016] At the same time, both Trichoderma harzianum T11-W and Burkholderia vietnamese P418 have rhizosphere growth-promoting effects. They can colonize and occupy positions on crop roots, inside plants or in soil ecology, and secrete secondary metabolites such as indoleacetic acid, indole-2-carboxylic acid, indole-3-acrylic acid and gibberellin A4 in the rhizosphere of plants. They can induce tomatoes to produce systemic defense responses and disease resistance, and have a nutritional superposition effect, thereby improving the microecological environment of tomato-growing soil, improving soil structure and physical and chemical properties, and increasing the biological yield and quality of tomatoes.

[0017] The co-culture fermentation liquid of Trichoderma and Burkholderia and the co-culture bacterial agent of the present invention save costs in the preparation process, have a stable fermentation method, are simple to use, and are environmentally friendly. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, for ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0019] Figure 1These are photos of the culture results of Trichoderma harzianum T11-W and Burkholderia vietnamensis P418, where A is the result of single-bacterium culture of P418, B is the result of single-bacterium culture of T11-W, and C is the result of co-culture of T11-W + P418.

[0020] Figure 2 These are photos of the inhibition and killing results of the co-culture fermentation filtrate of Trichoderma harzianum T11-W and Burkholderia vietnamese P418 on the eggs of southern root-knot nematode, among which A is the morphology of healthy eggs when the co-culture fermentation filtrate is added; B is the morphology of eggs 6 hours after the co-culture fermentation filtrate is added; C is the morphology of eggs 12 hours after the co-culture fermentation filtrate is added; D is the morphology of eggs 24 hours after the co-culture fermentation filtrate is added. DETAILED DESCRIPTION

[0021] In order to enable those skilled in the art to better understand the technical solutions in the present invention, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the drawings in the embodiments of the present invention. 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 creative work should fall within the scope of protection of the present invention.

[0022] The Trichoderma harzianum T11-W used in the present invention has been deposited in the General Microbiological Center of the China Microbiological Culture Collection Administration on July 19, 2013, with the deposit address being No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number being CGMCC No. 7938, and the classification name being Trichoderma harzianum Trichoderma harzianum , the strain has been disclosed in Chinese invention patent CN103484380B.

[0023] The Burkholderia vietnamese P418 used in the present invention was deposited in the General Microbiological Center of the China Microbiological Culture Collection Administration on August 30, 2004, with the deposit address being No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number being CGMCC No. 1212, and the classification name being Burkholderia vietnamese Burkholderia vietnamiensis , the strain has been disclosed in Chinese invention patent CN103478147B.

[0024] Each culture medium used in the present invention can be obtained using a homemade method, or commercially available culture medium products can be directly purchased. PDA culture medium components include 20% peeled potatoes, 2% glucose, 1.5% agar powder, 1L distilled water, and natural pH. TY culture medium components include 1% tryptone, 0.5% yeast powder, 0.02% calcium chloride, 1.5% agar powder, 1L distilled water, pH 7.2~7.4. TY liquid culture medium components include 1% tryptone, 0.5% yeast powder, 0.02% calcium chloride, 1L distilled water, pH 7.2~7.4. MKB fermentation culture medium components include 2% casein hydrolyzate, 1% glycerol, 0.15% dipotassium hydrogen phosphate, 0.15% magnesium sulfate, 1L distilled water, pH 7.2±0.2.

[0025] Example 1 Preparation of co-culture fermentation broth and fermentation filtrate of Trichoderma and Burkholderia (1) Strain activation: Inoculate Trichoderma harzianum T11-W onto PDA medium and place it in a constant temperature incubator at 28°C for 5 days for activation.

[0026] Pick a Vietnamese Burkholderia P418 colony and streak it on TY medium, then place it in a constant temperature incubator at 30°C in the dark until a single colony grows out (usually 24 to 48 hours).

[0027] (2) Seed solution preparation: Scrape the spores of Trichoderma harzianum T11-W with sterile water and dilute to 1.0×10 8 cfu / mL, as seed solution A.

[0028] A single colony of Burkholderia vietnamese P418 was transferred to TY liquid medium and cultured in a shaker at 30°C and 180 rpm until the logarithmic growth phase. 660 The value is about 0.6~0.8, and the concentration is 1.0×10 8 cfu / mL, as seed solution B.

[0029] (3) Antagonistic experiment between Trichoderma and Burkholderia: Three PDA culture media were prepared and inoculated with the seed solution A and seed solution B obtained in step (2) to obtain a single culture plate of Trichoderma harzianum T11-W, a single culture plate of Burkholderia vietnamese P418, and a co-culture plate of Trichoderma harzianum T11-W and Burkholderia vietnamese P418. The results of the three PDA culture media are shown in the figure. Figure 1 As shown, it is proved that there is no antagonism between Trichoderma harzianum T11-W and Burkholderia vietnamensis P418, and they can coexist symbiotically.

