KR101626801B1 - Beauveria bassiana having insect pathogenicityand agent for prevention of rice vermin using the same - Google Patents

Abstract

The present invention relates to a strain of B. bassiana ERL836, an insect pathogenic fungus, and a preparation for controlling water pests using the same. The water pest control agent according to the present invention is produced using Beauveria bassiana strain ERL836, which is an insect pathogenic fungus, so that it is easy to handle, economical, harmless to the human body, and has an effect of controlling water pests in an environmentally friendly manner .

Description

FIELD OF THE INVENTION [0001] The present invention relates to an insect pathogenic bovaria bacillus and an insect pest control agent,

The present invention relates to a bacterium bacterium Bacteriophage ERL836, which is an insect pathogenic fungus, and a preparation for controlling water pests using the same.

Leafhopper, brownfly, and rice weevil are widely distributed in tropical, subtropical, and rice cultivation areas including Korea, and they directly affect physical damage by feeding rice, and they inhibit growth inhibitory viruses such as rice blast foliage blight, It acts as an infectious insect, and is one of the major water pests that causes massive losses in rice yields.

The emergence of organic synthetic insecticides after World War II marked a milestone turning point in two aspects: increasing agricultural crops by agricultural pest control and improving human health by sanitary pest control. Insecticides used for pest control in Korea are increasing in usage every year, of which organic phosphorus and carbamate accounts for about 90% of total usage.

 However, due to decades of continuous use and abuse of these organic synthetic pesticides, various adverse effects such as the appearance of resistant pests, lethal toxicity, biological control disturbance by natural enemies, residual poison and environmental pollution have been caused. Recently, The use of synthetic insecticides is becoming increasingly restricted as the interest in the adverse effects and environmental effects of the insecticides is increasing. Biological control has been actively studied as part of the development of next-generation pesticides that can replace synthetic pesticides, but biological control is an obstacle to commercialization because it is difficult to handle microorganisms, natural enemies or natural extracts

Therefore, there is a desperate need to develop a new water pest control agent that is harmless to the human body, eco-friendly, easy to handle, and easy to manufacture, and which can be used commercially.

KR 10-2010-0111297

The present inventors newly discovered a strain of ERL836 of Beauveria bassiana and found that the strain has an insect pathogenicity and excellent spore forming ability, and when it is used in water pest control The present invention has been accomplished on the basis of the findings that it is possible to easily prepare pharmaceutical preparations and compositions with excellent control effect against water pests.

Accordingly, the present invention provides a strain of B. bassiana ERL836, an insect pathogenic fungus, and a preparation for controlling water pests using the same.

In order to achieve the above object,

The present invention provides a strain of beauveria bassiana ERL836 (KCCM11506P) having insect pathogenicity.

In addition, the present invention provides a microorganism preparation for water pest control, which comprises a strain of Beauveria bassiana ERL836 or a spore thereof.

In addition, the present invention provides a method for producing a bacterium, comprising culturing a bovaria bacillus strain ERL836 in a solid culture medium; And harvesting the culture. The present invention also provides a method for producing a microorganism preparation for controlling water pests.

The present invention also provides a water pest control method comprising the step of spraying a microorganism preparation for water pest control comprising a strain of Beauveria bassiana ERL836 or a spore thereof on a gravestone.

Hereinafter, the present invention will be described in detail.

The present invention provides a strain of beauveria bassiana ERL836 (KCCM11506P) having insect pathogenicity.

In addition, the present invention provides a composition for water pest control comprising a strain of Beauveria bassiana ERL836 or a spore thereof.

In addition, the present invention provides a microorganism preparation for water pest control, which comprises a strain of Beauveria bassiana ERL836 or a spore thereof.

" Beauveria bassiana ERL836 strain" of the present invention is a bacterium belonging to the genus Bacterium which shows excellent insecticidal effect against water insect pests such as, for example, spermatozoa, brownflies, white beetles, After isolating the strain of Beriabassiana through a biochemical assay, the isolated strain was morphologically identified, and the genomic DNA sequence was analyzed to show about 98% homology with the previously known Biberia basiana strain It was named as ERL836, and deposited on Jan. 22, 2014 with the Korean Society for Microbiological Protection (KCCM11506P).

The B. bovisiana ERL836 strain has a morphological characteristic of forming white clusters and spores (see FIG. 1). The ERL836 strain has particularly good spore forming ability, and the spore of the ERL836 strain is superior to the known strain of bovaria barbiana or GHA in spore stability.

