KR100781472B1 - Microorganism having the characteristic of antagonism to phytopathogenic fungi, desease curing microorganism composition comprising said microorganism and production method thereof - Google Patents

Abstract

A novel strain is provided which shows the antagonistic characteristic on phytopathogenic fungi without contaminating soil and water. And a microorganism composition comprising the same for controlling Rhizoctonia solani and Sclerotinia homoeocarpa is provided. A novel microorganism having the antagonistic function on phytopathogenic fungi is at least one selected from the group consisting of Bacillus-amyloliquefaciens KS-4(deposition no. KACC 91247P), Bacillus-agaradhaerens KS-5(deposition no. KACC 91248P), Paenibacillus-lentimorbus KS-6(deposition no. KACC 91249P), Bacillus laevolacticus KS-7(deposition no. KACC 91250P), Leuconostoc paramesenteroides KS-9(deposition no. KACC 91251P), Kurthia-sibirica KS-13(deposition no. KACC 91252P) and Sphingobacterium-spiritivorum KS-18(deposition no. KACC 91253P). A microorganism composition for treating and preventing large patch caused by Rhizoctonia solani and Sclerotinia homeocarpa comprises the microorganism as an effective ingredient. A method for preparing the composition comprises the steps of: (a) after suspending soil in brine to prepare a suspension, culturing the suspension in a culture medium to culture a mixture spawn of soil microorganisms; (b) after mixing 0.001-0.02 parts by weight of the mixture spawn of soil microorganisms, 2-5 parts by weight of rice bran, 2-4 parts by weight of molasses, and 2-4 parts by weight of brown sugar and the remaining parts of water to make a mixture to be 100 parts by weight, culturing the mixture with repetitively aerating the mixture for 1-3 hours 2 to 6 times per day with 5-10 m^3/hour of air with 2-6 hours intervals with maintaining a temperature at 20-25 deg.C for 15-21 days to prepare a mixture microorganism liquid formulation; (c) after mixing 20-30 wt.% of a culturing raw material consisting of rice bran or dried powder of agricultural products with 70-80 wt.% of the mixture microorganism liquid formulation, inoculating 0.01-0.1 wt.% of the mixture spawn of soil microorganisms thereinto; and (d) culturing the mixture obtained from the step(c) at an outside air temperature of 300 deg.C and an inner temperature of 80-85 deg.C for 2-8 hours.

Description

Strains having an antagonistic action against plant pathogens, microorganisms for controlling the same, and a method for producing the same, including the strains

1 is a photograph showing the results of the antimicrobial activity test of a novel strain isolated from the microbial agent of the present invention

Figure 2 is a photograph showing the growth state after 10 days of turf flower pots (treatment) and turf flower pots (control) not treated with microbial agents, each treated with a different throughput.

3 is a graph showing the incidence index displayed by investigating the incidence index according to the time of the treatment and non-microbial treatment treatment treated with different throughput of the microbial agent of the present invention, respectively

Figure 4 is a lawn photo contrasted with a control area treated with a microbial agent of the present invention and the surroundings not treated with a microbial agent

5 is a 16s RNA sequence of Bacillus amyloliquefaciens KS-4, a novel strain having an antagonistic action against the plant pathogens of the present invention.

Figure 6 is a 16s RNA sequence of Bacillus-agaradhaerens KS-5 (Bacillus-agaradhaerens KS-5), a novel strain having an antagonistic action against the plant pathogens of the present invention

Figure 7 is a 16s RNA sequence of Paenibacillus-lentimorbus KS-6 (Paenibacillus-lentimorbus KS-6), a novel strain having an antagonistic action against the plant pathogens of the present invention

8 is a 16s RNA sequence of Bacillus laebolaticus KS-7 (Bacillus-laevolacticus KS-7), a novel strain having an antagonistic action against the plant pathogens of the present invention.

Figure 9 is a 16s RNA sequence of the Leuconostoc paramesenteroides KS-9 (Reuconostoc paramesenteroides KS-9), a novel strain having an antagonistic action against the plant pathogens of the present invention

10 is a 16s RNA sequence of the Kurthia-sibirica KS-13, a novel strain having an antagonistic activity against the plant pathogens of the present invention.