[0030] (4) Co-culture fermentation: The seed solution A and the seed solution B obtained in step (2) are inoculated into the same MKB fermentation medium at an inoculation rate of 10% (volume percentage), and the co-culture fermentation is carried out at 28-30° C. and 180-200 rpm with shaking for 7 days to obtain a co-culture fermentation liquid of Trichoderma and Burkholderia; the co-culture fermentation liquid comprises fermentation cells of the co-cultured Trichoderma harzianum T11-W and Burkholderia vietnamese P418, fermentation supernatant and metabolites produced therefrom.

[0031] (5) Filtration and sterilization: The co-culture fermentation broth of Trichoderma and Burkholderia is sterilized by filtering through a 0.22 μm microporous filter membrane to obtain a co-culture fermentation filtrate of Trichoderma and Burkholderia; the co-culture fermentation filtrate comprises components such as the co-culture fermentation supernatant and the generated metabolites.

[0032] At the same time, seed liquid A and seed liquid B were inoculated into two MKB fermentation mediums at an inoculation rate of 10% (volume percentage) for single bacterial culture. The fermentation conditions were the same as those in step (3), i.e., 28-30°C and 180-200 rpm shaking for 7 days to obtain T11-W single bacterial fermentation liquid and P418 single bacterial fermentation liquid. The T11-W single bacterial fermentation liquid and the P418 single bacterial fermentation liquid were filtered and sterilized with a 0.22 μm microporous filter membrane to obtain T11-W single bacterial fermentation filtrate and P418 single bacterial fermentation filtrate.

[0033] (6) Analysis and identification of co-culture metabolites: The co-culture fermentation filtrate, T11-W single-bacteria fermentation filtrate, and P418 single-bacteria fermentation filtrate were placed in a freeze dryer for freeze drying, and 100 μL of 80% methanol aqueous solution was added. The mixture was vortexed for 30 seconds, placed in an ice bath for 5 minutes, and centrifuged at 4°C and 12,000 rpm for 15 minutes. The supernatant was diluted with mass spectrometry-grade water to a methanol content of 53%, and centrifuged at 4°C and 12,000 rpm for 15 minutes. The supernatant was taken for LC-MS analysis and identification.

[0034] The differential metabolites produced by the co-culture fermentation of Trichoderma and Burkholderia are shown in Table 1. Compared with the T11-W single-bacteria fermentation broth, the co-culture fermentation produced natamycin, ofloxacin, cyclo(L-pro-L-tyro) dipeptide, cyclo(L-pro-L-valine) dipeptide, and cyclo(D-leu-L-pro) dipeptide, which were not produced by T11-W single-bacteria fermentation, and significantly increased the content of puromycin, tanspiramycin, indoleacetic acid (IAA), indole-2-carboxylic acid (IC A), indole-3-acrylic acid (IA) and gibberellin A4 (GA4); compared with the P418 single bacteria fermentation broth, the co-culture fermentation produced puromycin, tanspiramycin, IAA, ICA, IA and GA4, which were not produced by the P418 single bacteria fermentation, and significantly increased the content of substances such as natamycin, ofloxacin, cyclo(L-pro-L-tyro) dipeptide, cyclo(L-pro-L-valine) dipeptide, and cyclo(D-leu-L-pro) dipeptide. Among them, IAA, ICA, IA and GA4 have the effect of promoting plant growth, and natamycin, puromycin, ofloxacin and tanspiramycin have antifungal effects, which laid the foundation for the development of co-culture bacterial agents.

[0035] Table 1 Differential metabolites produced by co-culture fermentation of Trichoderma and Burkholderia

[0036] (7) Determination of the inhibitory effect of co-culture metabolites on the eggs of southern root-knot nematode: 50 μL of co-culture fermentation filtrate, 50 μL of sterile water and 100 μL of southern root-knot nematode egg suspension (200 eggs / mL) were added to a 24-well plate and placed in a 25°C incubator in the dark for 24 hours. During this period, the hatching of eggs was observed and recorded under an inverted microscope, and the inhibition rate of eggs was calculated. Figure 2 As shown in the figure, the test found that 6 hours after adding the co-culture fermentation filtrate, the morphology of the southern root-knot nematode eggs changed irregularly, the egg shell became thinner, and there was an obvious protrusion; after 12 hours, the protrusion began to break and the egg material flowed out; after 24 hours, the egg material completely flowed out, leaving only the egg shell, and the nematode died. It was calculated that the inhibition rate of the co-culture fermentation filtrate on the southern root-knot nematode eggs reached 93.79%.

