CN114921364B - Pseudomonas brassicae, biochemical microbial inoculum and application thereof - Google Patents

Abstract

The invention discloses pseudomonas brassicae, a biochemical microbial agent and application thereof, and relates to the field of microbial application. The accession number of the bacterial strain of the pseudomonas brassicae species is: GDMCCNo.61435, which has strong biofilm formation capability, can perform rapid field planting, can effectively prevent and treat plant re-planting diseases, plant bacterial diseases, plant fungal diseases, plant soil-borne diseases and/or plant oomycete diseases, has the functions of relieving continuous cropping obstacle caused by diseases and/or soil self-toxicity, promoting seed germination, plant growth and soil improvement, and can be used for preparing biological bactericides, seed coating bactericides, root irrigation agents, plant growth regulators, soil improvers or seed soaking agents. In addition, the pseudomonas brassicae provided by the invention has good chemical compatibility, can be rapidly planted in a soil environment for long-term application of chemical, can limit pathogenic bacteria to generate drug resistance, and can generate efficient and stable biocontrol effect.

Description

A kind of Pseudomonas campestris, biochemical agent and application thereof

Technical field

The present invention relates to the field of microbial application, specifically to a kind of Pseudomonas campestris, biochemical bacteria and their application.

Background technique

In plant soils where continuous cultivation is the main method, continuous cropping obstacles/replanting problems are very likely to occur. On the one hand, the occurrence of plant continuous cropping obstacles is related to the deterioration of soil physical and chemical properties after plant continuous cropping and the autotoxicity caused by various stubble decomposition products or pesticide residues; on the other hand, it is related to the decline in the diversity of microbial populations in the soil and the abundance of beneficial microorganisms that can be cultivated. There is a significant decrease in the degree of infection and an increase in the number of harmful pathogenic microorganisms.

Studies have reported that bacterial soil-borne diseases caused by the pathogenic bacteria Ralstonia solanacearum are very likely to occur in tomato soil under long-term continuous cropping (see Zheng Xuefang, Soil Remediation of Facility Tomato Continuous Cropping Obstacles and Prevention and Control of Bacterial Wilt Disease Effect, DOI: 10.16409/j.cnki.2095-039x.2018.01.014). At present, the commonly used method to suppress Ralstonia solanacearum is to apply chemical pesticides. However, long-term application of chemical pesticides will cause irreversible damage to the soil environment, bring self-toxicity to the soil, reduce the content of beneficial bacteria in the soil, thereby exacerbating continuous cropping obstacles; on the other hand, On the one hand, pathogenic bacteria will develop resistance, causing chemical pesticides to lose their efficacy.

Pseudomonas brassicacearum is a type of plant growth-promoting rhizosphere bacteria (PGPR), which has growth-promoting and disease-preventing effects. For example, the patent CN109182199B discloses a P.b.YZX4 strain that can promote the growth and seed germination of pakchoi in saline-alkali soil by synthesizing IAA; another example is the nlsy003 strain disclosed in the patent CN103333843B, which has an inhibitory effect on fungal diseases such as rapeseed blight. However, there are significant differences in functional properties between different Pseudomonas campestris strains, so whether a specific strain has specific functions still needs to be experimentally verified (see Barquero M., Biocontrol of Fusarium oxysporum f.sp. Phaseoli and Phytophthora capsici with AutochthonousEndophytes in Common Bean and Pepper in Castilla y leon (spain), section 19.3.2, DIO: 10.1007/978-3-319-32528-6_19).

In view of this, the present invention is proposed.

Contents of the invention

The purpose of the present invention is to provide a Pseudomonas campestris with good chemical compatibility and strong biocontrol ability, biochemical inoculants and their applications to solve replanting diseases in continuous cropping soil and continuous cropping obstacles caused by soil autotoxicity. At the same time, Pseudomonas campestris is a growth-promoting bacterium with strong colonization ability. Long-term use can increase the abundance of beneficial bacteria in the soil, improve the soil/repair the soil environment, and promote plant growth and seed germination.

The invention is implemented as follows:

The invention provides the use of a bacterial strain of Pseudomonas brassicacearum species in preventing and controlling plant replanting diseases, plant bacterial diseases, plant fungal diseases, plant soil-borne diseases and/or plant oomycete diseases.

The bacterial strain of Pseudomonas campestris species was isolated by the inventor from agricultural soil collected from the sweet potato planting area in Wucun, Beizhang Township, Wenshui County, Shanxi Province (37°22′28.24″ north latitude, 112°05′34.32″ east longitude) . The strain was deposited in the Guangdong Provincial Microbial Culture Collection Center on January 15, 2021. The deposit number is: GDMCC No. 61435. The deposit address: Guangdong Institute of Microbiology, 5th Floor, Building 59, Courtyard 100, Xianlie Middle Road, Guangzhou. The biological material name is Pseudomonasbrassicacearum M29, and the taxonomic name is Pseudomonas brassicacearum. The identification result was survival.

The strain was inoculated into Luria-Bertani medium and cultured at 30°C for 1 day. The strain was round in shape, with smooth and shiny edges, dirty white color, and a diameter of 0.1cm.

The sequence of this strain was measured using 16S sequence fragments (amplification primer and sequencing primer were 27F: 5'-AGAGTTTGATCCTGGCTCAG-3' and 1492R: 5'-GGTTACCTTGTTACGACTT-3'), and the results were as shown in SEQ ID NO. 1. BLAST homology comparison determined that the nearest species of this strain is Pseudomonas brassicacearum, named Pseudomonas brassicae M29 (M29 below all refers to Pseudomonas campestris).

The SEQ ID NO.1 sequence is as follows:

AGTCGAGCGGTAGAGAGGTGCTTGCACCTCTTGAGAGCGGCGGACGGGTGAGTAAAGCCTAGGAATCTGCCTGGTAGTGGGGGATAACGCTCGGAAACGGACGCTAACCGCATACGTCCTACGGGAGAAAGCAGGGGACCTTCGGGCCTTGCGCTATCAGATGAGCCTAGGTCGGATTAGCTAGTTGGTGAGGTAATGGCTCACCAAGGCGACGATCCGTAACTGGTCTGAGAGGATGATCAGTC ACACTGGAACTGAGACACGGTCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGGACAATGGGCGAAAGCCTGATCCAGCCATGCCGCGTGTGTGAAGAAGGTCTTCGGATTGTAAAGCACTTTAAGTTGGGAGGAAGGGCATTAACCTAATACGTTAGTGTTTTGACGTTACCGACAGAATAAGCACCGGCTAACTCTGTGCCAGCAGCCGCGGTAATACAGAGGGTGCAAGCGTTAATCGGAATTACTGGGCGTAAA GCGCGCGTAGGTGGTTCGTTAAGTTGGATGTGAAATCCCCGGGCTCAACCTGGGAACTGCATTCAAAACTGTCGAGCTAGAGTATGGTAGAGGGTGGTGGAATTTCCTGTGTAGCGGTGAAATGCGTAGATATAGGAAGGAACACCAGTGGCGAAGGCGACCACCTGGACTGATACTGACACTGAGGTGCGAAAGCGTGGGGAGCAAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGTCAACTAGCCGT TGGGAGCCTTGAGCTCTTAGTGGCGCAGCTAACGCATTAAGTTGACCGCCTGGGGAGTACGGCCGCAAGGTTAAAACTCAAATGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGCCTTGACATCCAATGAACTTTCCAGAGATGGATTGGTGCCTTCGGGAACATTGAGACAGGTGCTGCATGGCTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTC CCGTAACGAGCGCAACCCTTGTCCTTAGTTACCAGCACGTCATGGTGGGCACTCTAAGGAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAGTCATCATGGCCCTTACGGCCTGGGCTACACACGTGCTACAATGGTCGGTACAGAGGGTTGCCAAGCCGCGAGGTGGAGCTAATCCCACAAAACCGATCGTAGTCCGGATCGCAGTCTGCAACTCGGTGAAGTCGGAATCGCTAGTAATCGCGA ATCAGAATGTCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGGGAGTGGGTTGCACCAGAAGTAGCTAGTCTAACCTTCGG.