The insect pathogenicity refers to the ability of a pathogen to infect a target insect (host insect) to cause disease. In the present invention, the bacterium ERL836 is infected with a water pest, It means to cause illness.

The insect pathogenicity may be an insecticidal activity. Specifically, the bovaria bacillus strain ERL836 or the spore of the strain is adhered to the outer shell of the host insect and germinates, and the chitinase, the protease, the lipase and / or the esterase And penetrates into the cuticle layer of the insect and propagates in the body of the insect so that small molecular weight produces a toxic protein such as toxic substance or glycoprotein to block the immune function of the insect and to kill the host insect .

The bovaria bacillus strain ERL836, which has excellent spore forming ability and has a remarkably excellent insecticidal activity against water insects, has been identified and can be usefully used for the control of water pests.

The insect may be a insect pest, and the insect pest may include a horticultural insect, a water insect, and the like.

The insect may preferably be a water pest, and the water pest is also referred to as a rice pest, and includes all the worms that harm rice. The water pest may preferably be any one selected from the group consisting of a liana, a brownfly, a white lizard, a rice weevil, a rice weevil, a rice leaf weevil and a combination thereof, more preferably a rice weevil, It can be one.

The spore of the bovaria bacillus strain ERL836 can be produced by culturing inoculum of the bovaria bacillus strain ERL836 in a solid culture medium. The bovaria bacillus ERL836 strain of the present invention can exhibit spore productivity of about 4 to 6 times and spore storage stability of about 2 to 3 times as compared with the conventional bovaria barbiana GHA strain.

The microorganism preparation for water-pest control may be provided as a granular preparation, a liquid preparation or a compost. The microorganism preparation for water-pest control has an excellent insecticidal effect when sprayed on water pests, and in particular, the microorganism control agent for controlling water pests has a remarkable insecticidal effect compared to the case of spraying with the liquid formulation of the known strain of bovaria bacillus or GHA strain And showed about 2 times better insecticidal effect.

The microorganism preparation may be preferably a granular preparation, and when the preparation is a granular preparation, it is easy to handle, and granular preparations can be easily produced by using the excellent sporulation ability of bovisia barbiana strain ERL836, The production efficiency of the control agent can be improved.

In addition, the present invention provides a method for producing a bacterium, comprising culturing a bovaria bacillus strain ERL836 in a solid culture medium; And harvesting the culture. The present invention also provides a method for producing a microorganism preparation for water pest control.

The solid culture medium preferably contains any one selected from the group consisting of millet, white rice, wheat bran, and combinations thereof, and more preferably comprises a millet.

The method for preparing a microorganism preparation for water-pest control may further comprise preparing a solid culture medium by mixing any one selected from organic acids with millet, white rice, wheat bran, millet, millet, and combinations thereof.

In addition, the step of preparing the solid culture medium may further comprise immersing any one selected from the group consisting of millet, white rice, wheat bran mixed with organic acid, and combinations thereof at 80 to 100 캜 for 0.5 to 1.5 hours in water.

In the present invention, the solid culture medium mixed with the organic acid is subjected to sterilization treatment for 20 minutes to 40 minutes at a temperature of 110 to 130 DEG C while water is absorbed in the water bath for 0.5 to 1.5 hours while maintaining the temperature at 80 to 100 DEG C To prepare a solid culture medium.

The organic acid is not particularly limited as long as it can be used as a carbon source. More specifically, it is preferably any one selected from citric acid, propionic acid, butyric acid, lactic acid, succinic acid, clonic acid, valeric acid, caproic acid and combinations thereof, more preferably citric acid.

The organic acid and the solid culture medium may be mixed by adding an organic acid to the solid culture medium and mixing the organic acid.

The culturing vessel of the strain may be a culturing vessel commonly used in the technical field of the present invention, and is not limited thereto. For example, a filter for supplying oxygen may be attached to a polyvinyl bag to be used for culturing.

The cultivation of the bovis larva and ERL836 strain is preferably carried out at 20 to 30 DEG C for 6 to 8 days, more preferably at 25 DEG C for 7 days.

The inoculum to be used for the solid culture medium may be obtained by subculturing the bovisia barbica strain ERL836, and the cultivation of the ERL836 strain may be performed according to a conventional subculture method. The medium required for the subculture is not limited to that kind.

As an example of the subculture medium, a liquid medium containing PDB (Potato Dextrose Broth) medium, rice bran or wheat bran can be used, but it is not limited thereto. The subculture medium may also include corn flour, rice bran or cornsteep powder, and the like.

