KR101994072B1 - Biological microbial treating agent for removing agrochemical - Google Patents

Abstract

The present invention provides a biological microbial treating agent for removing agrochemical components, including at least one microorganism selected among Bacillus amyloliquefaciens KS-R01, Bacillus siamensis KS-R02, Bacillus velezensis KS-R03, and Bacillus tequilensis KS-R04.

Description

TECHNICAL FIELD [0001] The present invention relates to a biological microbial treatment agent for removing pesticide ingredients,

More particularly, the present invention relates to a biological microorganism treating agent which is resistant to radioactive substances and pesticide ingredients and is excellent in ability to remove pesticide residues present in a contaminated area, thereby restoring contaminated water quality or soil And to a biological microorganism treating agent for removing pesticide ingredients capable of preventing environmental pollution and restoring the environment.

Pesticides are essential agricultural material for agriculture. Since 1990, 24 ~ 25,000 tons of pesticides have been distributed to farmland every year, contributing to the improvement of quality of agricultural products and the increase of yield.

However, contrary to the fact that pesticides have contributed greatly to the production of safe agricultural products, due to their inherent toxicity, the pesticides used must be rapidly degraded after controlling the target specific pests or weeds, so that the adverse effects on the environment can be minimized.

Only 10 to 20% of the pesticide applied to the crop is attached to the crop and the rest is absorbed into the crop again by the soil or remains in the soil. Some of the pesticide is transported to the water by the soil water, Or recirculated to soil or water.

Therefore, a large part of the pesticide applied to the soil is integrated into the soil, and understanding of the behavior of the integrated pesticide in the soil can be used as valuable data for evaluating the degree of the pesticide hazard as well as the conservation of the soil environment.

As a method for removing pesticide residues remaining in the soil up to now, there has been disclosed in Korean Patent Laid-Open No. 10-2012-0022346 (Mar. 12, 2012), but these methods are performed through various steps and chemical agents Which may lead to secondary environmental pollution, such as the need to be treated.

Although biological removal methods capable of removing pesticide residues in contaminated areas with resistance can be considered, research and report on biological microorganism treatment agents capable of preventing environmental pollution and restoring environment due to residual pesticides have been made yet It is not getting.

Disclosure of Invention Technical Problem [8] Accordingly, the present invention has been made in order to solve the problems of the prior art as described above, and its object is to provide a pesticidal composition which is resistant to radioactive materials and pesticide ingredients, The present invention provides a biological microorganism treating agent for removing pesticide components that can prevent environmental pollution and restore environment by restoring polluted water quality or soil.

The technical problem of the present invention as described above is achieved by the following means.

(1) Bacillus amyloliquefaciens KS-R01, Bacillus siamensis KS-R02, Bacillus velezensis KS-R03, and Bacillus tequilensis . A biological microorganism treating agent for removing an agricultural chemical component comprising at least one microorganism selected from KS-R04.

(2) The method according to claim 1,

Bacillus amyloliquefaciens KS-R01, Bacillus siamensis KS-R02, Bacillus velezensis KS-R03, and Bacillus tequilensis KS-R04 A biological microorganism treatment agent for removing the pesticide component

(3) A method for removing pesticide components by treating the microorganism treating agent of claim 1 or 2 with water or soil contaminated with pesticide ingredients.

According to the present invention, resistance to radioactive materials and pesticide ingredients is excellent and ability to remove pesticide ingredients is excellent, thereby restoring contaminated water quality or soil, thereby preventing environmental pollution and restoring the environment.

1 is a graph showing the degree of colony formation by pesticide treatment of four kinds of the starting strains according to the present invention,
Fig. 2 shows the survival curves of the four kinds of starting strains according to the present invention by treatment with the pesticide ingredient.

Hereinafter, the contents of the present invention will be described in more detail.

Bacillus amyloliquefaciens KS-R01, Bacillus siamensis KS-R02, Bacillus velezensis KS-R03, and Bacillus tezulensis as described above. ( Bacillus tequilensis ) KS-R04. The present invention also provides a biological microorganism treating agent for removing pesticide components comprising at least one microorganism selected from Bacillus tequilensis KS- RO4 .

The microorganism used in the present invention is preferably selected from the group consisting of Bacillus amyloliquefaciens KS-R01 (KCTC 13558BP), Bacillus siamensis KS-R02 (KCTC 13559BP), Bacillus buretensis Bacillus velezensis KS-R03 (KCTC 13560BP), and Bacillus tequilensis KS-R04 (KCTC 13561BP). The present invention also provides a biological microorganism treating agent for the removal of pesticide components comprising at least one microorganism selected from the group consisting of Bacillus velezensis KS-

The biological microorganism treating agent according to the present invention is not particularly limited, but each microorganism strain may be mixed in a ratio of 1 to 30% by weight. The biological microorganism treating agent according to the present invention is a mixture of the above four microorganisms, and exhibits a synergistic action to produce a synergistic effect in a region contaminated with radioactive material such as sea, river, lake or soil, contaminated with pesticide ingredient, And other objects contaminated with pesticide ingredients, it is possible to remove radioactive materials and pesticide components in an environmentally friendly manner.

Bacillus amyl Lowry Quebec Pacific Enschede (Bacillus amyloliquefaciens) KS-R01 ( KCTC 13558BP), Bacillus Xiamen sheath (Bacillus siamensis) KS-R02 ( KCTC 13559BP), Bacillus Belle Zen sheath (Bacillus constituting the biological microorganism agent according to the invention velezensis) KS-R03 (KCTC 13560BP ), and Bacillus Te kwilren system (Bacillus tequilensis) KS-R04 ( KCTC 13561BP) were both deposited with the Korea Research Institute of Bioscience and biotechnology gene bank dated June 21, 2018.

The biological microorganism treating agent according to the present invention containing the above microorganisms can be produced by the following method.

0.001 to 0.02% by weight of a microorganism strain consisting of the above microbial strains (at least one of the above four), 2 to 5% by weight of rice bran, 2 to 4% by weight of molasses and 2 to 4% 100 to 500 L / h of air is aerated for 1 to 3 hours at intervals of 2 to 6 hours while maintaining the temperature at 20 to 25 ° C. For 15 to 21 days to prepare a liquid microorganism. The conditions such as aeration are optimized results of the present inventors, and those skilled in the art will recognize that some modifications may be made to the above conditions, but such modifications do not depart from the scope of the present invention.

After the preparation of such a liquid microbicide, a mixed raw material composed of 20 to 30% by weight of at least one culture raw material selected from the group consisting of rice bran, agricultural by-products and agricultural forest product dry powder and 70 to 80% by weight of the liquid microbicide was prepared The seeding step of inoculating 0.01 to 0.1% by weight of the microorganism strain according to the present invention can be carried out.