The Pseudomonas campestris provided by the invention has excellent bactericidal performance and strong biofilm formation ability, can quickly colonize the soil rhizosphere, and effectively prevents and controls plant replanting diseases, plant bacterial diseases, plant fungal diseases, and plant soil-borne diseases. and/or plant oomycete diseases, especially to provide ideal biocontrol effects during continuous cropping of soils with disease-prone continuous cropping obstacles.

The Pseudomonas campestris provided by the invention has good compatibility with chemicals, can quickly colonize in soil environments where chemicals have been applied for a long time, limit the development of drug resistance of pathogenic bacteria, and bring efficient and stable biocontrol effects. At the same time, it can fully/partially replace the application of chemical pesticides, reduce the self-toxic effects of chemicals and residues on the soil and the destructive effects on soil beneficial bacteria, increase the abundance of beneficial bacteria in the soil, repair the soil environment, thereby solving/mitigating Soil autotoxicity and continuous cropping disorders caused by chemicals.

The Pseudomonas campestris provided by the invention has the effect of promoting seed germination and plant growth at the same time, and can be used to prepare biochemical inoculants, seed coating inoculants, root irrigation agents, seed soaking agents, plant growth regulators or soil conditioners.

The proposal of the present invention can not only solve the problem of continuous cropping obstacles caused by pathogenic bacteria and chemicals, and the shortcomings of pathogenic bacteria resistance caused by chemical pesticides, but also achieve the goal of being compatible with chemical pesticides and long-term effective in inhibiting various pathogenic bacteria and improving the physical and chemical properties of soil. , promote crop growth and seed germination, and its market value is huge.

In a preferred embodiment of the present invention, the above-mentioned plant soil-borne diseases include, but are not limited to, Ralstonia solanacearum, Rhizoctonia solani, Fusarium, Sclerotinia sclerotiorum, and Botrytis cirerea. , Cucumber Fusarium wilt fungus (Fusariumoxysporum.sp.cucumebrium Owen), Wheat total rot fungus (Gaeumannomycescritici), Wheat scab fungus (Fusariumgraminearum), apple tree rot fungus (Valsamali), apple anthracnose (Glomerellacingulata), rice sheath blight ( Rhizoctonia solan), Pyriculariagrisea, Alternariasolani, Exserohilumturcicum, Bipolariamaydis, Phytophthoracapsici and/or Phytophthoranicotianae ) caused by soil-borne plant diseases.

In an alternative embodiment, the Fusarium species is Fusarium graminearum.

In an optional implementation, the application temperature is 2-40°C; in an optional implementation, the application temperature is 31-35°C.

In a preferred embodiment of the present invention, the above-mentioned plant bacterial diseases include but are not limited to Ralstonia solanacearum, Bacillus subtilis (Bacillus subtilis), kiwi canker (Pseudomonas syringae), rice bacterial blight (Xanthomonas campestris), Bacterial diseases caused by Erwiniacarotorora, Xanthomonas campestris, Erwiniacarotovora and/or Staphylococcus aureus.

In an optional implementation, the application temperature is 2-40°C; in an optional implementation, the application temperature is 31-35°C.

In a preferred embodiment of the present invention, the above-mentioned plant fungal diseases or plant oomycete diseases include but are not limited to Rhizoctonia solani, Fusarium, Sclerotinia sclerotiorum, Botrytis cirerea, cucumber wilt Fusariumoxysporum.sp.cucumebrium Owen, Gaeumannomycescritici, Fusariumgraminearum, Valsamali, Glomerellacingulata, Rhizoctoniasolan, Pyriculariagrisea, Alternariasolani, Botrytis cirerea, Phytophthorainfestans, Exserohilumturcicum, Bipolariamaydis, Watermelon wilt Plant diseases caused by Fusarium oxysporum f. sp. niveum, Verticillium dahliae, Fusarium oxysporum f. sp. vasinfectum, Phytophthora capsici and/or Phytophthoranicotianae.

In an optional embodiment, the application temperature is 2-40°C; in another optional embodiment, the application temperature is 31-35°C, such as 31°C.

The present invention also provides the application of a bacterial strain of Pseudomonas campestris species in alleviating continuous cropping obstacles caused by diseases and/or soil autotoxicity, promoting plant growth, seed germination and/or soil improvement, Pseudomonas campestris The bacterial strain of this species was deposited in the Guangdong Provincial Microbial Culture Collection Center on January 15, 2021, with the preservation number: GDMCC No. 61435.

In a preferred embodiment of the present invention, the above-mentioned plants are grass crops or cash crops.

In an optional embodiment, the grass crops are corn, wheat, rice, sorghum, barley, oats, rye, millet, millet, barnyard grass, and buckwheat.

The economic crops are selected from at least one of the following economic crops: Solanaceae, Rosaceae, Rutaceae, Musa, Cucurbitaceae, Papilionaceae, Asteraceae, Liliaceae, Zingiberaceae, Passifloraceae, Bromeliaceae, Araliaceae and Cactaceae.

In an optional embodiment, Solanaceae is selected from at least one of the following Solanaceae crops: potato, pepper, and tomato; Rosaceae is selected from at least one of the following Rosaceae crops: strawberry and papaya; Rutaceae is selected from The following Rutaceae crops are: citrus; the Musa family crops are selected from the following Musa family crops: bananas; the Cucurbitaceae family is selected from cucumbers; the Papilionaceae family is selected from soybeans, the Asteraceae family is selected from lettuce, the Liliaceae family is selected from garlic, and the Zingiberaceae family is selected from Ginger, Passiflora family is selected from passion fruit, Bromeliaceae is selected from golden pineapple, Araliaceae is selected from Panax notoginseng, and Cactus family is selected from dragon fruit.