Figure 11 shows the 16s RNA sequence of the sphingoacterium-spiritivorum KS-18 (Sphingobacterium-spiritivorum KS-18), a novel strain having an antagonistic action against the plant pathogens of the present invention

The present invention relates to a novel strain having an antagonistic action against plant pathogens, a microorganism preparation for controlling the same, and a method for preparing the same, and more specifically, Bacillus amyloliquefaciens KS-4 (Bacillus-amyloliquefaciens KS-). 4, accession number: KACC 91247P), Bacillus-agaradhaerens KS-5, accession number: KACC 91248P, Paenibacillus lentimobus KS-6 (Paenibacillus-lentimorbus KS-6 , Accession No .: KACC 91249P), Bacillus Laevolacticus KS-7 (Bacillus-laevolacticus KS-7, Accession No .: KACC 91250P), Leukonostok paramesenteroid KS-9 (Leuconostoc paramesenteroides KS-9, Accession No. : KACC 91251P), Kursia-sibirica KS-13 (Accession No.:KACC 91252P) and Spingobacterium Spiriborum KS-18 (Sphingobacterium-spiritivorum KS-18, Accession No.:KACC 91253P) and at least one selected from the group consisting of Rhizoctoni a solani ) or a strain having an antagonistic action against plant pathogens of Sclerotinia homeocarpa , a microorganism preparation for control, and a method for producing the same.

Along with the increase in population, the crop cultivation environment has also developed remarkably. By the way, there is a lot of influence on the cultivation of such crops are damage caused by wild animals and insects and diseases caused by plant pathogens. As a method of controlling pests, chemical agents called pesticides are mainly used. On the other hand, with the increase of national income, golf is popularized, and as a result, many golf courses are being built. However, in the case of cold grass that is planted in a golf course, large patch disease caused by Rhizoctonia solani or sclerotinia homeocarpa bacteria is caused by a dollar spot pathogen. There is a problem vulnerable to turf disease. In order to reduce such turf diseases, most golf courses are spraying pesticides. However, due to soil and water pollution caused by excessive use of pesticides and pesticides scattered or remaining on grass after spraying, Of course, there is a problem that can adversely affect the health of residents living around the golf course. Therefore, there is a need for microbial agents useful for plant diseases. Korean Patent No. 397796 discloses Streptomyces spp. WYE 20 (KCTC 0341BP) and Streptomyces genus WYE 324 (KCTC 0342BP) for controlling plant growth by controlling fungal pathogens. Microbial agents comprising one or more microbial strains selected from the group consisting of lysatonia solani, piculeria aurase, lyctononia cellilis, sclerocthynia homeocapa, choletotricum graminicola, chol Antimicrobial microbial agents for use in the control of diseases caused by pathogens such as totricum nicotiane, botrytis cineria and spheroceca are disclosed. In addition, Korean Patent Publication No. 294023 has a bactericidal effect bacterium, Bacillus subtilis GB-0365 (Accession No .: KFCC-1071) and Bacillus sp. 017 (Accession No .: KFCC-11070), which provides and separates growth against mold-causing diseases such as gray mold, diarrhea, rash, wilting and brown patches, and large patches. A microbial agent containing the strain having the effect and the freshness maintaining effect after the fruit vegetable harvest is disclosed.

However, conventional microbial preparations including the above documents have not been able to replace chemical pesticides due to the poor persistence of the effects due to strains of microorganisms and difficult culture conditions.

Accordingly, it is an object of the present invention to provide a novel strain having an antagonistic action against plant pathogens without fear of land and water pollution, and to provide a microorganism preparation for control comprising the strain.

Another object of the present invention is to provide an economical method for producing the novel microbial agent.

In order to achieve the above object, the present invention is Bacillus amyloliquefaciens KS-4 (Bacillus-amyloliquefaciens KS-4, Accession No .: KACC 91247P), Bacillus- agaradaerens KS-5 (Bacillus-agaradhaerens KS-5 Accession number: KACC 91248P), Paenibacillus lentimobus KS-6 (Paenibacillus-lentimorbus KS-6, accession number: KACC 91249P), Bacillus Laebolactus KS-7 (Bacillus-laevolacticus KS-7, Deposit Number: KACC 91250P), Leuconostoc paramesenteroides KS-9 (Accession No .: KACC 91251P), Kursia-sibirica KS-13 (Accession No .: KACC 91252P) And Sphingobacterium-spiritivorum KS-18 (Accession Number: KACC 91253P), at least one selected from the group consisting of Rhizoctonia solani or Sclerotinia homeocarpa ) provides a strain having an antagonistic action against the plant pathogens.