The present invention also provides a water pest control method comprising the step of spraying a beauberia bassiana strain ERL836 or a spore thereof onto a grave plate.

The present invention also provides a water pest control method comprising the step of spraying a water pest control composition or a pest control microorganism preparation containing a spore of a strain of Beauveria bassiana ERL836 or a spore thereof on a tombstone .

It is preferred that the above-mentioned Beauveria bassiana ERL836 strain or a spore thereof, or a composition for controlling water pests or a microorganism preparation for controlling a water pest therefrom is sprayed on the tombstone before the transition, More preferably from 40 to 60 g / tray.

When the bovisia barbiana strain ERL836, its spore, composition or microorganism preparation is sprayed onto the gravestone, a two times higher control rate than that of the conventional bovaria barbiana or GHA strain is observed There is an effect indicating.

The water pest control agent according to the present invention is produced using Beauveria bassiana ERL836 strain, which is an insect pathogenic fungus, so that it is easy to handle, harmless to human body, and has an effect of controlling water pest environmentally.

Brief Description of the Drawings Fig. 1 is an optical micrograph (400X) of an Escherichia coli strain ERL836 cultured in Sabouraud dextrose agar (SDA) medium in a dark room at 25 캜 for 7 days.
FIG. 2 is a diagram showing DNA sequencing results of the bovaria bacillus and ERL836 strain.
FIG. 3 is a diagram showing microorganism preparation images of (a) length, (b) polyvinyl bag for solid culture, and (c) bovaria bacillus strain ERL836.
Fig. 4 is a diagram showing that the microorganism preparation of bovaria bacillus or ERL836 strain is sprayed on a pre-migration plate and then watered.
FIG. 5 is a graph showing the effect of microbial preparations of bovaria bacillus, ERL836 strain and GHA strain on the control of rice weevil.
FIG. 6 is a graph showing the effect of microbial preparations of bovaria bacillus, ERL836 strain and GHA strain on control of offspring.

Hereinafter, preferred embodiments of the present invention will be described in order to facilitate understanding of the present invention. However, the following examples are provided only for the purpose of easier understanding of the present invention, and the present invention is not limited by the examples.

Example 1 Isolation and Identification of Strain

1-1. Isolation of strain

Beauveria bassiana ERL836 strain is a bacteriocin that has excellent insecticidal effect against water insect pests, such as, for example, lycopene, brownfly, white lily, and rice weevil, among the fungi isolated from natural soil. Respectively.

1-2. Growth and morphological identification

FIG. 1 shows an optical microscope (400 times) image of the bovaria bacillus ERL836 strain cultured in Sabouraud Dextrose Agar (SDA) for 7 days in a dark room at 25 ° C.

As shown in Fig. 1, the formation of white mycelia, spores, and spores, typical features of the bovaria strain, was well observed.

1-3. Sequencing

After amplification of PCR (polymerase chain reaction, polymerase chain reaction) by ITS1 forward primer and ITS4 reverse primer, The results of DNA sequencing of E. bovisana ERL836 strain are shown in FIG.

Example 2. Confirmation of spore production ability

1 g of the microorganism preparation of strain ERL836 was diluted with 10 ml of 0.3% silwet, and the number of spores was counted using a hamacytometer to measure the spore productivity. The porosity coefficients of the bovisia basiana ERL836 strain are shown in Table 1.

Strain name Porosity B. bassiana ERL836 2.76 x 10 ⁸ conidia / g B. bassiana GHA 4.41 x 10 ⁷ conidia / g

Conduct 3. Confirm spore storage stability

The bovaria barbiana strain ERL836 and the GHA strain were cultured in a millet medium and stored at room temperature for 16 months to investigate the germination rate of the spores. Table 2 shows storage stability of spores of bovaria baicana strain ERL836 at room temperature.

Strain name Germination rate after 16 months at room temperature (%) B. bassiana ERL836 90.7 ± 2.5% B. bassiana GHA 58.1 ± 3.7%

As shown in Table 2, the ERL836 strain was found to have about twice the spore storage stability as compared with the GHA strain.