It should be noted that the above-mentioned crop types are only a few optional types listed by the inventor. In other embodiments, adaptive adjustments can also be made as needed and are not limited to the above-mentioned crop types.

The invention also provides Pseudomonas campestris. Pseudomonas campestris was deposited in the Guangdong Provincial Microbial Culture Collection Center on January 15, 2021, and the deposit number is: GDMCC No. 61435.

The above-mentioned Pseudomonas campestris includes, but is not limited to, cells of Pseudomonas campestris deposited with the deposit number GDMCC No. 61435 or its progeny strains or subclone strains.

The present invention also provides a biochemical bacterial agent, which includes the above-mentioned Pseudomonas campestris.

In an optional embodiment, the above-mentioned biochemical agents also include chemical agents.

In a preferred embodiment of the present invention, the above-mentioned chemical agent is a bacterial control chemical agent and/or a fungal control chemical agent.

In an optional embodiment, bacterial control chemical agents include, but are not limited to, cocidol, zhongshengmycin, copper thiacetin, thiocundazole, ethosalin, copper hydroxide, kasugamycin, and mycotoxin. , copper chloride, chlorobromoisocyanuric acid, trichloroisocyanuric acid, copper acetate or copper succinate; in an optional embodiment, the bacterial control chemical agent is selected from the group consisting of killable, killable and rapeseed The mixing volume ratio of Pseudomonas is 1000-10000:1; in an optional embodiment, the bacterial control chemicals are selected from Zhongshengmycin, and the mixture of Zhongshengmycin and Pseudomonas campestris The volume ratio is 1000-10000:1.

In an optional embodiment, the fungal control chemicals include, but are not limited to, carbendazim, mancozeb, ethyl aluminum, metalaxyl, redoxin, cymoxanil, dimethomorph, flumorphon pholine, argentil, benzamiconazole, myclobutanil, tebuconazole or propiconazole; in an optional embodiment, the fungal control chemical agent is selected from carbendazim, carbendazim and rapeseed The mixing volume ratio of Pseudomonas is 1000-10000:1.

The inventor found that M29 has good compatibility with chemicals, can quickly colonize in soil environments where chemicals have been applied for a long time, can limit the development of drug resistance of pathogenic bacteria, and has an efficient and stable biocontrol effect; at the same time, M29 and the fungicides can be used in whole or in part Substitute chemical chemicals are applied to soil plots with long-term continuous cropping to reduce the destructive effects of replanting diseases and chemicals on beneficial soil bacteria, increase the abundance of beneficial bacteria in the soil, and repair the soil environment, so as to reduce replanting diseases and chemical chemicals. Continuous cropping disorders caused by toxic effects.

The invention also provides a seed coating agent, a root irrigation agent, a seed soaking agent, a plant growth regulator and/or a soil conditioner. The seed coating agent includes a seed coating agent and Pseudomonas campestris, and the root irrigation agent, seed soaking agent The agent, plant growth regulator and/or soil conditioner includes Pseudomonas campestris, and the seed soaking agent includes Pseudomonas campestris. The seed coating agent can be selected from any commercially available seed coating agent.

In an optional embodiment, the seed soaking agent or root irrigation agent refers to a bacterial suspension of Pseudomonas campestris or a freeze-dried powder of Pseudomonas campestris.

In an optional embodiment, the bacterial suspension is a bacterial suspension of 10 ⁶ -10 ⁸ CFU/mL.

The invention has the following beneficial effects:

The Pseudomonas campestris provided by the invention has excellent bactericidal performance and strong biofilm formation ability, can quickly colonize the soil rhizosphere, and effectively prevents and controls plant replanting diseases, plant bacterial diseases, plant fungal diseases, and plant soil-borne diseases. and/or plant oomycete diseases, especially to provide ideal biocontrol effects during continuous cropping of soils with disease-prone continuous cropping obstacles.

In addition, the Pseudomonas campestris provided by the present invention has good chemical compatibility, can quickly colonize in soil environments where chemicals have been applied for a long time, limit the development of drug resistance of pathogenic bacteria, and can produce efficient and stable biocontrol effects. At the same time, it can replace the application of chemicals, reduce the autotoxic effect of chemicals on soil, increase the abundance of beneficial soil bacteria, improve soil physical and chemical properties, and solve the obstacles to continuous cropping of soil autotoxicity.

At the same time, the Pseudomonas campestris provided by the present invention has the effect of promoting seed germination and plant growth, and can be used to prepare seed coating agents, root irrigation agents, plant growth regulators or seed soaking agents.

The proposal of the present invention can not only solve the problem of continuous cropping obstacles caused by replanting diseases and chemicals and the shortcomings of pathogenic bacteria resistance caused by chemical pesticides, but also achieve the goal of being compatible with chemical pesticides and long-term effective in inhibiting various pathogenic bacteria and improving soil. Its physical and chemical properties, effects on promoting crop growth and seed germination, have huge market value.

Description of the drawings

In order to explain the technical solutions of the embodiments of the present invention more clearly, the drawings required to be used in the embodiments will be briefly introduced below. It should be understood that the following drawings only show some embodiments of the present invention and therefore do not It should be regarded as a limitation of the scope. For those of ordinary skill in the art, other relevant drawings can be obtained based on these drawings without exerting creative efforts.

Figure 1 shows the colony morphology of M29 strain on LB medium;

Figure 2 is a picture of the biological control effect of leaf cutting on bacterial wilt by M29 inoculant;

Figure 3 is a diagram showing the effectiveness of root irrigation and biological control of bacterial wilt by Pseudomonas campestris M29 strain;

Figure 4 shows the growth-promoting effect of Pseudomonas campestris M29 strain;

Figure 5 shows the biocontrol effect of Pseudomonas campestris M29 strain on Fusarium graminearum.

Detailed ways

In order to make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be described clearly and completely below. If the specific conditions are not specified in the examples, the conditions should be carried out according to the conventional conditions or the conditions recommended by the manufacturer. If the manufacturer of the reagents or instruments used is not indicated, they are all conventional products that can be purchased commercially.

The features and performance of the present invention will be described in further detail below with reference to examples.

Example 1

This example provides methods for screening and identifying Pseudomonas campestris M29 strains.

The screening method for M29 strains is as follows:

Pseudomonas brassicacearum M29 was isolated from agricultural soil collected from the sweet potato planting area in Wucun, Beizhang Township, Wenshui County, Shanxi Province (37°22′28.24″N, 112°05′34.32″E).

Weigh 5g of soil, add 45mL of sterile water, shake with a vortex shaker for 5 minutes, and then dilute it to a gradient of 10 ^-1 to 10 ^-6 . Take three gradients of bacterial liquid: 10 ^-4 , 10 ^-5 and 10 ^-6 . The plate is coated on Luria-Bertani (LB) solid medium (the formula is: 5g yeast extract powder, 10g tryptone, 10g sodium chloride, 15g agar, 1L water, pH=7), each coated Three replicates were made on the cloth, and the culture was inverted for 4 days in a constant temperature incubator at 30°C. After 4 days, single colonies of strains were picked out, streaked and inoculated on a new LB plate, and cultured at 30°C for 1 day to obtain a pure culture of the strain. Glycerol tubes were stored in a -80°C refrigerator.