In addition, the present invention provides a strain having an antagonistic action to the plant pathogen, characterized in that the plant pathogen is Rhizoctonia solani or Sclerotinia homeocarpa .

In addition, the present invention is Bacillus amyloliquefaciens KS-4 (Bacillus-amyloliquefaciens KS-4, accession number: KACC 91247P), Bacillus- agaradaerens KS-5 (Bacillus-agaradhaerens KS-5, accession number: KACC 91248P), Paenibacillus lentimobus KS-6 (Paenibacillus-lentimorbus KS-6, Accession No .: KACC 91249P), Bacillus Laevolactus KS-7 (Bacillus-laevolacticus KS-7, Accession No .: KACC 91250P) , Leuconostoc paramesenteroides KS-9 (Accession No .: KACC 91251P), Kursia-sibirica KS-13 (Accession No .: KACC 91252P) and Sphingobacterium It contains one or more strains selected from the group consisting of spiritiborum KS-18 (Sphingobacterium-spiritivorum KS-18, Accession No .: KACC 91253P) as an active ingredient, and provides a microbial agent having antagonistic activity against phytopathogenic bacteria. .

In addition, the present invention provides a microbial agent, characterized in that the plant pathogen is Rhizoctonia solani or Sclerotinia homeocarpa .

In addition, the present invention Bacillus subtilis ( Bacillus subtilis ), Bacillus sonorensis (B acillus sonorensis ), Bacillus TUT1206 ( Bacillus sp. TUT1206), Bacillus Q-12 ( Bacillus sp. Q-12) and Bacillus thermoamylo It provides a microbial agent, characterized in that it further comprises one or more soil microorganisms selected from the group consisting of borax ( Bacillus thermoamylovorans ) as an active ingredient.

Another aspect of the present invention comprises the steps of iii) suspending the soil in saline to prepare a suspension, followed by culturing in a medium to culture soil microbial mixed spawn; Ii) Soil microbial mixed spawn 0.001 to 0.02 parts by weight, 2 to 5 parts by weight of rice bran, 2 to 4 parts by weight molasses and 2 to 4 parts by weight of sugar sugar mixed with the remaining water to 100 parts by weight 20 to 20 Maintaining 25 ℃ and aeration of 5 to 10 ㎥ / h of air for 1 to 3 hours at intervals of 2 to 6 hours repeatedly performed 2 to 6 times a day and incubated for 15 to 21 days to prepare a complex microbial liquid step; Iii) inoculated to inoculate 0.01-0.1 wt% of the soil microbial mixed seed after preparing a mixed raw material consisting of 20-30 wt% of rice bran or agricultural by-product dry powder and 70-80 wt% of the mixed microbial solution. Step and iii) is prepared by a method comprising a high temperature culture step of incubating the inoculated mixed raw material for 2 to 8 hours at an ambient temperature of 300 ℃ internal temperature 80 to 85 ℃, Bacillus amyloliquefaciens KS-4 (Bacillus -amyloliquefaciens KS-4, Accession No .: KACC 91247P), Bacillus-agaradhaerens KS-5 (Bacillus-agaradhaerens KS-5, Accession No .: KACC 91248P), Paenibacillus Lentimobus KS-6 (Paenibacillus-lentimorbus KS-6, Accession No .: KACC 91249P), Bacillus Laevolacticus KS-7 (Bacillus-laevolacticus KS-7, Accession No .: KACC 91250P), Leukonostock paramesenteroid KS-9 (Leuconostoc paramesenteroides KS-9 , Accession number: KACC 91251P), Kursi From the group consisting of Ashibirica KS-13 (Kurthia-sibirica KS-13, Accession No .: KACC 91252P) and Sphingobacterium Spiriborum KS-18 (Accession No .: KACC 91253P) It provides a method for producing a microbial agent for controlling the selected one or more strains as an active ingredient.

In addition, the present invention provides a method for producing a microbial preparation, characterized in that the culture material comprises 35 to 75% by weight of rice bran and 25 to 65% by weight of agricultural product by-products.

