Co-culture fermentation liquid of Trichoderma and Burkholderia, bacterial agent and application thereof in preventing and controlling root rot of ginger

Abstract

The present invention relates to the field of biological control technology, and in particular to a co-culture fermentation liquid of Trichoderma and Burkholderia, a bacterial agent and an application thereof in preventing and treating ginger root rot. The preparation method of the co-culture fermentation liquid comprises: Trichoderma atroviride ) HB20111 and Burkholderia vietnamese ( Burkholderia vietnamiensis ) P418 co-culture fermentation, to obtain a co-culture fermentation liquid; the co-culture fermentation liquid contains fermented bacteria, fermentation supernatant and metabolites produced by co-culture. Compared with a single biocontrol strain, the dark green Trichoderma HB20111 and Vietnamese Burkholderia P418 used in the present invention can produce a wide range of antibacterial and growth-promoting secondary metabolites through co-culture fermentation, prevent and control ginger root rot, are not easy to produce drug resistance, have significant effects, and have high application value.

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 in preventing and controlling ginger root rot. Background Art

[0002] In recent years, biological control has received increasing attention and plays a key role in controlling plant diseases. Secondary metabolites are a class of low molecular weight compounds produced by microorganisms. Secondary metabolites such as antibiotics, alkaloids, terpenoids, flavonoids, lipopeptides and polyketides can not only regulate the responses of individuals and populations, but also shape the entire microbial community. With the development of genome sequencing technology, a large number of biosynthetic gene clusters for the synthesis of putative secondary metabolites have been identified in microbial genomes. However, there is a large gap between the genomic capacity of biosynthetic gene clusters and the secondary metabolites found in microorganisms, because many biosynthetic gene clusters remain silent or at low expression levels under normal laboratory culture conditions.

[0003] Co-cultivation of different species of microorganisms has become an effective method for mining new secondary metabolites. Co-cultivation simulates the growth environment of natural microbial communities and utilizes the complex ecological relationships between co-cultivated members, including synergistic metabolism, mutualism, competition and antagonism, and interactions between signal molecules, to activate the expression of silent biosynthetic gene clusters and induce the synthesis of new secondary metabolites. Velmourougane et al. observed that the biofilm produced by the co-cultivation of Trichoderma viride ITCC 2211 and Azotobacter foetida W5 helped the two to better colonize in the chickpea rhizosphere. Compared with single inoculation, co-cultivation increased the root biomass, plant enzyme activity and soil available nutrients of chickpeas, and could save 25% of nitrogen fertilizer application. Li Tingting et al. optimized the co-cultivation conditions of Trichoderma viride SG3403 and Bacillus subtilis 22 to secrete antifungal secondary metabolites, and found that the inhibition rate of the co-cultivation fermentation filtrate on Fusarium graminearum increased by 54.22%. Secondary metabolites such as mevastatin and Koninginin A in the co-cultivation were identified by LC-MS analysis. Yang Kai et al. used metabolomics to analyze the growth-promoting effect of the fermentation filtrate of co-culture of Trichoderma harzianum LTR-2 and ureabacter ureagenes DnL1-1 on wheat, and found that secondary metabolites such as amino acids produced by co-culture had a significant promoting effect on wheat seed germination.

[0004] As important biocontrol strains, Trichoderma and Burkholderia have good control effects on plant diseases caused by a variety of soil-borne pathogenic fungi. However, there is no research on the co-cultivation of Trichoderma and Burkholderia. Summary of the invention

[0005] In view of the current lack of technical problems related to the co-cultivation of Trichoderma and Burkholderia, the present invention provides a co-cultivation fermentation liquid of Trichoderma and Burkholderia, a bacterial agent and the application thereof in preventing and treating ginger root rot. Compared with a single biocontrol strain, the co-cultivation fermentation produces a wide range of antibacterial and growth-promoting secondary metabolites, is not easy to produce drug resistance in preventing and treating ginger root rot, has a significant effect, and has a high application value.