Identification of M29 strains:

The isolated strain was inoculated into Luria-Bertani medium and cultured at 30°C for 1 day. The morphology is as shown in Figure 1. The strain is round, with smooth and shiny edges, dirty white color, and a diameter of 0.1cm.

Subsequently, the 16S sequence fragment of the strain was determined (the amplification primer and the sequencing primer were both 27F: 5'-AGAGTTTGATCCTGGCTCAG-3' and 1492R: 5'-GGTTACCTTGTTACGACTT-3'). The determination results are shown in the sequence table SEQ ID NO. As shown in .1, through BLAST homology comparison, it was determined that the nearest species of this strain is Pseudomonas brassicacearum (Pseudomonas brassicacearum), named Pseudomonas brassicacearum M29. Send Pseudomonas campestris M29 to the Guangdong Microbial Culture Collection Center (GDMCC) for preservation. The preservation address is: 5th Floor, Building 59, Courtyard, No. 100 Xianlie Middle Road, Guangzhou. The preservation date is: January 15, 2021. Number: GDMCC No.61435.

Example 2

In this example, a fermentation culture of Pseudomonas campestris M29 strain was prepared.

After the frozen M29 strain in Example 1 was activated, liquid fermentation was performed to prepare a fermentation culture.

a. Activation of Pseudomonas campestris M29 strain

Use the coating method to spread the M29 strain stored in -80°C refrigerator on the LB plate (the formula is: 10g peptone, 5g yeast powder, 5g sodium chloride, 1g glucose, 15g agar, add water to 1L, sterilize at 121°C for 20 minutes) , cultured at 30°C for 1 day. Pick a single colony of M29 grown on the LB plate and inoculate it into LB liquid culture medium (the formula is: 10g peptone, 5g yeast powder, 5g sodium chloride, 1g glucose, add water to 1L, sterilize at 121°C for 20 minutes), sterilize at 30°C After culturing for 1 day, the M29 strain was obtained.

b. Preparation of M29 strain fermentation culture

Inoculate 1mL of M29 strain into LB liquid culture medium, and culture it aseptically on a shaking table at 30°C for 1 day to obtain a fermentation culture. The number of viable bacteria was measured to be up to 0.8×10 ¹¹ cfu/mL, which is consistent with the spore production amount or Strains with a viable bacterial count ≥10 billion spores/g or cfu/mL are determined as indicators of easily fermentable strains.

Example 3

This example conducts an experiment on the biological control effect of Pseudomonas campestris M29 strain on R. solanacearum R.s.

Preparation of R.s bacteria suspension:

Activate Ralstonia solanacearum Rs (a pathogenic bacterium preserved in the own bacterial bank) from the glycerol tube onto the TTC plate (the formula is: 5g glucose, 10g peptone, 1g hydrolyzed complexin, 15g agar, add water to 1L, Sterilize at 121°C for 20 minutes, cool to 60°C and then add TTC to make the final concentration 0.005% (W/V)). After culturing for 2 days, select colonies with high activity and transfer them to SPA liquid culture medium (the formula is: sucrose 20g, bacteriological peptone 5g, K ₂ HPO ₄ 0.5g, MgSO ₄ 0.25g, add water to 1L, adjust to pH 7.0~7.2, sterilize at 121°C for 20min), culture on a shaking table for 8-16h and then sterilize according to OD ₆₀₀ Dilute Rs to 10 ⁷ CFU/mL according to the absorbance value, and prepare Rs bacterial suspension for later use.

Preparation of M29 inoculant:

Use a spectrophotometer to measure the absorbance value at OD600 of the M29 strain fermentation culture prepared in Example 2, and adjust its concentration to 10 ⁸ CFU/mL using LB liquid culture medium to obtain the M29 bacterial agent.

Tomato seedling treatment:

After the 11-day-old tomato seedlings were thinned to 10 plants/pot, the leaves were cut to preliminarily judge the biocontrol effect of M29 on R.s. According to the difference in the soaking liquid of the scissors used when cutting leaves, a blank control group, a negative control group, a positive control group and a bacterial agent treatment group were set up, and each group was repeated three times. Before cutting the leaves of each tomato seedling, soak the sterile scissors in the corresponding soaking solution of each group for 1 second, and then use the soaked scissors to cut off half of the cotyledons of the tomato seedlings. Then, the tomato seedling pots after the leaves were cut were placed in a disease shed with high temperature and humidity of 30°C for 7 days. From the 4th day after leaf cutting, the number of dead tomato seedlings was recorded every day, and the mortality rate and relative control effect were calculated.

The soaking solutions used in each group are as follows:

The blank control group was 40 mL of clean water;

The negative control group consisted of 20 mL each of clean water and R.s bacterial suspension, mixed well;

The positive control group consisted of 20 mL each of 1000-fold dilution of neophytomycin and R.s bacteria suspension, and mixed well;

The bacterial agent treatment group contains 20 mL each of M29 bacterial agent and R.s bacterial suspension, and mix well.

Relative control effect = (negative control mortality rate - treatment group mortality rate)/negative control mortality rate × 100%.

Table 1 Biocontrol effects of Pseudomonas campestris M29 agent on bacterial wilt on leaf cuttings

Figure 2 and Table 1 show that under the leaf-cutting system, the mortality rate of tomato seedlings in the Pseudomonas campestris M29 inoculant treatment group was only 10%, which was 67% lower than the mortality rate of tomato seedlings in the positive control group (30%). Compared with the negative control group, the mortality rate of tomato seedlings (62.22%) was reduced by 84%. In addition, the control effect of the M29 bacterial agent treatment group on bacterial wilt reached 83.92%, while the control effect of the positive control group on bacterial wilt was 51.79%. This shows that M29 inoculant can significantly reduce the mortality of tomato seedlings and has very good biocontrol potential against tomato bacterial wilt.

Example 4

This example conducts a verification experiment on the potted biocontrol effect of Pseudomonas campestris M29 strain on R. solanacearum R.s.

Prepare R.s bacteria suspension and M29 bacteria agent according to the method described in Example 3 for later use.

Tomato seedling treatment:

Gently shake off the soil on the root surface of 20-day-old tomato seedlings, soak the roots in the treatment solution for 10 minutes, and then plant them into sterilized soil. One plant per pot, 6 pots per treatment, 5 repetitions. Set up a blank control group, a negative control group, a positive control group and a bacteria treatment group according to different treatment solutions. The treatment solution for the blank control group and the negative control group was clean water, the treatment solution for the positive control group was 1000-fold dilution of neophytomycin, and the treatment solution for the bacteria treatment group was M29 bacteria. Place the potted plants in an environment of 25-28°C to slow down the seedlings for 7 days. Then insert a garden spade into the soil around the roots to damage the tomato roots and cause wounds. Then, each seedling in the negative control group, positive control group and bacteria treatment group was filled with 50 mL of Rs bacterial suspension of 10 ⁷ cfu/mL, and the blank control group was filled with an equal amount of water.