The biological microorganism treating agent of the present invention can be used as a raw material for culturing microorganisms, such as rice bran, agricultural by-products or agro-forest products, which are relatively easy to obtain and cost less. The culture material may be used alone or in combination with the rice bran, agricultural by-products or agro-forest products, preferably 35-45 wt% rice bran, 25-35 wt% agricultural by-products, 20-30 wt% Of the composition. It is preferable that the culture material is pulverized to about 100 mesh in advance and used as a powder. The mixed and pulverized raw materials for cultivation may be put into a flask and the water may be adjusted with a liquid fermentation agent to maintain an appropriate environment for microbial fermentation.

After the inoculation step is performed, the inoculated mixed material is subjected to a high-temperature culture step of culturing at 80 to 85 ° C for 2 to 8 hours. The method for producing a biological microorganism treating agent of the present invention is characterized in that it is cultured at an ultra-high temperature of 80 ° C or higher as described above. Considering that ordinary microorganism culturing is generally performed at a temperature in the range of 20 to 40 ° C, it can be seen that the biological microorganism treating agent of the present invention is carried out at an ultra-high temperature condition. In particular, in one embodiment of the present invention, the cultured raw material inoculated with the mixed seed is cultured at 80 to 85 ° C for 2 to 8 hours at an ultra-high temperature of 30 to 80 rpm / min while stirring. At this time, the incubation temperature is preferably 80 ° C. or higher to inhibit the growth of unnecessary microorganisms, but it is preferably carried out at 85 ° C. or lower to maintain the activity of the microorganism complex according to the present invention.

The method of introducing the biological microorganism treating agent which can be produced as described above is characterized in that a culture tank containing the biological microorganism treating agent according to the present invention is directly sprayed on the pesticide contaminated site using an atomizer or the like, It can be simply handled by direct injection. The biological microorganism treating agent to be added varies depending on the pollutant source of the pesticide ingredient, but generally, the activating solution is made 10 to 30 times as much as the stock solution of the microorganism preparation, and is dosed several times every 1 to 3 days every day.

The microbial agent stock solution is prepared by adding 0.1 to 1.0 part by weight, preferably 0.2 to 0.7 part by weight, and most preferably 0.5 part by weight of the microbial preparation to 100 parts by weight of the water, adding 0.5 to 2.0 parts by weight of molasses, 0.8 to 1.2 parts by weight, and most preferably 1 part by weight, is used, which is cultured while being aerated for about 72 hours while maintaining the temperature at 18 to 25 占 폚. For example, in the preparation of the 10-times-activating solution, 10-fold amount of water may be added to the microbial agent stock solution at a weight ratio of 0.2: 1, and 0.2% by weight of molasses may be added. Thereafter, the mixture is incubated for about 48 hours while being maintained at 18 to 25 캜 to obtain the activated solution.

Hereinafter, the present invention will be described in detail with reference to Examples and Test Examples. However, these are for the purpose of illustrating the present invention in more detail, and the scope of the present invention is not limited thereto.

[Experimental Example 1] Isolation of radioactive resistant strains

1. Radioactive resistance of KS50 complex microorganism

The Co-60 (cobalt 60) gamma ray was irradiated at 0, 4, 6, 8, and 10 kGy for the KS50, KS50, and powder products and liquid products, which were developed by KOGAN CO., LTD.

As shown in Tables 1 and 2, there were 3.3 × ^{10 7} , 5.8 × 10 ⁶ , and 4.2 × 10 ⁶ CFU in the TSA medium for the KS50 seedlings (F2), the powdery products (F3) and the liquid product (F4) the ^{1.0x10 6, 4.8x10 6, 6.3x10 6} CFU were present in the YM medium

The maximum survivable dose in the TSA medium was 10 kGy for F2, 4 kGy for F3, and 10 kGy for F4, which were 5.0 × 10 ¹ , 1.9 × 10 ³ and 5 colonies, respectively. The YM medium is 1.0x10 ² CFU was identified as 5.0x10 ² CFU, F4 is 6 kGy as 5.0x10 ¹ CFU, F3 is 6 kGy as for F2 4 kGy.

From these results, it was confirmed that the KS50 complex microorganism seeds and products contain many radioactive resistant microorganisms.

The number of viable cells varied according to the composition of the media, but the number of viable cells decreased as the dose of radioactive irradiation increased. Especially, when the radioactive dose was increased to 4kGy, the viable cell number decreased sharply. Therefore, the radioactive dose to select effective radioactive resistant microorganisms was selected to be 4 kGy or more.

Measurement of viable cell count in TSA medium (CFU / mL) TSA (BD) Radiation dose (kGy) 0 0.5 One 2 4 6 8 10 F2 3.3 x 10 ⁷ 1.0 x 10 ⁶ 2.5x10 ⁶ 6.3x10 ⁵ 1.2x10 ⁴ 1.0x10 ² 1.0x10 ² 5.0x10 ¹ F3 5.8 x 10 ⁶ 1.5x10 ⁶ 3.8x10 ⁵ 6.3x10 ⁴ 1.9x10 ³ ND ND ND F4 4.2 x 10 ⁶ 6.8x10 ⁵ 2.9x10 ⁵ 4.5x10 ⁴ 3.5x10 ² 5.0x10 ¹ 5.0 5.0

* ND: not detected

Measurement of viable cell counts in YM medium (CFU / mL) YM (BD) Radiation dose (kGy) 0 0.5 One 2 4 6 8 10 F2 1.0 x 10 ⁶ 2.5x10 ⁶ 1.5x10 ⁶ 1.5x10 ⁵ 5.0x10 ¹ ND ND ND F3 4.8x10 ⁶ 8.0x10 ⁶ 8.5x10 ⁶ 2.5x10 ⁶ 4.5 x 10 ³ 5.0x10 ² ND ND F4 6.3x10 ⁶ 1.2x10 ⁶ 6.0x10 ⁵ 2.8 x 10 ⁴ 1.5x10 ² 1.0x10 ² ND ND * ND: not detected

2. Isolation of radioactive resistance strains from KS50 complex microorganism

The surviving strains were isolated from primary Co-60 (cobalt 60) radioactivity and researched at a dose of 4 to 15 kGy to isolate and isolate the surviving strains. A total of 22 microorganisms were isolated on TSA medium and total 14 microorganisms were isolated on YM medium and irradiated with gamma irradiation at doses of 4, 6, 8, 10, 11, 12, 13, and 15 kGy. As shown in Tables 3 and 4, T11-6k is 8 kGy, T12-11k is 11 kGy, T14-4k is 4 kGy, T15-4k is 4 kGy, T16-11k is 11 kGy, T17-6k is 6 kGy, T18 6k was survived at 6 kGy and T19-8k survived at 8 kGy dose. Y6-6k survived at 6 kGy, Y7-8k at 8 kGy, and Y11-6k at 6 kGy. As described above, 9 microorganisms were isolated from 6 kGy dose, 5 microorganisms survived at 8 kGy, and 2 microorganisms survived at 11 kGy.