[0006] In a first aspect, the present invention provides a co-culture fermentation broth of Trichoderma and Burkholderia, the preparation method comprising:

[0007] Dark green Trichoderma 绿木霉 ) HB20111 and Burkholderia vietnamese ( 越南伯克霍尔德氏菌 ) P418 co-culture fermentation to obtain co-culture fermentation liquid, the co-culture fermentation conditions are:

[0008] 28~30℃, 180~200rpm shaking culture for 5~7 days;

[0009] Dark green Trichoderma HB20111 was deposited in the General Microbiological Center of China Microbiological Culture Collection Administration on December 5, 2018. The deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing. The deposit number is CGMCC No.16963, and the classification name is dark green Trichoderma 绿木霉 ;

[0010] Vietnamese Burkholderia P418 was deposited in the General Microbiology Center of China Microbiological Culture Collection Committee on August 30, 2004. The deposit address is No. 13, Zhongguancun North 1st Road, Haidian District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, with the deposit number CGMCC No. 1212 and the classification name is Vietnamese Burkholderia 伯克霍尔德氏菌属 越南种 .

[0011] Furthermore, Trichoderma aureum HB20111 and Burkholderia vietnam P418 were inoculated into MKB fermentation medium at a live bacterial count of 1:1 for co-culture and fermentation.

[0012] Further, the co-culture fermentation method is as follows:

[0013] (1) Strain activation: Inoculate dark green Trichoderma HB20111 onto PDA medium and place it in a constant temperature incubator at 28°C for 5 days for activation;

[0014] Pick a colony of Burkholderia vietnamese P418 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 (usually 24-48 hours);

[0015] (2) Seed solution preparation: Scrape the spores of Trichoderma HB20111 with sterile water and dilute to a concentration of 1.0×10 8 cfu / mL, as seed solution A;

[0016] 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;

[0017] (3) Co-culture fermentation: The seed solutions A and B obtained in step (2) were inoculated into the MKB fermentation medium at an inoculation rate of 10% (volume percentage) respectively, and co-culture fermentation was carried out under the following culture conditions: 28-30°C, 180-200 rpm shaking culture for 5-7 days to obtain a co-culture fermentation liquid.

[0018] 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:

[0019] Cyclic (L-pro-L-tyro) dipeptide, cyclo (L-pro-L-valine) dipeptide, cyclo (D-leu-L-pro) dipeptide, indoleacetic acid, indole-2-acetic acid, DL-indole-3-lactic acid, natamycin, amikacin, ofloxacin and bafilomycin B1.

[0020] In a second aspect, the present invention provides an application of the co-cultured fermentation liquid of the above-mentioned Trichoderma and Burkholderia to prevent and treat ginger root rot, wherein the pathogen of ginger root rot is Fusarium oxysporum ginger-specific type ( 尖孢镰刀菌 f.sp. 姜属 , Foz).

[0021] In a third aspect, the present invention provides a co-culture bacterial agent of Trichoderma and Burkholderia, wherein a carrier is used to adsorb the co-culture fermentation liquid of the Trichoderma and Burkholderia to form a solid bacterial agent, and the solid bacterial agent is mixed with a binder to prepare the co-culture bacterial agent.

[0022] 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.

[0023] 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.

[0024] In a fourth aspect, the present invention provides an application of the co-cultured bacterial agent of Trichoderma and Burkholderia for preventing and treating ginger root rot, wherein the pathogen of ginger root rot is Fusarium oxysporum ginger-specific type.

[0025] Furthermore, the co-cultivation bacterial agent can also promote the growth of ginger and increase the yield of ginger.

[0026] The beneficial effects of the present invention are:

[0027] Compared with most "composite bacterial agent" products obtained by culturing and then mixing single bacteria, the dark green Trichoderma HB20111 and Vietnamese Burkholderia P418 in the co-culture fermentation liquid and co-culture bacterial 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.

[0028] Compared with single bacteria culture, the co-culture fermentation of dark green Trichoderma HB20111 and Vietnamese Burkholderia P418 of the present invention increases the yield of secondary metabolites that inhibit pathogenic fungi and promote plant growth. Among them, natamycin, amikacin, ofloxacin and bafilomycin B1 and cyclic dipeptides such as cyclic (L-pro-L-valine) dipeptide, cyclic (L-pro-L-tyro) dipeptide and cyclic (D-leucine-L-pro) dipeptide show a strong inhibitory effect on the ginger root rot pathogen Fusarium oxysporum ginger special type, thereby achieving the effect of preventing and controlling ginger root rot.