In another aspect, the present invention provides a method for producing a microbial agent, characterized in that further comprising the step of producing a powder by drying using hot air drying or spray dryer after the high temperature culture step.

In addition, the present invention, the soil microbial mixed species Bacillus subtilis ( Bacillus subtilis ), Bacillus sonorensis ( Bacillus sonorensis ), Bacillus TUT1206 ( Bacillus sp. TUT1206), Bacillus Q-12 ( Bacillus sp. Q-12), Bacillus thermoamylovorans , Leuconostoc paramesenteroides and Pediococcus pentosaceis strain LM2632 It provides a method for producing a microbial agent, characterized in that at least one.

Hereinafter, the present invention will be described in detail.

The novel strains and microbial agents of the present invention are effective for the treatment and prevention of diseases caused by plant pathogens, in particular, Rhizoctonia solani or Sclerotinia homeocarpa . That is, when the strain of the present invention and the microbial agent containing the strain are used for the treatment and prevention of the vegetable diseases caused by Laytonia bacteria or Sterlotini bacteria, it exhibits excellent control effect and excellent durability.

In order to prepare the microbial preparation of the present invention, the present invention is prepared by suspending the soil in saline to prepare a suspension and then cultured in the medium to culture soil microbial mixed seed, the soil microbial mixed seed 0.001-0.02 parts by weight, rice bran 2 to 5 parts by weight, molasses 2 to 4 parts by weight and 2 to 4 parts by weight of brown sugar to mix the remaining water to 100 parts by weight after mixing to maintain a temperature of 20 to 25 ℃ 5 to 5 hours at intervals of 2 to 6 hours Aeration of 10 m 3 / h of air for 1 to 3 hours is carried out repeatedly 2 to 6 times a day and 15 to 21 days of culturing to prepare a microbial liquid preparation. The soil microbial mixed seed means a microorganism obtained by culturing microorganisms grown in a general soil in a medium. In one embodiment of the present invention, the soil microbial mixed seed is Bacillus subtilis ( Bacillus subtilis) or Bacillus sp. subtilis ), B acillus sonorensis , Bacillus TUT1206 ( Bacillus sp. TUT1206), Bacillus Q-12 ( Bacillus sp. Q-12), Bacillus thermoamylovorans , Leukonostock para At least one member selected from the group consisting of Leuconostoc paramesenteroides and Pediococcus pentosaceis strain LM2632. The culture condition is an optimized result of the study of the inventors, and those skilled in the art can add some modifications to the above conditions, but it will be appreciated that such modifications do not depart from the scope of the present invention.

After preparing the composite microbial liquid, the mixed raw material consisting of 20 to 30% by weight of the culture raw material consisting of rice bran or agricultural product by-product dry powder and 70 to 80% by weight of the composite microbial solution, and then 0.01 to 0.01 of the soil microbial mixed seed An inoculation step of inoculating 0.1 wt% is performed. The microbial agent of the present invention may be used as a cultivating material, rice bran or agricultural by-products, etc., which are relatively easy to obtain and have a low cost. The culture raw material may be used alone or in combination with the rice bran or agricultural by-products, but preferably it is made of 35 to 75% by weight of rice bran and 25 to 65% by weight of agricultural by-products. The culture raw material is preferably ground to about 100 mesh to use in powder form.

The mixed and pulverized raw materials are put into a direct incubator, and the water is adjusted with a liquid microorganism in order to maintain an environment suitable for microbial fermentation, and then inoculated with a mixed seed comprising three or more kinds of Bacillus strains or lactic acid bacteria. The mixed spawn is produced as a powder by removing the harmful microorganisms of the soil in the presence of seasonal and environmental diversity, without artificial filtration, through the environmental adaptation process for 6 months on the natural product medium. In the preparation of the microbial preparation according to the present invention, the mixed seed is more preferably inoculated with 0.01 to 0.1% by weight of the total weight of the raw material.