The microorganisms survived above the irradiation dose of 4 kGy were found to be microorganisms detected in the liquid product cultured in the liquid medium using KS50 complex microorganism seeds.

The radioactive resistance (CFU / mL) of strains isolated from TSA medium Name Primary radioactive irradiation Secondary irradiation (kGy) Id  Information Viable cell count 4 6 8 10 11 12 13 15 T1 F2-10kGy 5.0x10 ¹ - - - - - - - - T2 F2-6kGy 5.0x10 ² - - - - - - - - T3 F2-6kGy 5.0x10 ² - - - - - - - - T4 F2-6kGy 5.0x10 ² - - - - - - - - T5 F2-4kGy 4.5 x 10 ³ - - - - - - - - T6 F2-4kGy 4.5 x 10 ³ - - - - - - - - T7 F2-4kGy 4.5 x 10 ³ - - - - - - - - T8 F2-4kGy 2.0x10 ⁴ - - - - - - - - T9 F2-4kGy 2.0x10 ⁴ - - - - - - - - T10 F2-4kGy 2.0x10 ⁴ - - - - - - - - T11 F4-8kGy 2.5x10 ¹ O O O - - - - - T11-6k T12 F4-8kGy 2.5x10 ¹ O O O O O - - - T12-11k T13 F4-8kGy 2.5x10 ¹ O - - - - - - - T14 F4-8kGy 2.5x10 ¹ O - - - - - - - T14-4k T15 F4-6kGy 5.0x10 ¹ O - - - - - - - T15-4k T16 F4-6kGy 5.0x10 ¹ O O O O O - - - T16-11k T17 F4-6kGy 2.0x10 ² O O - - - - - - T17-6k T18 F4-6kGy 2.0x10 ² O O - - - - - - T18-6k T19 F4-6kGy 2.0x10 ² O O O - - - - - T19-8k T20 F3-8kGy 2 - - - - - - - - T21 F3-8kGy 2 - - - - - - - - T22 F2-8kGy 2.5x10 ² - - - - - - - -

The radioactivity (CFU / mL) of microorganisms isolated from YM medium Name Primary radioactive irradiation Secondary radioactive irradiation Id  Information Viable cell count 4 6 8 10 11 12 13 15 Y1 F4-8kGy <9 - - - - - - - - Y2 F4-8kGy <9 - - - - - - - - Y3 F4-8kGy <9 - - - - - - - - Y4 F4-8kGy <9 - - - - - - - - Y5 F4-6kGy 1x10 ² - - - - - - - - Y6 F4-6kGy 1x10 ² O O - - - - - - Y6-6k Y7 F4-4kGy 1.5x10 ² O O O O O - - - Y7-8k Y8 F4-4kGy 1.5x10 ² - - - - - - - - Y9 F4-4kGy 1.5x10 ² - - - - - - - - Y10 F4-4kGy 1.5x10 ² - - - - - - - - Y11 F4-4kGy 1.5x10 ² O O - - - - - - Y11-6k Y12 F4-4kGy 1.5x10 ² - - - - - - - - Y13 F3-2kGy 2.5x10 ⁶ - - - - - - - - Y14 F2-2kGy 1.5x10 ⁵ - - - - - - - -

* Information notation: Sample type - Radiation dose * TSA medium: Tryptic soy agar, incubation at 35 ℃ for 24 hours

* YM medium: Yeast and mold media, culture at 30 ° C for 48 hours

* Colony Generation Notation - "O"

* The identification of the final ID for identification of strains should be based on the secondary irradiation survey

3. Identification of radioactive resistant microorganisms

A total of 36 strains were isolated from the surviving samples (Table 3 and 4) and the radioactivity was irradiated at a dose ranging from 4 to 15 kGy. The viable microorganisms were analyzed for 16S rRNA gene sequences The microorganisms were identified on the basis of.

As shown in Table 6, the microorganisms surviving at an irradiation dose of 11 kGy were Bacillus amyloliquefaciens (T16-11k) and Bacillus siamensis (T16-11k) T12-11k), the surviving microorganisms in the microbial survival in 8kGy Bacillus velezensis (T19-8k), 6kGy was identified to be Bacillus tequilensis (Y11-6k). Radioactive resistant microorganism is shone all appear as Bacillus bacteria, Bacillus spore forms a resistance in extreme environments such as ultraviolet light, high temperature, low temperature and high pressure is known to have resistance to extreme environmental conditions. Microorganisms surviving at a radiation dose of 4 kGy or more are believed to be resistant to resistant spores. The analyzed 16S base sequences are shown in Tables 7, 8, 9 and 10.

Results of identification of 16S strains in 28 strains No. ID Flour Similarity Diff / Total One T16-11k Bacillus amyloliquefaciens 99.86 2/1472 2 T12-11k Bacillus siamensis 99.86 2/1472 3 T19-8k Bacillus velezensis 99.86 2/1403 4-1 T16-8k-b-1 Bacillus velezensis 99.86 2/1403 4-2 T16-8k-b-2 Bacillus amyloliquefaciens 99.8 3/1472 4-3 T16-8k-b-3 Bacillus amyloliquefaciens 99.86 2/1471 4-4 T16-8k-b-4-a Bacillus amyloliquefaciens 99.86 2/1471 4-5 T16-8k-b-4-b Bacillus amyloliquefaciens 99.86 2/1471 5-1 T12-8k-b-1 Bacillus velezensis 99.86 2/1403 5-2 T12-8k-b-2 Bacillus velezensis 99.86 2/1403 5-3 T12-8k-b-3 Bacillus velezensis 99.86 2/1403 6 T19-6k Bacillus velezensis 99.86 2/1403 7 T18-6k Bacillus velezensis 99.86 2/1403 8 T17-6k Bacillus siamensis 99.93 1/1472 9 T11-6k Bacillus amyloliquefaciens 99.8 3/1472 10-1 T15-4k-a-1 Bacillus tequilensis 99.93 1/1472 10-2 T15-4k-a-2 Bacillus tequilensis 99.93 1/1472 10-3 T15-4k-b Bacillus tequilensis 99.93 1/1472 10-4 T15-4k-c-1 Bacillus tequilensis 99.93 1/1472 10-5 T15-4k-c-2 Bacillus tequilensis 99.93 1/1472 10-6 T15-4k-d Bacillus tequilensis 99.93 1/1472 10-7 T15-4k-e Bacillus tequilensis 99.93 1/1472 10-8 T15-4k-f Bacillus tequilensis 99.93 1/1472 11 T14-4k Bacillus amyloliquefaciens 99.8 3/1472 12 Y7-8k Bacillus amyloliquefaciens 99.8 3/1472 13 Y11-6k Bacillus tequilensis 99.93 1/1472 14 Y6-6k Bacillus amyloliquefaciens 99.8 3/1472 15 Y7-4k Bacillus amyloliquefaciens 99.8 3/1472