Observation record:

At the first sign of symptoms (withering symptoms on tomato seedlings) (usually the onset of symptoms occurs on the 5th day after root filling), observe and record the status of the tomato seedlings once a day (record the incidence according to the grading standards), until the 10th to 13th day (Determined based on the actual disease situation), record the number of plants at each disease level in each treatment, and calculate the disease index and control effect. The results are shown in Table 2 and Figure 3.

Plant disease levels are divided into the following five levels:

Level 0: Asymptomatic

Level 1: One leaf wilted

Level 2: 2-3 leaves wilted

Level 3: The whole plant wilts

Level 4: Plant death

Disease index (%) = (∑ (number of plants in each disease level × number of levels)/(number of total plants × number of highest levels)) × 100%.

Prevention and treatment effect (%) = ((negative control group’s condition index – experimental group’s condition index)/negative control group’s condition index) × 100%.

Table 2 Root irrigation biocontrol effects of Pseudomonas campestris M29 strain on bacterial wilt

As can be seen from Table 2 and Figure 3, because the tomato seedlings in the negative control group were only soaked in clean water without adding any pharmaceutical ingredients, the incidence rate of infection by R. solanacearum R.s was 41.7%. The tomato seedlings showed signs of plant failure. Severe wilting or even death of the entire plant. Compared with the negative control group, the tomato seedlings in the bacterial agent treatment group grew better, with almost no leaf wilting. The plant incidence rate (16.7%) was 60% lower than that in the negative control group, and the control effect on bacterial wilt was as high as 63.84%. %. In addition, the incidence rate of the bacterial treatment group was the same as that of the positive control group, but the disease index was slightly lower than that of the positive control. The control effect was equivalent to that of the positive control, and was much higher than the control effect of the same strain J12 from different sources (DOI: 10.1016 /j.micres.2012.01.003, section 3.4). It shows that M29 root soaking treated tomato seedlings can not only effectively prevent tomato bacterial wilt, but its control effect is even better than that of neophytomycin, a traditional chemical pesticide. It can be used to replace/partially replace traditional chemical pesticides and has broad market prospects.

In addition, this example also found that the Pseudomonas campestris M29 strain showed a significant promoting effect on the growth of tomato seedlings. As can be seen from Figure 4, the number of leaves grown by the tomato seedlings in the bacterial agent treatment group was significantly more than that in the blank control group, and by measuring the dry weight of the tomato seedlings, it was found that the average dry weight of the tomato seedlings in the bacterial agent treatment group was higher than that in the blank control group. 40%, Pseudomonas campestris M29 strain has significant growth-promoting effect.

Example 5

In this example, a verification experiment was conducted on the biofilm formation ability of Pseudomonas campestris M29 strain.

Prepare the M29 strain fermentation culture according to the method described in Example 2, use a spectrophotometer to measure its absorbance at OD ₆₀₀ , and use LB liquid culture medium to adjust the OD ₆₀₀ value of the fermentation culture to 1, and then draw 100 μL of the fermentation culture therefrom. cultured in a 96-well plate for 24 h. After washing twice with 200 μL PBS solution, the 96-well plate was placed in an oven at 60°C for 1 hour to fix the biofilm. Then, 50 μL 0.4% crystal violet solution was added to the wells for staining for 15 min, then washed three times with 200 μL PBS, and finally incubated at 37°C. Dry the liquid at low temperature, add 200 μL of 70% ethanol, and measure the absorbance at 600 nm with a microplate reader to evaluate the biofilm-forming ability of Pseudomonas campestris M29. During the entire experimental process, a negative control group and a bacterial agent treatment group were set up. In the negative control group, LB liquid culture medium was added to the 96-well plate, and in the bacterial agent treatment group, fermentation culture (OD ₆₀₀ = 1) was added. Three parallel groups were set up in each group. .

Table 3 Biofilm-forming ability of Pseudomonas campestris M29 strain

According to the measured absorbance, the average value (D) of three repetitions was taken, and 2 times the average value of the negative control group was used as the limit value (Dc). When the average value of D in the bacterial treatment group _is >2×Dc, the strain is judged to be a strong biofilm-forming strain; when Dc< _D =≤2×Dc, the strain is judged to be a weak biofilm-forming strain; when D _is ≤Dc, the strain is judged to be a weak biofilm-forming strain. The strain was non-biofilm forming.

It can be seen from Table 3 that D=0.48, D _pair =0.09, Dc=0.18, 2×Dc=0.36, _{D >} 2×Dc, so Pseudomonas campestris M29 strain is a strain with strong biofilm formation ability. strains. The strong ability to form biofilm indicates that Pseudomonas campestris M29 strain is easy to colonize in the plant soil environment, thus forming an excellent competitive advantage. This advantage can be used to quickly capture water, nutrients, and occupy space to weaken and eliminate the same The growth of other pathogenic bacteria in the growth environment thus produces a more rapid, stable and lasting biocontrol effect.

Example 6

In this example, a compatibility test between Pseudomonas campestris M29 and chemical drugs was conducted.

In view of the fact that there are varying degrees of chemical pesticide residues in the soil where most crops are currently grown, and the production of biocontrol agents is often used in conjunction with chemical agents, in order to ensure that the Pseudomonas campestris M29 strain is still stable and effective when put into field use. In order to improve the biocontrol effect, we specially organized a compatibility test between Pseudomonas campestris M29 strain and chemical drugs. The test methods and results are as follows.

Preparation of chemical-added LB plates

We selected two commonly used chemicals for bacterial diseases, Keshid and Zhongshengmycin, and two commonly used chemicals for fungal diseases, carbendazim and mancozeb, and selected the highest concentration dilution ratio and its 10-fold dilution of the recommended dosage. Concentration, add the four chemicals to the liquid LB agar medium at 50°C, shake well, and pour the plate. For example, the recommended dilution factor for the field that can be killed is 1000 times, then we choose two dilution concentrations of 1000 times and 10000 times. Zhongshengxin and mancozeb have strong antibacterial properties, so only the 10-fold dilution treatment of their highest concentration dilution is retained.

dilution plate

After preparing the fermentation culture with OD ₆₀₀ =1 in the manner described in Example 5, absorb 1 mL of the fermentation culture into a 15 mL centrifuge tube, add 9 mL of sterile Tween water to dilute to 10 ^-1 , and then use a gradient dilution method to dilute to 10 ^{- 7.} Select the fermentation culture diluted to 10 ^-5 -10 ^-7 and spread it on a plate with different chemicals. Use the LB plate without chemicals as a control. After culturing in a 30°C incubator overnight, use plate counting. method to calculate the antibacterial rate of each treatment group.