[0029] At the same time, both dark green Trichoderma HB20111 and Vietnamese Burkholderia 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 DL-indole-3-lactic acid in the rhizosphere of plants. They can induce ginger to produce systemic defense responses and disease resistance, and have a nutritional superposition effect, thereby improving the microecological environment of ginger-growing soil, improving soil structure and physical and chemical properties, and increasing the biological yield and quality of ginger.

[0030] 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

[0031] 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.

[0032] Figure 1 It is the culture result of Trichoderma aureum HB20111 and Burkholderia vietnamese P418; Figure 1 In the figure, A is the result of single bacterial culture of P418; B is the result of single bacterial culture of HB20111; C is the result of co-culture of HB20111+P418.

[0033] Figure 2 The results of the plate inhibition of Trichoderma aureum HB20111 and Burkholderia vietnam P418 against Fusarium oxysporum ginger-specific type Foz; Figure 2 In the figure, A is the antibacterial result of co-culture fermentation filtrate on Foz; B is the antibacterial result of HB20111 single bacteria fermentation filtrate on Foz; C is the antibacterial result of P418 single bacteria fermentation filtrate on Foz; D is the Foz blank plate control. DETAILED DESCRIPTION

[0034] 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.

[0035] The dark green Trichoderma HB20111 used in the present invention has been deposited in the General Microbiological Center of the China Microbiological Culture Collection Administration on December 5, 2018, with the deposit address being No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number being CGMCC No. 16963, and the classification name being dark green Trichoderma 绿木霉 , which has been disclosed in the Chinese invention patent CN110564627B. Vietnamese Burkholderia P418 was deposited in the General Microbiological Center of the China Microbiological Culture Collection Administration on August 30, 2004. The deposit address is No. 13, Zhongguancun North 1st Road, Haidian District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, with the deposit number CGMCC No. 1212 and the classification name is Vietnamese Burkholderia 越南伯克霍尔德氏菌 , the strain has been disclosed in Chinese invention patent CN103478147B.

[0036] Each culture medium used in the present invention can be obtained by a homemade method, or commercially available culture medium products can be directly purchased. The PDA culture medium components include 20% peeled potatoes, 2% glucose, 1.5% agar powder, and 1L distilled water, with a natural pH. The TY culture medium components include 1% tryptone, 0.5% yeast powder, 0.02% calcium chloride, 1.5% agar powder, and 1L distilled water, with a pH of 7.2 to 7.4. The TY liquid culture medium components include 1% tryptone, 0.5% yeast powder, 0.02% calcium chloride, and 1L distilled water, with a pH of 7.2 to 7.4. The MKB fermentation culture medium components include 2% casein hydrolyzate, 1% glycerol, 0.15% dipotassium hydrogen phosphate, 0.15% magnesium sulfate, and 1L distilled water, with a pH of 7.2±0.2.

[0037] Example 1 Preparation of Trichoderma and Burkholderia co-culture fermentation broth and fermentation filtrate

[0038] (1) Strain activation: Inoculate dark green Trichoderma HB20111 onto PDA medium and place it in a constant temperature incubator at 28°C for 5 days for activation.

[0039] Pick a colony of Burkholderia vietnamese P418 and streak it on TY medium, then place it in a constant temperature incubator at 30°C in the dark and culture it until a single colony grows out.

[0040] (2) Seed solution preparation: Scrape the dark green Trichoderma HB20111 spores with sterile water and dilute to 1.0×10 8 cfu / mL, as seed solution A.

[0041] 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.

[0042] (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 single culture plates of Trichoderma HB20111, single culture plates of Burkholderia vietnamese P418, and co-culture plates of Trichoderma HB20111 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 and Burkholderia, and they can coexist symbiotically.

[0043] (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 dark green Trichoderma HB20111 and Vietnamese Burkholderia P418, fermentation supernatant and metabolites produced therefrom.

[0044] (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.

[0045] 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) respectively 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 HB20111 single bacterial fermentation liquid and P418 single bacterial fermentation liquid. The HB20111 single bacterial fermentation liquid and P418 single bacterial fermentation liquid were filtered and sterilized with a 0.22 μm microporous filter membrane to obtain HB20111 single bacterial fermentation filtrate and P418 single bacterial fermentation filtrate.