Of the 28 strains, four non-redundant strains with high radioactive resistance No. ID Flour Similarity Diff / Total One T16-11k Bacillus amyloliquefaciens 99.86 2/1472 2 T12-11k Bacillus siamensis 99.86 2/1472 3 T19-8k Bacillus velezensis 99.86 2/1403 4 Y11-6k Bacillus tequilensis 99.93 1/1472

SEQ ID NO : 1 : 16S nucleotide sequence of T16-11k ( Bacillus amyloliquefaciens )

GACGAACGCTGGCGGCGTGCCTAATACATGCAAGTCGAGCGGACAGATGGGAGCTTGCTCCCTGATGTTAGCGGCGGACGGGTGAGTAACACGTGGGTAACCTGCCTGTAAGACTGGGATAACTCCGGGAAACCGGGGCTAATACCGGATGCTTGTTTGAACCGCATGGTTCAGACATAAAAGGTGGCTTCGGCTACCACTTACAGATGGACCCGCGGCGCATTAGCTAGTTGGTGAGGTAACGGCTCACCAAGGCGACGATGCGTAGCCGACCTGAGAGGGTGATCGGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCGCAATGGACGAAAGTCTGACGGAGCAACGCCGCGTGAGTGATGAAGGTTTTCGGATCGTAAAGCTCTGTTGTTAGGGAAGAACAAGTGCCGTTCAAATAGGGCGGCACCTTGACGGTACCTAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGAATTATTGGGCGTAAAGGGCTCGCAGGCGGTTTCTTAAGTCTGATGTGAAAGCCCCCGGCTCAACCGGGGAGGGTCATTGGAAACTGGGGAACTTGAGTGCAGAAGAGGAGAGTGGAATTCCACGTGTAGCGGTGAAATGCGTAGAGATGTGGAGGAACACCAGTGGCGAAGGCGACTCTCTGGTCTGTAACTGACGCTgAGGAGCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAAGTGTTAGGGGGTTTCCGCCCCTTAGTGCTGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGGTCGCAAGACTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCTCTGACAATCCTAGAGATAG GACGTCCCCTTCGGGGGCAGAGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGgTTAAGTCCCGCAACGAGCGCAACCCTTGATCTTAGTTGCCAGCATTCAGTTGGGCACTCTAAGGTGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGGCAGAACAAAGGGCAGCGAAACCGCGAGGTTAAGCCAATCCCACAAATCTGTTCTCAGTTCGGATCGCAGTCTGCAACTCGACTGCGTGAAGCTGGAATCGCTAGTAATCGCGGATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTTTTGGAGCCAGCCGCCGAAGGTGGGACAGATGATTGGGGTG

Sequence 2 : 16S base sequence of T12-11k ( Bacillus siamensis )

GACGAACGCTGGCGGCGTGCCTAATACATGCAAGTCGAGCGGACAGATGGGAGCTTGCTCCCTGATGTTAGCGGCGGACGGGTGAGTAACACGTGGGTAACCTGCCTGTAAGACTGGGATAACTCCGGGAAACCGGGGCTAATACCGGATGGTTGTCTGAACCGCATGGTTCAGACATAAAAGGTGGCTTCGGCTACCACTTACAGATGGACCCGCGGCGCATTAGCTAGTTGGTGAGGTAACGGCTCACCAAGGCGACGATGCGTAGCCgACCTGAGAGGGTGATCGGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCGCAATGGACGAAAGTCTGACGGAGCAACGCCGCGTGAGTGATGAAGGTTTTCGGATCGTAAAGCTCTGTTGTTAGGGAAGAACAAGTGCCGTTCAAATAGGGCGGCACCTTGACGGTACCTAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGAATTATTGGGCGTAAAGGGCTCGCAGGCGGTTTCTTAAGTCTGATGTGAAAGCCCCCGGCTCAACCGGGGAGGGTCATTGGAAACTGGGGAACTTGAGTGCAGAAGAGGAGAGTGGAATTCCACGTGTAGCGGTGAAATGCGTAGAGATGTGGAGGAACACCAGTGGCGAAGGCGACTCTCTGGTCTGTAACTGACGCTgAGGAGCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAAGTGTTAGGGGGTTTCCGCCCCTTAGTGCTGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGGTCGCAAGACTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCTCTGACAATCCTAGAGATAG GACGTCCCCTTCGGGGGCAGAGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGATCTTAGTTGCCAGCATTCAGTTGGGCACTCTAAGGTGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGACAGAACAAAGGGCAGCGAAACCGCGAGGTTAAGCCAATCCCACAAATCTGTTCTCAGTTCGGATCGCAGTCTGCAACTCGACTGCGTGAAGCtGGAATCGCTAGTAATCGCGGATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTTATGGAGCCAGCCGCCGAAGGTGGGACAGATGATTGGGGTG

Sequence 3 : 16S base sequence of T19-8k ( Bacillus velezensis )

GACGAACGCTGGCGGCGTGCCTAATACATGCAAGTCGAGCGGACAGATGGGAGCTTGCTCCCTGATGTTAGCGGCGGACGGGTGAGTAACACGTGGGTAACCTGCCTGTAAGACTGGGATAACTCCGGGAAACCGGGGCTAATACCGGATGGTTGTTTGAACCGCATGGTTCAGACATAAAAGGTGGCTTCGGCTACCACTTACAGATGGACCCGCGGCGCATTAGCTAGTTGGTGAGGTAACGGCTCACCAAGGCGACGATGCGTAGCCGACCTGAGAGGGTGATCGGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCGCAATGGACGAAAGTCTGACGGAGCAACGCCGCGTGAGTGATGAAGGTTTTCGGATCGTAAAGCTCTGTTGTTAGGGAAGAACAAGTGCCGTTCAAATAGGGCGGCACCTTGACGGTACCTAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGAATTATTGGGCGTAAAGGGCTCGCAGGCGGTTTCTTAAGTCTGATGTGAAAGCCCCCGGCTCAACCGGGGAGGGTCATTGGAAACTGGGGAACTTGAGTGCAGAAGAGGAGAGTGGAATTCCACGTGTAGCGGTGAAATGCGTAGAGATGTGGAGGAACACCAGTGGCGAAGGCGACTCTCTGGTCTGTAACTGACGCTgAGGAGCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAAGTGTTAGGGGGTTTCCGCCCCTTAGTGCTGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGGTCGCAAGACTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCTCTGACAATCCTAGAGATAG GACGTCCCCTTCGGGGGCAGAGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGATCTTAGTTGCCAGCATTCAGTTGGGCACTCTAAGGTGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGACAGAACAAAGGGCAGCGAAACCGCGAGGTTAAGCCAATCCCACAAATCTGTTCTCAGTTCGGATCGCAGTCTGCAACTCGACTGCGTGAAGCTGGAATCGCTAGTAATCGCGGATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTTTAGGAGCCAGCCGCCGAAGGTGGGACAGATGATTGGGGTG