Antibacterial rate = ((number of bacteria on LB plate - number of bacteria on plate with chemicals)/number of bacteria on LB plate) × 100% Table 4 Antibacterial spectrum of chemical drugs for Pseudomonas campestris M29 strain

Note: "KexiDe" represents the highest recommended field concentration dilution ratio of "KeKeDe"; "KeKeDe -10" represents the 10-fold dilution of "KeKeDe".

As can be seen from Table 4, in addition to being very sensitive to mancozeb, M29 has very good tolerance to cocid and carbendazim. The strain is not inhibited by them. The highest concentration dilution of Zhongshengmycin is recommended in the field. The 10-fold dilution is also fully tolerated. M29 has very good tolerance to the overall chemical, which provides a theoretical basis for subsequent adaptation to complex field conditions.

Example 7

This example conducts a verification experiment on the biocontrol effect of Pseudomonas campestris M29 strain on bacterial wilt in tomato continuous cropping field plots.

A multi-year continuous cropping plot with long-term application of mesozoicin to control tomato bacterial wilt disease was used as the experimental site (experimental location: Xingteng Experimental Base, Shiqi Town, Panyu District, Guangzhou City, Guangdong Province). The previous crop of the experimental plot was tomatoes, and the plot was continuously cropped for 4 years. , bacterial wilt disease is serious, commonly used chemicals include mesomycin, thiazole zinc, etc.), in order to verify the inhibitory or alleviating effect of M29 strain on continuous cropping obstacles caused by replanting diseases and soil autotoxicity.

Test method: Plan the test area according to the area of a single test area of 18m2. A total of nine test areas are set up and divided into bacterial agent treatment groups, positive controls and blank control groups. Each group repeats three test areas. An equal number of uniformly grown tomato seedlings were transplanted into each test area, and the tomato seedlings were irrigated with fixed root water. Seven days after transplanting, the tomato seedlings in each test area were rhizosphere flushed according to the treatment method shown in Example 4. Treatment, and apply compound fertilizer once 30 days after transplanting and once in the early flowering stage. Observe and record the disease incidence of tomato plants in each plot 30 days after transplantation according to the method of Example 4, and calculate the disease index and control effect.

Table 5 Control effect of Pseudomonas campestris M29 strain on soil bacterial wilt disease in continuous cropping fields

The results in Table 5 further verify the chemical compatibility of the M29 strain. Its rapid colonization in soil environments where chemicals are heavily used effectively solves the continuous cropping obstacles caused by replanting diseases and can alleviate the damage caused by chemicals to a certain extent. Soil autotoxicity is an obstacle to continuous cropping, which is conducive to continuous cropping of crops and ensures crop yields.

Example 8

This example conducts a verification experiment on the control effect of Pseudomonas campestris M29 strain on cucumber blight.

Pathogen preparation:

Use a hole punch to make holes on the Rhizoctonia solani culture medium under sterile conditions to obtain a 5mm diameter mushroom cake, and connect it to PDA (the formula is: 200g of potatoes, boiled in water, Filter, add 20g of glucose, 15g of agar, add water to 1L, sterilize at 121°C for 20min) on the culture medium plate, place one bacterial cake per dish, and culture in a constant temperature incubator at 28°C for 7 days for later use.

Watering treatment:

After 7 days of cucumber seedling cultivation, water-soluble fertilizer is poured into the seedling tray. After 12 days of sowing, pull it out and use scissors to cut off all the fibrous roots and part of the main root. Select cucumber seedlings with consistent growth and plant them in small flower pots with sterilized soil. , 3 plants per pot. Use a spectrophotometer to measure the absorbance at OD ₆₀₀ of the M29 strain fermentation culture obtained in Example 2, and use LB liquid culture medium to adjust the OD ₆₀₀ value of the fermentation culture to 0.12, then use it to water cucumber seedlings, each pot A total of 6 × 10 ⁸ cfu was poured into the pots with 50 mL. Every 6 pots were treated as one treatment, and each treatment was repeated 3 times. The control group was irrigated with the same amount of water. Two hours after inoculation, two small holes were poked at opposite positions in each pot, and a pathogen cake with a diameter of 5 mm was connected to each hole, and then placed in a greenhouse at 28-30°C for 7 days to develop the disease.

Clean the soil at the roots of cucumber seedlings that have experienced high temperature and high humidity, and record the incidence according to the disease classification standards. Finally, put the cleaned cucumber seedlings into a 75°C oven to dry, weigh the dry weight, and calculate the dry weight growth rate. .

Disease grade grading standards:

Level 0: There are no lesions at the base of the cucumber stems.

Level 1: The lesions at the base of the cucumber stem occupy less than 1/4 of the stem circumference.

Level 2: The lesions at the base of the cucumber stem occupy between 1/4 and 1/2 of the stem circumference.

Level 3: The lesions at the base of the cucumber seedling occupy more than 1/2 of the stem circumference, but do not destroy the entire stem circumference.

Level 4: Symptoms such as depression or breakage at the junction of the roots and stems of cucumber seedlings, rot and browning, etc.

Incidence index = (∑ (number of diseased plants at each level × representative value of each disease level)/(total number of plants investigated × highest level of disease incidence value)) × 100%

Relative control effect (%) = (disease grade index of the control group - disease grade index of the treatment group)/disease grade index of the control group × 100%.

Table 6 The control effect of Pseudomonas campestris M29 strain on cucumber blight

After irrigating with Pseudomonas campestris M29 agent, the blight incidence index of cucumber seedlings was significantly lower than that of the control group. The control effect against cucumber blight reached 30.27%, and the dry weight increased by 9.89%. It shows that Pseudomonas campestris M29 agent can prevent and control cucumber blight and has a growth-promoting effect.

Example 9

In this example, a verification experiment was conducted on the biocontrol effect of Pseudomonas campestris M29 strain on Fusarium graminearum on wheat.

F.g bacterial suspension preparation:

Fusarium graminearum Fg (Fusarium graminearum) was activated on the PDA plate. After culturing for 5 days at 28°C, CMC culture medium (the formula was: CMC-Na 15g, NH ₄ NO ₃ 1g, yeast extract 1g, MgSO ₄ ·7H ₂ O 0.5g, KH ₂ PO ₄ 1g, add water to 1L, sterilize at 121℃ for 20min), incubate for 7 days at 25-28℃, 200rpm, filter with gauze to obtain the spore suspension, count with a hemocytometer, and then prepare to a concentration The spore suspension is 10 ⁶ CFU/mL.

Preparation of seed coating agent

Formula: sucrose 5%, PVPK30 10%, PEG 3%, dispersant MF 2%, polyethylene glycol 5%. Each component is proportioned according to mass fraction.

Take a 50mL centrifuge tube, first add PVPK30 and a small amount of water according to the above proportion, shake to dissolve, then add other reagents in proportion, dissolve completely, and adjust to volume to obtain the required seed coating agent.