[0037] Example 2 Preparation of co-culture agent of Trichoderma and Burkholderia Diatomaceous earth was used as a carrier to adsorb the co-culture fermentation broth of Example 1, the mass ratio of the co-culture fermentation broth to the diatomaceous earth was 5:1, and a solid bacterial agent was formed. 3wt% sodium carboxymethyl cellulose and 3wt% sodium alginate were added to the solid bacterial agent and mixed with a blender to prepare a co-culture bacterial agent A of Trichoderma and Burkholderia. The co-culture bacterial agent A contained fermentation cells of co-cultured Trichoderma harzianum T11-W and Burkholderia vietnamese P418, fermentation supernatant, metabolites produced therefrom, and other components.

[0038] Peat soil was used as a carrier to adsorb the co-culture fermentation broth of Example 1, the mass ratio of the co-culture fermentation filtrate to diatomaceous earth was 5:1, and a solid bacterial agent was formed. 6 wt % sodium carboxymethyl cellulose was added to the solid bacterial agent and mixed with a stirrer to prepare a co-culture bacterial agent B of Trichoderma and Burkholderia. The co-culture bacterial agent B contained fermentation cells of Trichoderma harzianum T11-W and Burkholderia vietnamese P418, fermentation supernatant, metabolites produced therefrom, and other components.

[0039] Medical stone was used as a carrier to adsorb the co-culture fermentation broth of Example 1, the mass ratio of the co-culture fermentation filtrate to diatomaceous earth was 5:1, and a solid bacterial agent was formed. 6 wt % sodium alginate was added to the solid bacterial agent and mixed with a stirrer to prepare a co-culture bacterial agent C of Trichoderma and Burkholderia, wherein the co-culture bacterial agent C contained fermentation cells of Trichoderma harzianum T11-W and Burkholderia vietnamese P418, fermentation supernatant, metabolites produced therefrom, and other components.

[0040] Example 3 Application of the co-cultured bacterial agent of Trichoderma and Burkholderia in the prevention and control of tomato root-knot nematode disease Experimental location: Qudi Street, Jiyang District, Jinan City, Shandong Province, a planting greenhouse with many years of continuous cropping and high incidence of soil root-knot nematodes, with an experimental area of ​​100 square meters.

[0041] Test crop: Tomato.

[0042] The experiment was divided into 6 treatment groups, including blank control group, thiazophos treatment group (positive control), P418 single bacterial agent treatment group, T11-W single bacterial agent treatment group, P418 and T11-W compound bacterial agent treatment group and co-culture bacterial agent treatment group.

[0043] The blank control group did not receive any pretreatment.

[0044] Thiabendazole treatment group: During the tomato seedling stage, 10% thiabendazole granules were mixed with soil at a dosage of 2kg / mu and applied to plant tomato seedlings.

[0045] P418 single-bacterial agent treatment group: During the tomato seedling stage, P418 single-bacterial agent was evenly applied into the soil by mixing with soil at a dosage of 20kg / mu, and tomato seedlings were transplanted.

[0046] T11-W single bacterial agent treatment group: During the tomato seedling stage, T11-W single bacterial agent was evenly applied into the soil at a dosage of 20kg / mu by mixing with the soil, and tomato seedlings were transplanted.

[0047] P418 and T11-W compound treatment group: P418 single bacterial agent and T11-W single bacterial agent were compounded in a 1:1 ratio. During the tomato seedling stage, the compound bacterial agent was evenly applied into the soil by mixing with soil at a dosage of 20kg / mu, and tomato seedlings were transplanted.

[0048] Co-cultivation bacterial agent treatment group: During the tomato seedling stage, the co-cultivation bacterial agent A prepared in Example 2 was evenly applied to the soil at a dosage of 20 kg / mu by mixing with the soil, and tomato seedlings were transplanted.

[0049] The planting and management of tomatoes in the six treatment groups were consistent with local cultivation conditions. 90 days after planting, the protective efficacy and growth-promoting effect of the co-cultured bacterial agent on tomato field plots were investigated and tested.

[0050] The tomato root-knot nematode disease was investigated plant by plant according to the 5-level classification method. The severity classification standards are as follows: Level 0: No root knot; Level 1: 0<The number of roots with knots accounts for <10% of the total root system; Level 2: 10%≤The number of roots with root knots accounts for <25% of the total root system; Level 3: 25%≤The number of roots with root knots accounts for <50% of the total root system; Level 4: 50%≤The number of roots with root knots accounts for <75% of the total root system; Level 5: The number of roots with root knots is 75% ≤ 100% of the total root system.

[0051] The disease index and disease prevention effect are calculated according to the following formula: ; .