Sequence 4 : 16S base sequence of Y11-6k ( Bacillus tequilensis )

GACGAACGCTGGCGGCGTGCCTAATACATGCAAGTCGAGCGGACAGATGGGAGCTTGCTCCCTGATGTTAGCGGCGGACGGGTGAGTAACACGTGGGTAACCTGCCTGTAAGACTGGGATAACTCCGGGAAACCGGGGCTAATACCGGATGGTTGTTTGAACCGCATGGTTCAAACATAAAAGGTGGCTTCGGCTACCACTTACAGATGGACCCGCGGCGCATTAGCTAGTTGGTGAGGTAACGGCTCACCAAGGCAACGATGCGTAGCCGACCTGAGAGGGTGATCGGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCGCAATGGACGAAAGTCTGACGGAGCAACGCCGCGTGAGTGATGAAGGTTTTCGGATCGTAAAGCTCTGTTGTTAGGGAAGAACAAGTACCGTTCGAATAGGGCGGTACCTTGACGGTACCTAACCAGAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGAATTATTGGGCGTAAAGGGCTCGCAGGCGGTTTCTTAAGTCTGATGTGAAAGCCCCCGGCTCAACCGGGGAGGGTCATTGGAAACTGGGGAACTTGAGTGCAGAAGAGGAGAGTGGAATTCCACGTGTAGCGGTGAAATGCGTAGAGATGTGGAGGAACACCAGTGGCGAAGGCGACTCTCTGGTCTGTAACTGACGCTGAGGAGCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAAGTGTTAGGGGGTTTCCGCCCCTTAGTGCTGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGGTCGCAAGACTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCTCTGACAATCCTAGAGATAG GACGTCCCCTTCGGGGGCAGAGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGATCTTAGTTGCCAGCATTCAGTTGGGCACTCTAAGGTGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGACAGAACAAAGGGCAGCGAAACCGCGAGGTTAAGCCAATCCCACAAATCTGTTCTCAGTTCGGATCGCAGTCTGCAACTCGACTGCGTGAAGCTGGAATCGCTAGTAATCGCGGATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTTTAGGAGCCAGCCGCCGAAGGTGGGACAGATGATTGGGGTG

4. Radioactive resistance of selected strain

1). In the case of the T16 and T19 strains, colony was not formed at an irradiation dose of 2 kGy or more, whereas Y11 and T12 strains (Fig. 1) were irradiated with four kinds of strains irradiated with radioactivity at a dose of 0.5 to 4 kGy The surviving Colony was observed even at a dose of 2 kGy. The survival rate of the strain Y11 was about 0.001% at 2 kGy, while the survival rate of T12 strain was 2% at 2 kGy and 0.7% at 4 kGy. In addition, the results of confirming the survival rate trend according to dose (FIG. 2) showed that the T16 strain was -1.1, the Y11 strain was -1.9, and the T19 strain was -2.5, while the T12 strain was -0.48. Respectively.

To determine quantitative radioactive resistance, the D10 value of the strain was calculated (Table 7). Based on the survival rate, the growth curve trend line of each strain could be obtained as follows.

Radioactive resistance of isolated microorganism Sample b a D10 value (kGy) T16 0.078 -1.13 0.95 T19 0.053 -2.00 0.53 Y11 0.135 -1.94 0.58 T12 -0.46 -0.48 1.13

Based on the survival curves of each strain, the D10 value (the dose at which 90% of the strain was killed) was calculated. As a result, the D10 value of the T12 strain was 1.13, which was higher than that of the other strains. In conclusion, T12 isolates have higher radioactivity than other isolates. The radioactive susceptibility of microorganisms differs depending on the species of each microorganism, the growth condition, and the environment. Gram positive bacteria, resistant spores, and some radioactive resistant microorganisms in Clostridium or Basillus genus exhibit high radioactivity resistance above 1 kGy D10. In contrast, food poisoning bacteria ( Salmonella , Staphylococcus , Listeria, Escherichia, etc.) that do not form spores exhibit a D10 value of about 0.5 kGy due to low radioactive resistance. The radioactive sterilization dose of 3 kGy, ^-6 level of germicidal activity.

[Experimental Example 2] Resistance to pesticides

Twenty - two kinds of pesticides were selected from 11 kinds of bactericides, 7 kinds of insecticides and 4 kinds of herbicides, which are used in farmland with high frequency. The resistance to pesticides was measured by inoculating a paper disc with 50 μl of the bacterial culture on the TSA plate containing the pesticide added at the concentration (Table 1) in the cultivation area, and then culturing at 30 ° C. for 2 days.

The four strains selected in Experimental Example 1 showed high growth in solid medium supplemented with 21 kinds of pesticides except two kinds of bactericides (Table 8). There was no resistance to the disinfectants Tebuconazole and Copper hydroxide, and it was found to be highly resistant to the other 21 pesticides. Tebuconazole belongs to the antibiotic-based triazole system, and it is known to be effective against not only fungi but also bacteria as a demethylase activity inhibitor. It has a wide range of pests to be applied and has excellent effect on resistant bacteria caused by other pharmacological agents. In particular, the Bacillus line is known to be inhibited by tebuconazole.