After performing the inoculation step, the inoculated mixed raw material is subjected to a high temperature culture step of incubating for 2 to 8 hours at 80 to 85 ℃. As described above, the microbial preparation method of the present invention is characterized by culturing at ultra high temperature of 80 ℃ or more. Considering that the general microbial culture is generally made in the range of 20 to 40 ℃, it can be seen that the microbial preparation of the present invention is carried out under ultra-high temperature conditions. In particular, in one embodiment of the present invention, the culture material inoculated with the composite spawn was incubated at high temperature while stirring at 80 to 85 ° C. for 2 to 8 hours at a rate of 30 to 80 rpm / min. At this time, the culture temperature is carried out at 80 ℃ or more to suppress the growth of unnecessary microorganisms, it is preferable to perform at 85 ℃ or less to maintain the activity of the microorganism complex according to the present invention. After the high temperature culture, it is possible to obtain a liquid microbial agent through filtration and dilution if necessary, and various excipients and additives, such as minerals (coagulants), alginic acid and salts thereof, organic acids, protective colloid thickeners, etc. Can be added.

After the high temperature culture may further comprise the step of drying the cultured microbial preparation to a solid or powdered microbial preparation. The drying may be any known drying method, such as hot air drying, scattering drying using a spray dryer, etc. within a range in which the microbial agent does not have a concern such as thermal deformation, and there is no particular limitation. In one embodiment of the present invention was carried out hot air drying in the range of 50 to 80 ℃.

Hereinafter, the present invention will be described in more detail with reference to preferred embodiments of the present invention. However, the following examples are merely to help the understanding of the present invention, the scope of the present invention is not limited only to the following examples.

Example (cultivation of complex microbial strain)

After the microorganism-containing soil samples by taking the following, 1g sample prepared based finely ground in a mortar and then heat treatment (composting soil of Chungbuk Jechon Forest) in a 60 ℃ for 30 minutes, suspended in 0.85% NaCl 9㎖, 10 ⁰ Diluted to 10 ⁻⁷ . 100 μl of each dilution suspension was plated in TSA, BL, BBL medium (DIFCO Co.) and incubated at 28 ° C. to incubate the soil microbial complex seedling. As a result of separating and identifying the mixed soil microorganism, Bacillus subtilis ( Bacillus subtilis ), Bacillus sonorensis (B acillus sonorensis ), Bacillus TUT1206 ( Bacillus sp. TUT1206), Bacillus Q-12 ( Bacillus) sp. Q-12), Bacillus thermoamylovorans , Leuconostoc paramesenteroides and Pediococcus pentosaceis strain LM2632. Could know.

Example 2 (Preparation of Microorganisms)

Soil microbial culture 0.01kg, rice bran 4kg, molasses 2kg and 4kg of brown sugar cultured as described above is mixed with water suitable for drinking water standards and mixed so that the weight is 100kg, maintaining 20 to 25 ℃ and 10㎥ every 4 hours Aeration of the air of / h for 2 hours was repeated four times a day and cultured for 21 days to prepare a complex microbial liquid. The complex microbial solution was 4,3 × 10 ⁹ cfu / g.

Separately, the pre-crushed 65kg rice bran and 35kg agricultural by-products were placed in an incubator with a mixing function and mixed for 30 minutes. 30 kg of the mixed microbial liquid was added to the mixed raw material, and the water concentration was adjusted to 70%. The soil microbial complex spawn was inoculated at 0.01% of the total weight of the culture raw material. Ultra-high temperature culture was performed for 4 hours while stirring the inoculated culture raw material at an outside temperature of 300 ° C., an incubator internal temperature of 80 to 85 ° C., and 30 to 80 rpm / min.

Example 3 (Identification of Microorganisms)

Among the microorganisms prepared as described above, colonies of different types were selected and passaged at least three times in TSA medium to separate them into single colonies, and the isolated strains were cultured in liquid and purified genomic DNA was purified by 16S. After amplification using a primer of RNA, the amplified portion was used as a template, and the sequence was sequenced using 519r and 785r primers. The resulting species were identified in the NCBI DB to identify species. The primer sets used to identify bacteria with 16S RNA were described in "Nucleic acid techniques in bacterial systematics (John Wiley and Sons, England)" and "nucleic acid research (2000). 28 (1): 173-174.

In addition, GC-MIDI analysis and BIOLOG analysis were identified in addition to the 16s RNA sequencing. When the results of the 16s RAN sequencing and the results of the GC-MIDI are different, they were identified in a direction following the results of the GC-MIDI. Table 1 below shows the identification results of the novel strains included in the microorganisms of the present invention identified through the 16s RNA sequencing results and GC-MIDI and Biolog analysis results. As shown in Table 1 below, the microbial agent of the present invention includes a total of seven novel strains having antagonism against plant pathogens. 5 to 11 are 16s RNA analysis results for the following novel strains, respectively.