[0046] (6) Analysis and identification of co-culture metabolites: The co-culture fermentation filtrate, HB20111 single-bacteria fermentation filtrate, and P418 single-bacteria fermentation filtrate were placed in a freeze dryer and freeze-dried. 100 μL of 80% methanol aqueous solution was added and vortexed for 30 seconds. After standing in an ice bath for 5 minutes, the mixture was 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%. The mixture was centrifuged at 4°C and 12,000 rpm for 15 minutes. The supernatant was sampled for LC-MS analysis and identification.

[0047] The differential metabolites produced by co-culture fermentation of Trichoderma and Burkholderia are shown in Table 1 below. It can be seen that co-culture fermentation significantly increased the contents of various substances such as natamycin, amikacin, ofloxacin, bafilomycin B1, cyclo(L-pro-L-tyro) dipeptide, cyclo(L-pro-L-valerian) dipeptide, cyclo(D-leucine-L-pro) dipeptide, indoleacetic acid (IAA), indole-2-carboxylic acid (ICA) and DL-indole-3-lactic acid (ILA). Among them, natamycin, amikacin, ofloxacin and bafilomycin B1 have antifungal effects, and IAA, ICA and ILA have plant growth-promoting effects, which lays a foundation for the development of co-culture bacterial agents.

[0048] Table 1 Differential metabolites produced by co-culture fermentation of Trichoderma and Burkholderia

[0049]

[0050] (7) Determination of the antagonistic activity of co-culture metabolites against Fusarium oxysporum: The co-culture fermentation filtrate, HB20111 single-bacteria fermentation filtrate, and P418 single-bacteria fermentation filtrate were mixed with PDA medium at 10% (volume percentage) and poured into plates as three treatment groups. PDA medium supplemented with 10% sterile water was used as a blank plate control. The ginger-specific Foz bacterial cake of Fusarium oxysporum was inoculated in the center of each plate and cultured at 28°C for 5 days. The results are as follows: Figure 2 shown.

[0051] The cross method was used to measure the distance between the center and edge hyphae of the ginger-specific type of Fusarium oxysporum colony, and the inhibition rate of the ginger-specific type of Fusarium oxysporum was calculated. The calculation formula is as follows:

[0052] .

[0053] After calculation, the inhibition rate of the co-culture fermentation filtrate on the ginger-specific type of Fusarium oxysporum reached 94.1%, which was significantly higher than the 88.02% of the HB20111 single-bacteria fermentation filtrate and the 18.19% of the P418 single-bacteria fermentation filtrate. This shows that under the co-culture fermentation mode, Trichoderma HB20111 and Burkholderia vietnam P418 can produce more secondary metabolites with antibacterial effects, and improve the antagonistic activity against the ginger-specific type of Fusarium oxysporum Foz.

[0054] Example 2 Preparation of Trichoderma and Burkholderia co-culture agent

[0055] 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 dark green Trichoderma HB20111 and Vietnamese Burkholderia P418, fermentation supernatant, metabolites produced therefrom, and other components.

[0056] 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 dark green Trichoderma HB20111 and Vietnamese Burkholderia P418, fermentation supernatant, metabolites produced therefrom, and other components.

[0057] 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. The co-culture bacterial agent C contained fermentation cells of dark green Trichoderma HB20111 and Vietnamese Burkholderia P418, fermentation supernatant, metabolites produced therefrom, and other components.

[0058] Example 3 Application of Trichoderma and Burkholderia co-culture agent in preventing and controlling ginger root rot

[0059] Experimental location: Zhaili Town, Laiwu District, Jinan City, Shandong Province, a ginger-growing area with a high incidence of ginger root rot in the early stage, with an experimental area of ​​100 square meters.

[0060] Test crop: Laiwu ginger.

[0061] The experiment was divided into three treatment groups: blank control group, carbendazim treatment group (positive control) and co-culture bacterial agent treatment group.

[0062] The blank control group did not receive any pretreatment.

[0063] Carbendazim-treated group: During the ginger sowing period, 50% carbendazim wettable powder was mixed with soil at a dosage of 2kg / mu and applied to plant ginger seed buds.