Resistance to pesticides on selected strains Pesticides (ppm) T19-8k T16-11k T12-11k Y11-6k Benomyl (250) +++ ++ +++ +++ Flusilazole (20) ++ ++ +++ +++ Tebuconazole (250) - - - - Oxadixyl (250) +++ +++ +++ +++ Edifenphos (300) +++ +++ +++ +++ Pencycuron (250) +++ +++ +++ +++ Azoxystrobin (100) ++ +++ +++ +++ Isoprothiorane (400) +++ +++ +++ +++ Iprobenphos (50) +++ +++ +++ +++ Procymidone (250) +++ +++ +++ +++ Chlorothalonil (250) +++ +++ +++ +++ Copper hydroxide (800) - - - - Chlorfenaphyr (50) ++ +++ +++ +++ Imidaclopride (50) +++ +++ +++ +++ Carbofuran (300) ++ +++ +++ +++ Chlorpyrifos (200) +++ ++ +++ +++ Fenpyroximate (25) +++ +++ +++ +++ Ethoprophos (250) +++ +++ +++ +++ Fenobucarb (500) +++ +++ +++ +++ Butachlor (50) +++ +++ +++ +++ Glyphosate (50) +++ +++ +++ +++ Alachlor (250) +++ ++ +++ +++ Pendimethalin (250) +++ +++ +++ +++

0 to 39%: - (no resistance), 40 to 59%: + (mild-), 60 to 79%: ++ (good-), 80%

[Example 1]

0.01 kg of Bacillus amyloliquefaciens KS-R01 (KCTC 13558BP), 4 kg of rice bran, 2 kg of molasses and 4 kg of sulfur sugar were mixed with water suitable for drinking water and weighed 100 kg The mixture was kept at 20 to 25 ° C. and aerated with 100 L / h of air for 2 hours at intervals of 4 hours. This procedure was repeated four times a day for 21 days to prepare a liquid fermentation product. The total amount of the liquid fermentation broth was 4.3 x 10 ⁷ cfu / g.

Separately, pre-milled 65 kg of rice bran and 35 kg of agricultural by-products were placed in an incubator with mixing function and mixed for 30 minutes. 30 kg of the liquid fermentation agent was added to the mixed raw materials to adjust the moisture concentration to 70%, and then the microorganism mixed germs according to the present invention was inoculated at 0.01% of the total weight of the culture raw materials. The inoculated culture material was agitated at an internal temperature of the incubator of 80 to 85 ° C at 30 to 80 rpm / min while the ultra-high temperature culture was continued for 4 hours to prepare a complex microorganism preparation of the present invention.

One ton of water was added to 5 kg of the complex microorganism preparation, 10 kg of molasses was added thereto, and the mixture was incubated with aeration at 25 ° C for 72 hours to obtain a stock solution. Twenty times the weight of water was added to the stock solution, and 0.2 wt% of molasses was added thereto to obtain an activity of 20 times.

[Example 2]

A microorganism preparation was prepared by the same procedure as in Example 1 except that T12-11k ( Bacillus siamensis KS-R02 (KCTC 13559BP)) was used as the microorganism strain.

[Example 3]

A microorganism preparation was prepared by the same procedure as in Example 1, except that T19-8k ( Bacillus velezensis KS-R03 (KCTC 13560BP)) was used as the microorganism strain.

[Example 4]

A microorganism preparation was prepared by the same procedure as in Example 1 except that Y11-6k ( Bacillus tequilensis KS-R04 (KCTC 13561BP)) was used as the microorganism strain.

[Example 5]

Bacillus amyloliquefaciens KS-R01 (KCTC 13558BP), Bacillus siamensis KS-R02 (KCTC 13559BP), Bacillus velezensis KS-R03 (KCTC 13558BP ), Bacillus amyloliquefaciens KC- ( Bacillus tequilensis KS-13560BP), and Bacillus tequilensis KS-R04 (KCTC 13561BP) were used in the same manner as in Example 1.

[Experimental Example 3]

Experimental Method

In order to optimize the performance of the microbial agent, 20 times the volume of the original microbial agent solution was injected at 1/50 of the volume of the soil microbial agent.

The soil samples were taken from the granite - based Danyang area, where the amount of radon produced exceeds the standard value. The average value of naturally occurring radon in the soil collected in this area is 180 becquerel (Bq / ㎥) Respectively.

Experiment result

Results of radon measurement division Radon level (Bq / ㎥) 0 60 minutes 120 minutes 180 minutes Example 1 182 166 125 86 Example 2 179 165 122 82 Example 3 183 164 123 80 Example 4 184 166 120 79 Example 5 183 142 108 58 Comparative Example 182 179 172 168

Table 9 above shows the result of continuously measuring the radon level in the sample soil during the experiment using the radon meter at intervals of 60 minutes.

From the above experimental results, it can be confirmed that the microbial agent according to the present invention significantly reduces the amount of radon produced, which is expected to be very effective for preventing or restoring environmental pollution in a radioactive contaminated site.

[Experimental Example 4]

The results of measurement of residual pesticide component content after 14 days from the pesticide treatment in Experimental Example 2 are as shown in Table 10 below, and it is seen that the pesticide component is removed by about 50% .

Pesticide removal rate (after 14 days) Pesticides (ppm) T19-8k T16-11k T12-11k Y11-6k Benomyl (250) 53 68 81 55 Flusilazole (20) 2 2 3 3 Oxadixyl (250) 60 62 57 41 Edifenphos (300) 66 54 72 69 Pencycuron (250) 103 111 125 98 Azoxystrobin (100) 39 48 44 37 Isoprothiorane (400) 129 154 201 133 Iprobenphos (50) 18 20 16 11 Procymidone (250) 92 88 100 76 Chlorothalonil (250) 80 59 115 66 Chlorfenaphyr (50) 26 13 17 12 Imidaclopride (50) 23 11 18 27 Carbofuran (300) 53 61 63 59 Chlorpyrifos (200) 109 112 89 90 Fenpyroximate (25) 10 8 9 11 Ethoprophos (250) 88 93 91 101 Fenobucarb (500) 216 198 173 204 Butachlor (50) 18 13 12 21 Glyphosate (50) 14 11 16 22 Alachlor (250) 67 84 75 74 Pendimethalin (250) 109 82 123 94

From the above experimental results, it can be confirmed that the microbial agent according to the present invention significantly reduces the content of the pesticide component, and thus it is expected to be highly effective in preventing or restoring environmental pollution in the area contaminated with the pesticide ingredient.

Although the preferred embodiments of the present invention have been disclosed for illustrative purposes, those skilled in the art will appreciate that various modifications, additions and substitutions are possible, without departing from the scope and spirit of the invention as disclosed in the accompanying claims. It will be understood that the present invention can be changed.