GC Analysis 16s RNA sequencing Biolog analysis No Similarity ks-4 0.769 Bacillus-amyloliquefaciens ← ← ks-5 0.761 Bacillus-agaradhaerens Bacillus sp. Bacillus subtilis C ks-6 0.789 Paenibacillus-lentimorbus ← ← ks-7 0.794 Bacillus-laevolacticus Bacillus sonorensis Bacillus licheniformis ks-9 Leuconostoc paramesenteroides Leuconostoc paramesenteroides ks-13 0.294 Kurthia-sibirica Bacillus oleronius Streptococcus iniae ks-18 0.674 Sphingobacterium-spiritivorum Uncultured bacterium PE02 Sphingobacterium spiritovorum

Bacillus amyloliquefaciens KS-4 (Bacillus-amyloliquefaciens KS-4, Accession No .: KACC 91247P), Bacillus-agarada Eren KS-5 (Bacillus-agaradhaerens KS-5) Accession number: KACC 91248P), Paenibacillus lentimobus KS-6 (Paenibacillus-lentimorbus KS-6, accession number: KACC 91249P), Bacillus Laebolactus KS-7 (Bacillus-laevolacticus KS-7, Deposit Number: KACC 91250P), Leuconostoc paramesenteroides KS-9 (Accession No .: KACC 91251P), Kursia-sibirica KS-13 (Accession No .: KACC 91252P) And Sphingobacterium-spiritivorum KS-18 (Accession Number: KACC 91253P), and as of June 14, 2006, the Korean Institute of Agricultural Microbiology The deposit number was assigned to (KACC).

Example 4 Preparation of Powdered Microbial Agents

The liquid microbial agent prepared in Example 2 was cooled to room temperature (20 to 25 ° C.), followed by hot air drying (60 ° C.) to prepare 92 kg of the microbial agent having a water content of 12%.

Example 5 (antimicrobial activity test of microorganisms)

The microorganisms identified in the above example were tested for antimicrobial activity through an aluminum ring test. The strains isolated and identified in the above example were stored in a -20 ° C. freezer, tryptic soia was transferred to medium (trypticsoy agar, TSA), incubated for 24 hours in a 28 ° C. incubator, and the strains grown in the medium were tryptic soy broth, TSB) and incubated for 48 hours at 28 ℃ in shaking culture.

Separately, the Rhizoctonia solani AG2-2 and Sclerotinia homeocarpa, which are large patch pathogens, were cultured in potato agar medium (PDA).

Wrap one side of a ring with a diameter of 2.7 mm (DO 2.9 mm, DI 2.7 mm) and a height of 10 mm flat with a foil, and then wrap the foil in a petri dish with the ring facing down. Sterilization for 15 minutes at 121 ℃. Potato agar medium (PDA) and TSA medium medium were poured into sterile rings (base agar-PDA 2ml, top agar-TSA 2ml). Each strain identified in the above examples was inoculated onto TSA and incubated at 28 ° C. for 48 hours. After confirming that the bacteria were sufficiently incubated, remove the foil under the inverting ring, and then, the hyphae of Rhizoctonia solani AG2-2 and Sclerotinia homeocarpa were cultured on PDA medium. The cork bore (9 mm diameter) was applied and topped off. Then, after incubating for at least 48 hours at 25 ℃ was tested for antimicrobial activity according to the presence or absence of mycelial growth of pathogens. 1 is a photograph showing the results of the antimicrobial test of the novel strain isolated from the microbial agent of the present invention. The results shown in the photograph are summarized in Table 2 below.

Strain name Lyzoctonia solani AG2-2 Sclerotinia homeocarpa Bacillus-amyloliquefaciens KS-4 + + Bacillus-agaradhaerens KS-5 + + Paenibacillus-lentimorbus KS-6 + + Bacillus-laevolacticus KS-7 + + Leuconostoc paramesenteroides KS-9 + - Kurthia-sibirica K-13 + + Sphingobacterium-spiritivorum K-18 + -

Example 6-1 (Test of Control Effects of Microbial Agents)