[0064] Co-cultivation bacterial agent treatment group: During the ginger sowing period, 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 ginger seed sprouts were planted.

[0065] The planting and management of ginger in the three treatment groups were consistent with local cultivation conditions. 180 days after planting, the protective efficacy and growth-promoting effect of the co-cultivated bacterial agent on ginger field plots were investigated and tested.

[0066] The root rot of ginger was investigated plant by plant according to the 7-level classification method. The classification standards for the severity of the disease are as follows:

[0067] Level 0: The whole plant is disease-free;

[0068] Level 1: The base of the stem is slightly discolored, or less than 1 / 4 of the leaves are slightly faded or wilted;

[0069] Level 3: The base of the stem is obviously discolored, or 1 / 4 to 1 / 2 of the leaves are slightly wilted;

[0070] Level 5: Browning at the base of the stem covering 1 / 2 of the entire stem, or more than 2 / 3 of the leaves withering;

[0071] Level 7: All diseased plants wilt or die.

[0072] The disease index and disease prevention effect are calculated according to the following formula:

[0073] ;

[0074] .

[0075] The field plot control results of ginger root rot are shown in Table 2 below. It can be seen that the co-cultured bacterial agent of the present invention can use dark green Trichoderma HB20111 and Vietnamese Burkholderia P418 to build a microecological balance in the soil. The antibacterial substances produced by the two can also antagonize the growth of pathogens and effectively reduce the incidence of ginger root rot. The control effect of ginger root rot reached 77.05%, which is significantly higher than the 63.88% of the chemical pesticide carbendazim treatment. The application of the co-cultured bacterial agent can also significantly improve the growth indicators of ginger. The plant height and single plant yield are significantly different from the blank control ( p <0.05), and the promotion effect on ginger plant height and single plant yield increased by 23.87% and 28.30% respectively compared with the control group; and significantly higher than the carbendazim treatment (18.83% and 24.76% respectively). This shows that the application of co-cultured bacterial agents can not only effectively prevent and control ginger root rot, but also play a significant role in promoting ginger yield, thereby reducing the use of pesticides and fertilizers, and has great application value and economic and social benefits.

[0076] Table 2 Statistics of ginger field plot test

[0077]

[0078] 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: The preparation method comprises: Dark green Trichoderma Trichoderma atroviride ) HB20111 and Burkholderia vietnamese ( Burkholderia vietnamiensis ) P418 co-culture fermentation to obtain co-culture fermentation liquid, the co-culture fermentation conditions are: 28~30℃, 180~200rpm shaking culture for 5~7 days; Dark green Trichoderma HB20111 was deposited in the General Microbiological Center of China Microbiological Culture Collection Administration on December 5, 2018. The deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing. The deposit number is CGMCC No. 16963 and the classification name is dark green Trichoderma Trichoderma atroviride ; Vietnamese Burkholderia P418 was deposited in the General Microbiology Center of China Microbiological Culture Collection Committee on August 30, 2004. The deposit address is No. 13, Zhongguancun North 1st Road, Haidian District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, with the deposit number 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 Dark green Trichoderma HB20111 and Vietnamese Burkholderia P418 were inoculated into MKB fermentation medium at a live bacterial count of 1:1 for co-culture and fermentation.

3. An application of the co-culture fermentation liquid as claimed in any one of claims 1 to 2 for preventing and treating ginger root rot, characterized in that: The pathogen of ginger root rot is Fusarium oxysporum, a special strain of ginger ( Fusarium oxysporum f. sp. Zingiberi ).

4. A co-culture agent of Trichoderma and Burkholderia, characterized in that: The co-culture fermentation liquid according to any one of claims 1 to 2 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.

5. The co-cultivation bacterial agent according to claim 4, 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.

6. The co-cultivation bacterial agent according to claim 4, 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.

7. An application of the co-cultured bacterial agent according to any one of claims 4 to 6 for preventing and treating ginger root rot, characterized in that: The pathogen of ginger root rot is Fusarium oxysporum, a ginger-specific type.

8. The use according to claim 7, characterized in that The application of co-culture bacterial agents in promoting ginger growth and increasing ginger yield.

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