[0052] The field plot control results of tomato root knot nematode disease are shown in Table 2 below. It can be seen that the co-cultivated bacterial agent of the present invention can use Trichoderma harzianum T11-W and Burkholderia vietnam P418 to build a microecological balance in the soil. The antibacterial substances produced by the two can also antagonize the growth of root knot nematodes and effectively reduce the incidence of tomato root knot nematode disease. The control effect of tomato root knot nematode disease reached 72.32%, which is higher than 68.90% of the chemical thiathiazolyl treatment group, 65.97% of the P418 and T11-W bacterial agent compound treatment group, 59.29% of the T11-W single bacterial agent treatment group, and 49.16% of the P418 single bacterial agent treatment group. The application of the co-cultivated bacterial agent can also significantly improve various growth indicators of tomatoes. The plant height, stem thickness, single fruit weight and single plant yield are significantly different from the blank control (p <0.05), and the promoting effects on tomato plant height, stem thickness, single fruit weight and single plant yield were increased by 19.31%, 13.70%, 18.53% and 27.71% respectively compared with the blank control group; all of them were higher than those in the thiazophos treatment group (17.23%, 12.74%, 17.73% and 23.14% respectively) and the P418 and T11-W combined treatment group (17.94%, 12.33%, 17.57% and 21.71% respectively). This shows that the application of co-cultured bacterial agents can not only effectively prevent and control tomato root-knot nematode diseases, but also play a significant role in promoting tomato yield, thereby reducing the use of pesticides and fertilizers, and has great application value and economic and social benefits.

[0053] Table 2 Statistics of tomato field plot test

[0054] Although the present invention has been described in detail with reference to the accompanying drawings and in combination with the preferred embodiments, the present invention is not limited thereto. Without departing from the spirit and essence of the present invention, a person of ordinary skill in the art may make various equivalent modifications or substitutions to the embodiments of the present invention, and these modifications or substitutions shall be within the scope of the present invention. Any person of ordinary skill in the art may easily think of changes or substitutions within the technical scope disclosed by the present invention, and these shall be within the scope of protection of the present invention.

Claims

1. A co-culture fermentation broth of Trichoderma and Burkholderia, characterized in that: By Trichoderma harzianum ( Trichoderma harzianum )T11-W and Burkholderia vietnamese ( Burkholderia vietnamiensis ) P418 was obtained by co-culturing and fermenting at 28-30°C and 180-200 rpm for 5-7 days; Among them, Trichoderma harzianum T11-W was deposited in the General Microbiological Center of China Microbiological Culture Collection Administration on July 19, 2013, with the deposit address at No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No. 7938, and the classification name is Trichoderma harzianum Trichoderma harzianum ; Vietnamese Burkholderia P418 was deposited in the General Microbiological Center of China Microbiological Culture Collection Administration on August 30, 2004. The deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing. The deposit number is CGMCC No. 1212 and the classification name is Vietnamese Burkholderia Burkholderia vietnamiensis .

2. The co-culture fermentation broth according to claim 1, characterized in that Trichoderma harzianum T11-W and Burkholderia vietnam P418 were inoculated into MKB fermentation medium at a live cell count of 1:1 for co-culture and fermentation.

3. The co-culture fermentation broth according to claim 2, characterized in that The concentration was 1.0×10 8 cfu / mL of Trichoderma harzianum T11-W seed solution and a concentration of 1.0×10 8 The seed liquid of Burkholderia vietnamese P418 with a volume percentage of 10% was inoculated into MKB medium for co-cultivation.

4. The co-culture fermentation broth according to claim 1, characterized in that The co-culture fermentation broth includes fermentation bacteria, fermentation supernatant and metabolites produced by co-culture, and the metabolites include the following main secondary metabolites: Cyclic (L-pro-L-tyro) dipeptide, cyclo (L-pro-L-valine) dipeptide, cyclo (D-leu-L-pro) dipeptide, indoleacetic acid, indole-2-carboxylic acid, indole-3-acrylic acid, gibberellin A4, natamycin, puromycin, ofloxacin and tanspiramycin.

5. A co-culture agent of Trichoderma and Burkholderia, characterized in that: The preparation method is: The co-culture fermentation liquid of Trichoderma and Burkholderia as claimed in any one of claims 1 to 4 is adsorbed on a carrier to form a solid bacterial agent, and the solid bacterial agent is mixed with a binder to prepare a co-culture bacterial agent.

6. The co-cultivation bacterial agent according to claim 5, characterized in that The carrier is selected from diatomaceous earth, medical stone or peat soil, and the mass ratio of the fermentation liquid to the carrier is 5:

1.

7. The co-culture bacterial agent according to claim 5, characterized in that The binder is selected from sodium hydroxymethyl cellulose and / or sodium alginate, and the mass ratio of the solid bacterial agent to the binder is 100:

6.

8. A use of the co-culture fermentation broth of Trichoderma and Burkholderia according to any one of claims 1 to 4 and / or the co-culture bacterial agent according to any one of claims 5 to 7, characterized in that: It is used to prevent and control tomato root-knot nematode diseases, promote tomato growth, and increase tomato yield.

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