Preparation of inoculation agent

The absorbance at OD ₆₀₀ of the M29 strain fermentation culture obtained in Example 2 was measured with a spectrophotometer, and after the OD ₆₀₀ value of the fermentation culture was adjusted to 1 using LB liquid culture medium, 40 μL of the fermentation culture with OD ₆₀₀ = 1 was taken. The culture is mixed with 360 μL of seed coating agent to obtain M29 chlamydia agent. An equal amount of LB liquid medium was used to replace the fermentation culture and an equal amount of seed coating agent was mixed to prepare a control seed coating agent.

seed treatment

Set up a blank control group, negative control group and bacteria treatment group. In the inoculant treatment group, the seeds were coated with inoculant (400 μL of inoculant/20g wheat) to allow M29 to adhere to the surface of the seeds. The seeds in the blank control group were not treated in any way, and the seeds in the negative control group were coated with a seed coating agent without the addition of M29 fermentation culture.

soil preparation

Sterile soil was added to each seedling pot. 20 mL of F.g bacterial suspension was mixed into the sterile soil of the negative control group and the bacterial treatment group in advance, and the same amount of water was mixed into the blank control group. Then plant the coated seeds into seedling pots.

The germination rate of wheat was counted after germination, and the condition of wheat was counted after 14 days.

The specific classification of the condition is as follows:

Level 0, no disease;

Level 1, mild symptoms appear on the first leaf sheath (brown stripes or blackening appear on the first leaf sheath, but the degree of blackening does not exceed 50%);

Level 2, the first leaf sheath is seriously affected (the degree of blackening of the first leaf sheath exceeds 50%);

Level 3, mild symptoms appear on the second leaf sheath (brown stripes or blackening appear on the second leaf sheath, but the degree of blackening does not exceed 50%);

Level 4, the second leaf sheath is seriously affected (the degree of blackening of the second leaf sheath exceeds 50%);

Level 5, the third leaf sheath is mildly affected (the third leaf sheath appears brown stripes to blackened, but the degree of blackening does not exceed 50%);

Level 6: The disease on the third leaf sheath is so severe that the plant is close to death.

Table 7 Biocontrol effects of Pseudomonas campestris M29 strain on Fusarium graminearum

As can be seen from Table 7 and Figure 5, the germination rate of the wheat seedlings in the negative control group was significantly reduced (28.7%) after being infected with Fusarium graminearum, and the late growth of the germinated seeds was also affected by the toxicity of Fusarium graminearum. The harvested wheat Plant death was severe. In the bacterial agent treatment group, due to the M29 bacterial agent coating, the wheat seeds were protected from pathogenic interference, and the germination rate was maintained at about 60%, which was 106% higher than the negative control group. The effect of M29 in promoting seed germination was extremely significant. On the other hand, M29 reduced the disease index of wheat plants to a certain extent, and its control effect was 17.06%.

In summary, the Pseudomonas campestris M29 strain of the present invention has ideal bactericidal properties and can not only prevent and control pathogenic bacteria such as tomato bacterial wilt, cucumber damping off and wheat fusarium, but also carry out continuous cropping in soils with disease continuous cropping obstacles. It provides ideal biocontrol effects at the same time; it also has good chemical compatibility and rhizosphere colonization ability, which can limit the development of drug resistance of pathogenic bacteria, improve soil physical and chemical properties, and reduce continuous cropping obstacles caused by chemical toxicity; and it can also Significantly promotes seed germination and plant growth, and has broad application prospects.

The above descriptions are only preferred embodiments of the present invention and are not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and changes. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention shall be included in the protection scope of the present invention.

SEQUENCE LISTING

<110> Munn (Guangzhou) Biotechnology Co., Ltd.

<120> A kind of Pseudomonas campestris, biochemical agent and its application

<160> 1

<170> PatentIn version 3.5

<210> 1

<211> 1390

<212> DNA

<213> Artificial sequence

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Claims (31)

1. Pseudomonas brassicaePseudomonas brassicacearum) The use of bacterial strains of the species for controlling plant bacterial diseases, characterized in that,

the bacterial strain of the Pseudomonas brassicae is preserved in the microorganism strain collection of Guangdong province in 2021, 1 and 15 days, and the preservation number is: GDMCC No.61435.

2. Pseudomonas brassicaePseudomonas brassicacearum) The use of bacterial strains of the species for controlling fungal plant diseases, characterized in that,

the bacterial strain of the Pseudomonas brassicae is preserved in the microorganism strain collection of Guangdong province in 2021, 1 and 15 days, and the preservation number is: GDMCC No.61435.

3. Pseudomonas brassicaePseudomonas brassicacearum) The application of bacterial strains of species in controlling plant oomycete diseases is characterized in that,

The bacterial strain of the Pseudomonas brassicae is preserved in the microorganism strain collection of Guangdong province in 2021, 1 and 15 days, and the preservation number is: GDMCC No.61435.

4. Pseudomonas brassicaePseudomonas brassicacearum) The use of bacterial strains of the species for controlling bacterial re-plant diseases of plants, characterized in that,

the bacterial strain of the Pseudomonas brassicae is preserved in the microorganism strain collection of Guangdong province in 2021, 1 and 15 days, and the preservation number is: GDMCC No.61435.

5. Pseudomonas brassicaePseudomonas brassicacearum) The use of bacterial strains of the species for controlling fungal reimplantation diseases of plants, characterized in that,

the bacterial strain of the Pseudomonas brassicae is preserved in the microorganism strain collection of Guangdong province in 2021, 1 and 15 days, and the preservation number is: GDMCC No.61435.

6. Pseudomonas brassicaePseudomonas brassicacearum) The use of bacterial strains of the species for controlling bacterial soil-borne diseases of plants, characterized in that,

the bacterial strain of the Pseudomonas brassicae is preserved in the microorganism strain collection of Guangdong province in 2021, 1 and 15 days, and the preservation number is: GDMCC No.61435.

7. Rape falseMonomonas @Pseudomonas brassicacearum) The application of bacterial strains of species in preventing and controlling fungal soil-borne diseases of plants is characterized in that,

The bacterial strain of the Pseudomonas brassicae is preserved in the microorganism strain collection of Guangdong province in 2021, 1 and 15 days, and the preservation number is: GDMCC No.61435.

8. Pseudomonas brassicaePseudomonas brassicacearum) The use of a bacterial strain of a species for controlling at least two diseases, characterized in that,

the disease is selected from the group consisting of plant bacterial disease, plant fungal disease and plant oomycete disease;

the bacterial strain of the Pseudomonas brassicae is preserved in the microorganism strain collection of Guangdong province in 2021, 1 and 15 days, and the preservation number is: GDMCC No.61435.