Korea Biotechnology Research Institute KCTC13558BP 20180621 Korea Biotechnology Research Institute KCTC13559BP 20180621 Korea Biotechnology Research Institute KCTC13560BP 20180621 Korea Biotechnology Research Institute KCTC13561BP 20180621

<110> GEON-NONG CO., LTD. <120> BIOLOGICAL MICROBIAL TREATING AGENT FOR REMOVING RADIOACTIVE          MATERIAL <130> SP1008 <160> 4 <170> KoPatentin 3.0 <210> 1 <211> 1473 <212> DNA <213> Unknown <220> <223> Bacillus amyloliquefaciens <400> 1 gacgaacgct ggcggcgtgc ctaatacatg caagtcgagc ggacagatgg gagcttgctc 60 cctgatgtta gcggcggacg ggtgagtaac acgtgggtaa cctgcctgta agactgggat 120 aactccggga aaccggggct aataccggat gcttgtttga accgcatggt tcagacataa 180 aaggtggctt cggctaccac ttacagatgg acccgcggcg cattagctag ttggtgaggt 240 aacggctcac caaggcgacg atgcgtagcc gacctgagag ggtgatcggc cacactggga 300 ctgagacacg gcccagactc ctacgggagg cagcagtagg gaatcttccg caatggacga 360 aagtctgacg gagcaacgcc gcgtgagtga tgaaggtttt cggatcgtaa agctctgttg 420 ttagggaaga acaagtgccg ttcaaatagg gcggcacctt gacggtacct aaccagaaag 480 ccacggctaa ctacgtgcca gcagccgcgg taatacgtag gtggcaagcg ttgtccggaa 540 ttattgggcg taaagggctc gcaggcggtt tcttaagtct gatgtgaaag cccccggctc 600 aaccggggag ggtcattgga aactggggaa cttgagtgca gaagaggaga gtggaattcc 660 acgtgtagcg gtgaaatgcg tagagatgtg gaggaacacc agtggcgaag gcgactctct 720 ggtctgtaac tgacgctgag gagcgaaagc gtggggagcg aacaggatta gataccctgg 780 tagtccacgc cgtaaacgat gagtgctaag tgttaggggg tttccgcccc ttagtgctgc 840 agctaacgca ttaagcactc cgcctgggga gtacggtcgc aagactgaaa ctcaaaggaa 900 ttgacggggg cccgcacaag cggtggagca tgtggtttaa ttcgaagcaa cgcgaagaac 960 cttaccaggt cttgacatcc tctgacaatc ctagagatag gacgtcccct tcgggggcag 1020 agtgacaggt ggtgcatggt tgtcgtcagc tcgtgtcgtg agatgttggg gttaagtccc 1080 gcaacgagcg caacccttga tcttagttgc cagcattcag ttgggcactc taaggtgact 1140 gccggtgaca aaccggagga aggtggggat gacgtcaaat catcatgccc cttatgacct 1200 gggctacaca cgtgctacaa tgggcagaac aaagggcagc gaaaccgcga ggttaagcca 1260 atcccacaaa tctgttctca gttcggatcg cagtctgcaa ctcgactgcg tgaagctgga 1320 atcgctagta atcgcggatc agcatgccgc ggtgaatacg ttcccgggcc ttgtacacac 1380 cgcccgtcac accacgagag tttgtaacac ccgaagtcgg tgaggtaacc tttttggagc 1440 cagccgccga aggtggga gatgattggg gtg 1473 <210> 2 <211> 1472 <212> DNA <213> Unknown <220> <223> bacillus siamensis <400> 2 gacgaacgct ggcggcgtgc ctaatacatg caagtcgagc ggacagatgg gagcttgctc 60 cctgatgtta gcggcggacg ggtgagtaac acgtgggtaa cctgcctgta agactgggat 120 aactccggga aaccggggct aataccggat ggttgtctga accgcatggt tcagacataa 180 aaggtggctt cggctaccac ttacagatgg acccgcggcg cattagctag ttggtgaggt 240 aacggctcac caaggcgacg atgcgtagcc gacctgagag ggtgatcggc cacactggga 300 ctgagacacg gcccagactc ctacgggagg cagcagtagg gaatcttccg caatggacga 360 aagtctgacg gagcaacgcc gcgtgagtga tgaaggtttt cggatcgtaa agctctgttg 420 ttagggaaga acaagtgccg ttcaaatagg gcggcacctt gacggtacct aaccagaaag 480 ccacggctaa ctacgtgcca gcagccgcgg taatacgtag gtggcaagcg ttgtccggaa 540 ttattgggcg taaagggctc gcaggcggtt tcttaagtct gatgtgaaag cccccggctc 600 aaccggggag ggtcattgga aactggggaa cttgagtgca gaagaggaga gtggaattcc 660 acgtgtagcg gtgaaatgcg tagagatgtg gaggaacacc agtggcgaag gcgactctct 720 ggtctgtaac tgacgctgag gagcgaaagc gtggggagcg aacaggatta gataccctgg 780 tagtccacgc cgtaaacgat gagtgctaag tgttaggggg tttccgcccc ttagtgctgc 840 agctaacgca ttaagcactc cgcctgggga gtacggtcgc aagactgaaa ctcaaaggaa 900 ttgacggggg cccgcacaag cggtggagca tgtggtttaa ttcgaagcaa cgcgaagaac 960 cttaccaggt cttgacatcc tctgacaatc ctagagatag gacgtcccct tcgggggcag 1020 agtgacaggt ggtgcatggt tgtcgtcagc tcgtgtcgtg agatgttggg ttaagtcccg 1080 caacgagcgc aacccttgat cttagttgcc agcattcagt tgggcactct aaggtgactg 1140 ccggtgacaa accggaggaa ggtggggatg acgtcaaatc atcatgcccc ttatgacctg 1200 ggctacacac gtgctacaat ggacagaaca aagggcagcg aaaccgcgag gttaagccaa 1260 tcccacaaat ctgttctcag ttcggatcgc agtctgcaac tcgactgcgt gaagctggaa 1320 tcgctagtaa tcgcggatca gcatgccgcg gtgaatacgt tcccgggcct tgtacacacc 1380 gcccgtcaca ccacgagagt ttgtaacacc cgaagtcggt gaggtaacct ttatggagcc 1440 agccgccgaa ggtgggacag atgattgggg tg 1472 <210> 3 <211> 1472 <212> DNA <213> Unknown <220> <223> Bacillus velezensis <400> 3 gacgaacgct ggcggcgtgc ctaatacatg caagtcgagc ggacagatgg gagcttgctc 60 cctgatgtta gcggcggacg ggtgagtaac acgtgggtaa cctgcctgta agactgggat 120 aactccggga aaccggggct aataccggat ggttgtttga accgcatggt tcagacataa 180 aaggtggctt cggctaccac ttacagatgg acccgcggcg cattagctag ttggtgaggt 240 aacggctcac caaggcgacg atgcgtagcc gacctgagag ggtgatcggc cacactggga 300 ctgagacacg gcccagactc ctacgggagg cagcagtagg gaatcttccg caatggacga 360 aagtctgacg gagcaacgcc gcgtgagtga tgaaggtttt cggatcgtaa agctctgttg 420 ttagggaaga acaagtgccg ttcaaatagg gcggcacctt gacggtacct aaccagaaag 480 ccacggctaa ctacgtgcca gcagccgcgg taatacgtag gtggcaagcg ttgtccggaa 540 ttattgggcg taaagggctc gcaggcggtt tcttaagtct gatgtgaaag cccccggctc 600 aaccggggag ggtcattgga aactggggaa cttgagtgca gaagaggaga gtggaattcc 660 acgtgtagcg gtgaaatgcg tagagatgtg gaggaacacc agtggcgaag gcgactctct 720 ggtctgtaac tgacgctgag gagcgaaagc gtggggagcg aacaggatta gataccctgg 780 tagtccacgc cgtaaacgat gagtgctaag tgttaggggg tttccgcccc ttagtgctgc 840 agctaacgca ttaagcactc cgcctgggga gtacggtcgc aagactgaaa ctcaaaggaa 900 ttgacggggg cccgcacaag cggtggagca tgtggtttaa ttcgaagcaa cgcgaagaac 960 cttaccaggt cttgacatcc tctgacaatc ctagagatag gacgtcccct tcgggggcag 1020 agtgacaggt ggtgcatggt tgtcgtcagc tcgtgtcgtg agatgttggg ttaagtcccg 1080 caacgagcgc aacccttgat cttagttgcc agcattcagt tgggcactct aaggtgactg 1140 ccggtgacaa accggaggaa ggtggggatg acgtcaaatc atcatgcccc ttatgacctg 1200 ggctacacac gtgctacaat ggacagaaca aagggcagcg aaaccgcgag gttaagccaa 1260 tcccacaaat ctgttctcag ttcggatcgc agtctgcaac tcgactgcgt gaagctggaa 1320 tcgctagtaa tcgcggatca gcatgccgcg gtgaatacgt tcccgggcct tgtacacacc 1380 gcccgtcaca ccacgagagt ttgtaacacc cgaagtcggt gaggtaacct tttaggagcc 1440 agccgccgaa ggtgggacag atgattgggg tg 1472 <210> 4 <211> 1472 <212> DNA <213> Unknown <220> <223> Bacillus tequilensis <400> 4 gacgaacgct ggcggcgtgc ctaatacatg caagtcgagc ggacagatgg gagcttgctc 60 cctgatgtta gcggcggacg ggtgagtaac acgtgggtaa cctgcctgta agactgggat 120 aactccggga aaccggggct aataccggat ggttgtttga accgcatggt tcaaacataa 180 aaggtggctt cggctaccac ttacagatgg acccgcggcg cattagctag ttggtgaggt 240 aacggctcac caaggcaacg atgcgtagcc gacctgagag ggtgatcggc cacactggga 300 ctgagacacg gcccagactc ctacgggagg cagcagtagg gaatcttccg caatggacga 360 aagtctgacg gagcaacgcc gcgtgagtga tgaaggtttt cggatcgtaa agctctgttg 420 ttagggaaga acaagtaccg ttcgaatagg gcggtacctt gacggtacct aaccagaaag 480 ccacggctaa ctacgtgcca gcagccgcgg taatacgtag gtggcaagcg ttgtccggaa 540 ttattgggcg taaagggctc gcaggcggtt tcttaagtct gatgtgaaag cccccggctc 600 aaccggggag ggtcattgga aactggggaa cttgagtgca gaagaggaga gtggaattcc 660 acgtgtagcg gtgaaatgcg tagagatgtg gaggaacacc agtggcgaag gcgactctct 720 ggtctgtaac tgacgctgag gagcgaaagc gtggggagcg aacaggatta gataccctgg 780 tagtccacgc cgtaaacgat gagtgctaag tgttaggggg tttccgcccc ttagtgctgc 840 agctaacgca ttaagcactc cgcctgggga gtacggtcgc aagactgaaa ctcaaaggaa 900 ttgacggggg cccgcacaag cggtggagca tgtggtttaa ttcgaagcaa cgcgaagaac 960 cttaccaggt cttgacatcc tctgacaatc ctagagatag gacgtcccct tcgggggcag 1020 agtgacaggt ggtgcatggt tgtcgtcagc tcgtgtcgtg agatgttggg ttaagtcccg 1080 caacgagcgc aacccttgat cttagttgcc agcattcagt tgggcactct aaggtgactg 1140 ccggtgacaa accggaggaa ggtggggatg acgtcaaatc atcatgcccc ttatgacctg 1200 ggctacacac gtgctacaat ggacagaaca aagggcagcg aaaccgcgag gttaagccaa 1260 tcccacaaat ctgttctcag ttcggatcgc agtctgcaac tcgactgcgt gaagctggaa 1320 tcgctagtaa tcgcggatca gcatgccgcg gtgaatacgt tcccgggcct tgtacacacc 1380 gcccgtcaca ccacgagagt ttgtaacacc cgaagtcggt gaggtaacct tttaggagcc 1440 agccgccgaa ggtgggacag atgattgggg tg 1472