The therapeutic effect of the liquid and solid microbial preparations prepared in Example 2 and Example 3 was examined in grass infected with large patches. First of all, the test was conducted in an indoor greenhouse. The indoor greenhouse test was used for pots, the pots used were 0.5m × 0.15m wide, and the soil filled in pots was sterilized for 1 hour at 121 ° C. The pollen was removed from the seedling packaging of New Seoul CC in the potted plants with a hole cutter and transplanted by four bags per pot. One day after transplanting the herds were treated with the liquid and solid microbial preparation of Examples 2 and 3. The throughput is as described in Figure 2 below. Liquid microbial products were irrigated throughout, and solid microbial products were evenly sprayed between grass aerations. The throughput of the microbial preparation was 2 liters / m 2 for the liquid formulation, 100 g / m 2 for the solid formulation, and 4 repetitions were made for each treatment. The turf was grown at 25 ° C. and 53% humidity. 2 and 3 are photographs showing the state of growth after 10 days of turf flower pots (treatment) and turf flower pots (control) not treated with microbial agents, respectively, with different throughputs of the microbial agent of the present invention. It is a graph of the incidence index displayed by investigating the incidence index according to the time of the treatment group treated with different throughputs and the control group not treated with microbial agents. As can be seen in Figures 2 and 3, it can be seen that the microbial agent including the novel strain of the present invention has a low incidence index and a small change over time as its usage increases. The incidence index is 4 repetition averages, 0 in case of no disease, 1 in cases of less than 10% of grass, 2, 30 in case of 10 to 30% of cases, and 3 in cases of 30 to 50% of cases. In the case of 50-75% of cases, 4 and 75% of cases were defined as 5 and quantified.

Example  6-2 (test for controlling effectiveness of microorganisms)

The therapeutic effect of the liquid and solid microbial preparations prepared in Example 2 and Example 3 was examined in grass infected with large patches. Unlike Example 6-1, the test was conducted on an outdoor lawn. In the test, the liquid and solid microbial agents were treated once on the basis of 20 liter / 100 pyeong and 10 Kg / 100 pyeong on the lawn, and the incidence was confirmed at 20 days intervals after the treatment. Figure 4 is a lawn photo contrasted with the control area treated with the microbial agent of the present invention and the microbial agent is not surrounding. As can be seen in Figure 4, it can be seen that the part treated with the microbial agent including the novel strain of the present invention exhibits a therapeutic and preventive effect of Rajpatch disease. Table 3 below summarizes the results of investigation of the incidence index of lawns treated with the microbial agent of the present invention (treatment zone) and lawns not treated with the microbial agent (control, untreated zone) at 20-day intervals.

Onset date after treatment Liquid microbial agent Solid Microorganisms Treatment No treatment Treatment No treatment After 20 days - ++ + ++ 40 days later - +++ + +++ 60 days later - +++ + +++ After 80 days ^* - + - + In 100 days - +++ + +++ 120 days later + +++ + +++

(-: 0%, +: <5%, ++: 5-20%, +++:> 20%) After 80 days of irradiation, the disease is temporarily absent due to the high temperature precursor.

As can be seen in Table 3, as a result of the treatment with the microbial agent of the present invention it can be seen that the incidence is very low and there is almost no change over time even if only one treatment.

As described above, the present invention is a novel soil-derived strain having a antagonism against plant pathogens, in particular, Rhizoctonia solani or Sclerotinia homeocarpa , and a microbial agent comprising the strain. And it provides a method for producing the microbial agent.

The embodiments of the present invention described above should not be construed as limiting the technical idea of the present invention. The protection scope of the present invention is limited only by the matters described in the claims, and those skilled in the art can change and change the technical idea of the present invention in various forms. Therefore, such improvements and modifications will fall within the protection scope of the present invention, as will be apparent to those skilled in the art.

Claims (9)