Example 4: Preparation of a microorganism preparation for water pest control

100 g of millet was placed in a polyvinyl bag for culture, and then 0.16 ml of 50% citric acid was further added. After that, the length was incubated at 90 DEG C for 1 hour by using a water bath, so that moisture was sufficiently absorbed in the long field. A paper cup and a filter for supplying oxygen using sterilized gauze were attached to the entrance of the polyvinyl bag and sterilized by heating at 121 ° C for 30 minutes. The microbial preparation of ERL836 strain was prepared by cooling the sterilized capillary to room temperature, inoculating 1 ml of the Bovrieria basiana ERL836 strain cultured in PDB medium for 3 days at room temperature, shaking it, and culturing at 25 ° C for 7 days. Respectively.

Images of the microorganism preparations of the millet, the solid culture polyvinyl bag and the bovaria barbiana strain ERL836 are shown in Figs. 3 (a) to 3 (c), respectively.

Example 5: Examination of insecticidal effect of microorganism preparation on rice weevil

The bacterium (50 ml / tray) was sprayed with a microbial formulation of boveriac basiana ERL836 at a dose of 50 g / tray (58 x 28 x 2 cm ³ ) to the basal plate three days before transplantation (Fig. 4). In contrast, the liquid formulation of GHA strain was diluted 500-fold and then treated with 1 L / tray. After 3 days (10 x 5 m ² ), the damage rate of rice was investigated on the 14th day of transplantation. After 38 days, the root of rice was extracted to investigate the density of rice weevil larvae present in the rhizosphere. The control effect of the rice weevil was investigated in Fig.

As shown in FIG. 5, the control effect of the microorganism preparation of the ERL836 strain was two times higher than that of the conventional liquid formulation of the GHA strain.

Table 3 shows the weevil larva density according to the spraying treatment of the microorganism preparation of the bovaria barbiana strain ERL836.

Strain name Larvae density (number of grains / 5 m ² ) B. bassiana ERL836 37.1 ± 12.8 B. bassiana GHA 68.1 ± 11.1 No treatment 97.7 ± 15.1

As shown in Table 3, when the microorganism preparation of strain ERL836 was treated, the lowest density larvae were examined.

Example 6 Identification of the Insecticidal Effect of the Microorganism Preparation on Leptospira

The microbial preparation of bovaria bacillus strain ERL836 was sprayed on the base plate three days before transplantation in a dose of 50 g / tray (58 x 28 x 2 cm ³ ), and then subjected to irrigation (50 ml / tray). (See FIG. 4). In contrast, the liquid formulation of GHA strain was diluted 500-fold and treated with 1 L / tray. The growth density of the larvae was examined at 3, 7, and 14 days after transplantation to 3 repetitions (10 x 5 m ² ) in the young seedling packaging conditions, and the results are shown in Fig.

As shown in FIG. 6, the microbial treatment of ERL836 showed a high control effect on the 14th day, and the control effect of the GHA strain on microbial treatment was as low as 35%.

Korea Microorganism Conservation Center (overseas) KCCM11506 20140122

Claims (9)

Beauveria bassiana strain ERL836 (KCCM11506P) with insect pathogenicity to rice pests. The method according to claim 1,
The rice pests aemyeolgu, BPH, huindeung myeolgu, byeomul weevil, rice stem gulpari, byeoip gulpari and would in any one of the rice pest selected from a combination thereof, bonded Beria Bridge Ana (Beauveria bassiana) ERL836 strain (KCCM11506P). A microbial agent for controlling rice insect pests of a rice weevil, a ladybug or a combination thereof, comprising a strain of the ERL836 strain (KCCM11506P) or a spore of the beaververia bassiana having an insect pathogenicity to a rice pest. (1) culturing a boveriac basiana strain ERL836 (KCCM11506P) having an insect pathogenicity in a rice insect pest in a solid culture medium;
(2) harvesting the culture; And
(3) spraying the harvested culture onto the gravestone. 5. The method of claim 4,
Wherein the solid culture medium comprises any one selected from the group consisting of millet, white rice, wheat bran, millet, millet, and combinations thereof. 5. The method of claim 4,
Wherein the solid culture medium is prepared by mixing any one selected from the group consisting of organic acid, millet, white rice, wheat bran, millet, millet, and combinations thereof. The method according to claim 6,
Wherein the solid culture medium is prepared by immersing any one selected from the group consisting of millet, white rice, wheat bran, millet, millet, and combinations thereof with organic acid at 80 to 100 DEG C for 0.5 to 1.5 hours in water , A method for controlling rice insect pests. The method according to claim 6,
Wherein the organic acid is any one selected from the group consisting of citric acid, propionic acid, butyric acid, lactic acid, succinic acid, clonic acid, valeric acid, caproic acid and combinations thereof. delete

Images