Claims (3)

Bacillus amyloliquefaciens KS-R01 (KCTC 13558BP), Bacillus siamensis KS-R02 (KCTC 13559BP), Bacillus velezensis KS-R03 (KCTC 13560BP), Bacillus amyloliquefaciens KS- And Bacillus tequilensis KS-R04 (KCTC 13561BP), and include Benomyl, Flusilazole, Oxadixyl, Edifenphos, Pencycuron ), Azoxystrobin, isoprothiorane, Iprobenphos, Procymidone, Chlorothalonil, Chlorfenaphyr, Imidaclopride, But are not limited to, Carbofuran, Chlorpyrifos, Fenpyroximate, Ethoprophos, Fenobucarb, Butachlor, Glyphosate, Allachlor Alachlor), and Pendimethali n). &lt; / RTI &gt; delete Water or soil is treated with the microorganism treating agent of claim 1 to prepare a solution containing at least one of Benomyl, Flusilazole, Oxadixyl, Edifenphos, Pencycuron, Azoxystrobin ), Isoprothiolane, Iprobenphos, Procymidone, Chlorothalonil, Chlorfenaphyr, Imidaclopride, Carbofuran, Chlorophyllone, But are not limited to, Chlorpyrifos, Fenpyroximate, Ethoprophos, Fenobucarb, Butachlor, Glyphosate, Alachlor, and Pendimethalin (Pendimethalin). &Lt; / RTI &gt;

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