Bacillus-amyloliquefaciens KS-4 (Accession No .: KACC 91247P), Bacillus-agaradhaerens KS-5 (Bacillus-agaradhaerens KS-5, Accession No .: KACC 91248P) Bacillus lentibusus KS-6 (Paenibacillus-lentimorbus KS-6, Accession No .: KACC 91249P), Bacillus Laevolactus KS-7 (Bacillus-laevolacticus KS-7, Accession No .: KACC 91250P), Leukonostock Para Mesenteroid KS-9 (Leuconostoc paramesenteroides KS-9, Accession No .: KACC 91251P), Kursia-sibirica KS-13 (Accession No .: KACC 91252P) and Sphingobacterium Spiriborum KS At least one selected from the group consisting of -18 (Sphingobacterium-spiritivorum KS-18, Accession No .: KACC 91253P), the strain having antagonistic activity against vegetable pathogens. The method of claim 1, The plant pathogen is a strain having antagonism to the plant pathogen, characterized in that the Rhizoctonia solani or Sclerotinia homeocarpa . Bacillus-amyloliquefaciens KS-4 (Accession No .: KACC 91247P), Bacillus-agaradhaerens KS-5 (Bacillus-agaradhaerens KS-5, Accession No .: KACC 91248P) Bacillus lentibusus KS-6 (Paenibacillus-lentimorbus KS-6, Accession No .: KACC 91249P), Bacillus Laevolactus KS-7 (Bacillus-laevolacticus KS-7, Accession No .: KACC 91250P), Leukonostock Para Mesenteroid KS-9 (Leuconostoc paramesenteroides KS-9, Accession No .: KACC 91251P), Kursia-sibirica KS-13 (Accession No .: KACC 91252P) and Sphingobacterium Spiriborum KS It contains at least one strain selected from the group consisting of -18 (Sphingobacterium-spiritivorum KS-18, Accession No .: KACC 91253P) as an active ingredient, the plant pathogen Rhizoctonia solani or Sclerotinia homeocarpa It has the effect of the treatment and prevention of large patch disease by the bacteria Microbial agent to a gong. delete delete Iii) suspending the soil in saline to prepare a suspension, and then culturing in a medium to culture soil microbial mixed spawn; Ii) Soil microbial mixed spawn 0.001 to 0.02 parts by weight, 2 to 5 parts by weight of rice bran, 2 to 4 parts by weight molasses and 2 to 4 parts by weight of sugar sugar mixed with the remaining water to 100 parts by weight 20 to 20 Maintaining 25 ℃ and aeration of 5 to 10 ㎥ / h of air for 1 to 3 hours at intervals of 2 to 6 hours repeatedly performed 2 to 6 times a day and incubated for 15 to 21 days to prepare a complex microbial liquid step; Iii) inoculated to inoculate 0.01-0.1 wt% of the soil microbial mixed seed after preparing a mixed raw material consisting of 20-30 wt% of rice bran or agricultural by-product dry powder and 70-80 wt% of the mixed microbial solution. step; Iii) the inoculated mixed raw material is prepared by a method comprising a high-temperature culturing step of incubating for 2 to 8 hours at an ambient temperature of 300 ° C. and an internal temperature of 80 to 85 ° C., Bacillus amyloliquefaciens KS-4 (Bacillus-amyloliquefaciens) KS-4, Accession No .: KACC 91247P), Bacillus-agaradhaerens KS-5 (Accession No .: KACC 91248P), Paenibacillus Lentimobus KS-6 (Paenibacillus-lentimorbus KS- 6, accession number: KACC 91249P), Bacillus laevolacticus KS-7 (Bacillus-laevolacticus KS-7, accession number: KACC 91250P), leukonostock paramesenteroid KS-9 (Leuconostoc paramesenteroides KS-9, deposit KACC 91251P), Kurthia-sibirica KS-13 (Accession No .: KACC 91252P) and Spingobacterium Spiriborum KS-18 (Sphingobacterium-spiritivorum KS-18, Accession No .: KACC 91253P) room containing one or more strains selected from the group consisting of The method of microbial preparations. The method of claim 6, The culture material is a method for producing a microbial agent, characterized in that it comprises 35 to 75% by weight of rice bran and 25 to 65% by weight of the by-product powder. The method according to claim 6 or 7, After the high temperature culture step, using a hot air drying or spray dryer to dry the manufacturing method of the microbial agent further comprising the step of preparing in powder form. The method according to claim 6 or 7, The soil microbial mixed species is Bacillus subtilis ( Bacillus subtilis ), Bacillus sonorensis (B acillus sonorensis ), Bacillus TUT1206 ( Bacillus sp. TUT1206), Bacillus Q-12 ( Bacillus sp. Q- 12), Bacillus thermoamylovorans , Leuconostoc paramesenteroides and Pediococcus pentosaceis strain LM2632, characterized in that at least one selected from the group consisting of. Method for producing a microbial agent.

Images