9. The use according to claim 1, 4 or 8, wherein the plant bacterial reignition disease or plant bacterial disease comprises the species lactobacillus solanacearum @Ralstonia solanacearum) Bacillus subtilisBacillus subtilis) Kiwi fruit canker pathogenPseudomonas syringae) Bacterial leaf blight of riceXanthomonas campestris pv.oryzae) Chinese cabbage soft rot fungusErwinia carotorora) Black spot germ of walnutXanthomonas campestris) Konjak soft rot fungusErwinia carotovora) And/or staphylococcus aureus @ sStaphylococcus aureus) Bacterial disease caused.

10. The use according to claim 9, wherein the application temperature is 2-40 ℃.

11. The use according to claim 10, wherein the application temperature is 31-35 ℃.

12. Use according to any one of claims 2-3, use according to claim 5 orThe use according to claim 8, wherein said plant fungal disease or plant oomycete disease comprises a disease selected from the group consisting of rhizoctonia solani @Rhizoctonia solani) Fusarium (Fusarium)Fusarium) Sclerotinia sclerotiorum (L.) KuntzeSclerotinia sclerotiorum) Tomato gray mold fungusBotrytis cirerea) Cucumber fusarium wiltFusarium oxysporum.sp.cucumebrium Owen) Wheat take-all germGaeumannomyces critici) Wheat gibberella germFusarium graminearum) Apple tree rot pathogenValsamali) Apple anthracnose pathogenGlomerella cingulata) Rhizoctonia solani of riceRhizoctonia solan) Pyricularia oryzaePyricularia grisea) Tomato early blight germAlternaria solani) Botrytis cinerea of strawberryBotrytis cirerea) Potato late blight germPhytophthora infestans) The corn big spot germ isExserohilum turcicum) Small spot germ of cornBipolaria maydis) Watermelon fusarium wiltFusarium oxysporum f.sp.niveum) Verticillium wilt bacteria of eggplantVerticillium dahliae) Cotton fusarium wilt bacteriaFusarium oxysporum f.sp.vasinfectum) Phytophthora capsiciPhytophthora capsici) And/or phytophthora nicotianae bacteriaPhytophthora nicotianae) Caused plant diseases.

13. The use according to claim 12, wherein the application temperature is 2-40 ℃.

14. The use according to claim 13, wherein the application temperature is 31-35 ℃.

15. The use according to claim 6 or 7, wherein the plant bacterial soil-borne disease or plant fungal soil-borne disease comprises the species lactobacillus solanacearum @ Ralstonia solanacearum) Rhizoctonia solani (wall.) kuntzeRhizoctonia solani) Fusarium (Fusarium)Fusarium) Sclerotinia sclerotiorum (L.) KuntzeSclerotinia sclerotiorum) Tomato gray mold fungusBotrytis cirerea) Cucumber fusarium wiltFusarium oxysporum.sp.cucumebrium Owen) Wheat take-all germGaeumannomyces critici) Wheat gibberella germFusarium graminearum) Apple tree rot pathogenValsamali) Apple anthracnose pathogenGlomerella cingulata) Rhizoctonia solani of riceRhizoctonia solan) Pyricularia oryzaePyricularia grisea) Tomato early blight germAlternaria solani) The corn big spot germ isExserohilum turcicum) Small spot germ of cornBipolaria maydis) Phytophthora capsiciPhytophthora capsici) And/or phytophthora nicotianae bacteriaPhytophthora nicotianae) And (3) the caused plant soil-borne diseases.

16. The use according to claim 15, wherein the fusarium is fusarium graminearum.

17. The use according to claim 15, wherein the application temperature is 2-40 ℃.

18. The use according to claim 17, wherein the application temperature is 31-35 ℃.

19. Use of a bacterial strain of the species pseudomonas brassicae for alleviating continuous cropping obstacles caused by disease and/or soil autotoxicity, promoting plant growth, promoting seed germination and/or soil improvement, characterized in that said bacterial strain of the species pseudomonas brassicae is deposited at the microorganism strain deposit center in the cantonese province at 1 month 15 of 2021, deposit number: GDMCC No.61435.

20. The use according to claim 19, wherein the plant is a gramineous crop or a commercial crop.

21. The use according to claim 20, wherein the gramineous crop is maize, wheat, rice, sorghum, barley, oat, rye, millet, barnyard grass, buckwheat;

the cash crop is selected from at least one of the following cash crops: solanaceae, rosaceae, rutaceae, musaceae, cucurbitaceae, papilionaceae, compositae, liliaceae, zingiberaceae, passifloraceae, pineapple, araliaceae, and Cactaceae.

22. Use according to claim 21, wherein said solanaceae is selected from the group consisting of at least one solanaceous crop of the family: potatoes, peppers, and tomatoes; the rosaceae is selected from at least one rosaceae crop of the following: strawberry and papaya; the Rutaceae is selected from Rutaceae crops as follows: citrus fruit; the musaceae is selected from the following musaceae crops: bananas; the cucurbitaceae is selected from cucumber; the Papilionaceae is selected from soybean, the Compositae is selected from lettuce, the Liliaceae is selected from garlic, the Zingiberaceae is selected from ginger, the Passifloraceae is selected from passion fruit, the pineapple family is selected from golden pineapple, the Araliaceae is selected from pseudo-ginseng, and the Cactaceae is selected from dragon fruit.

23. A pseudomonas rape, characterized in that the pseudomonas rape is deposited with the cantonese province microorganism strain collection at 1-15 of 2021 under the deposit number: GDMCC No.61435.

24. A biochemical microbial agent comprising pseudomonas brassicae of claim 23; the biochemical microbial inoculum also comprises a chemical agent, wherein the chemical agent is at least one of zinc thiazole, zhongshengmycin, spinosad and carbendazim.

25. The biochemical microbial agent according to claim 24, wherein the mixing volume ratio of said shakable and said pseudomonas brassicae is 1000-10000:1.

26. The biochemical microbial agent according to claim 24, wherein the mixing volume ratio of the mesogenic agent to the pseudomonas brassicae is 1000-10000:1.

27. The biochemical microbial agent according to claim 24, wherein the mixing volume ratio of carbendazim to pseudomonas brassicae is 1000-10000:1.

28. The use of the biochemical fungicide according to claim 24 for controlling plant diseases, wherein said diseases are selected from at least one of plant bacterial diseases, plant fungal diseases and plant oomycete diseases.

29. A seed coating agent, root irrigation agent, seed soaking agent, plant growth regulator and/or soil conditioner, characterized in that the seed coating agent comprises a seed coating agent and the pseudomonas brassicae of claim 23, and the root irrigation agent, seed soaking agent, plant growth regulator and/or soil conditioner comprises the pseudomonas brassicae of claim 23.

30. A seed coating agent, root canal filling agent, seed soaking agent, plant growth regulator and/or soil amendment according to claim 29, wherein the seed soaking agent or root canal filling agent is a bacterial suspension of pseudomonas brassicae or a lyophilized powder of pseudomonas brassicae.

31. A seed coating agent, root canal filling agent, seed soaking agent, plant growth regulator and/or soil conditioner according to claim 30, wherein the bacterial suspension is 10 ⁶ -10 ⁸ CFU/mL of bacterial suspension.