JP2002293708A - Bacillus popilliae, method for controlling insect of family scarabaeidae by using the same and controlling agent - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a microorganism comprising microbial cell, spore or sporangium exhibiting excellent controlling activity against insects of the family Scarabaeidae including cupreous chafer and oriental beetle, provide a method and an agent for controlling the insects of the family Scarabaeidae using the microorganism and provide a controlling agent exhibiting similar insecticidal effect independent of the species on the insects of the family Scarabaeidae, especially representative and serious harmful insects comprising cupreous chafer, oriental beetle, soy bean beetle and Japanese beetle and applicable over a wide application period. SOLUTION: The microorganism is Bacillus popilliae var. popilliae DIC-2001 (FERM P-18250) exhibiting insecticidal activity against oriental beetle and cupreous chafer and strong insecticidal action against oriental beetle. The method for controlling the insects of the family Scarabaeidae comprises the application of the microorganism to the insect. The invention further provides a controlling agent for the insects of the family Scarabaeidae containing the sporangium of the above microorganism.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a bacterial cell, a spore, or a sporangia having a spore and a parasporal body of a microorganism belonging to Bacillus populiae, and a method and an agent for controlling scarabid insects using the same. About. Microorganisms belonging to the Bacillus popiliae can induce emulsifying disease on larvae of the insects of the order Scarabaeidae, which are plant pests, and thus can be used as a control agent against Scarabaeidae.

[0002]

2. Description of the Related Art Conventionally, the insects of the family Scarabaeidae, in particular, scrophulariid beetles, dung beetle buoy, bean beetle, and beetle, have been serious plant pests in lawns, pastures, farms, orchards, and gardens. It is common to use pesticides that have been used. Since these larvae live in the ground, chemical pesticides sprayed from the ground are difficult to work and their habitats are difficult to identify, so in order to obtain a sufficient insecticidal effect, a large amount of insecticidal activity is required. Pesticides must be sprayed and penetrated into the ground. As a result, environmental pollution is likely to be caused, so that its control and control have been practically difficult. In particular, it was even more difficult to control and control large-sized dung beetles and scabies.

[0003] However, under the background of the times in which environmental problems are regarded as important, instead of chemical pesticides that are likely to adversely affect the natural environment and the human body, a highly safe biological control method that can contribute to environmental conservation has been desired. .

In view of the above, microbial pesticides using microorganisms that exhibit insecticidal properties against insects have been developed. For example, Bacillus thuringiensis, which has pathogenicity against lepidoptera and mosquito larvae, has been developed. A so-called BT agent using a living bacterium or an insecticidal component thereof as an agrochemical is known as a typical one.

It is known that a microorganism belonging to Bacillus popiliae isolated from a beetle (Popilliae japonica) larvae affected by an emulsifying disease induces an emulsifying disease to the beetle larvae. Microbial preparations using the microorganism are commercially available in the United States. However, these microorganisms have no effect on Douganebuibui, an important pest in Japan (Agriculturally useful microorganisms-their use and prospects-Kenji Umetani, Hajime Kato, p.236)
990).

As a microorganism belonging to the genus Bacillus popiliae that causes an emulsifying disease against Semaphora kogane and Douganebuui, Japanese Patent Application Laid-Open No. H11-332556, the insecticidal effect was not sufficient. further,
An industrial production method of the microorganism of Bacillus popirie which exhibits insecticidal activity to Scarabaeid insects has not been established,
The only production method is culture in the larva of an insect of the order Scarabaeidae, which is extremely low in productivity. Therefore, a microorganism having a higher insecticidal activity has been desired.

Bacteria belonging to the genus Bacillus having an insecticidal activity against the buddha buoy (Bacillus thuringiensis japonensis 141 strain, Japanese Patent Application Laid-Open No. 8-228783)
However, it is not a microorganism belonging to Bacillus populiae. In addition, these microorganisms have a feature that they are susceptible to ultraviolet rays, and it is difficult to maintain the control effect when formulated, and a more practical microorganism control agent is desired.

[0008] In addition, conventionally known bacteria belonging to Bacillus popiliae have a problem in that there is a great variation in their effects on representative and serious plant pests such as mamekogane, himekogane, samadaragane, and douganebuui. That is, although it has an extremely high insecticidal activity against bean pods and corn beetles, it has a low insecticidal activity against beetle buoys and semadara bean, and its control effect is selective. Therefore, when applied as a microbial pesticide, the amount of application must be appropriately adjusted after confirming the type of insects, which has been a burden on operators for handling.

[0009] Furthermore, the bacterium belonging to Bacillus popiliae, which is conventionally known, has a relatively narrow width of a period in which the control effect is effectively exerted on Semaphora kogane and Douganebuuibu, so-called "appropriate period of application". That is, the bacterium belonging to the Bacillus popirie exhibits relatively herbicidal activity against Semaphora kogane and Douganebuui during the application up to the second stage, but is applied to the stage after the second stage, particularly to the second to third stages. In this case, the control effect is extremely low. For this reason, on grass at golf courses where these insects breed, busy golf course management work and irregular weather have passed the effective application period of the pesticide, and sufficient control activity is not obtained, and the lawn landscape is not obtained. May be impaired.

[0010]

The problem to be solved by the present invention is to provide a microorganism exhibiting excellent control activity against Scarabaeidae insects, in particular, Scarabaeidae and Semaphoridae, and a method for controlling Scarabaeidae insects using the microorganism. And a control agent. In addition, to provide a control agent having a similar insecticidal effect to Scarabaeidae insects, particularly the typical and serious plant pests Douganebuui, Semadarakogane, Himekogane, and Mamekogane irrespective of the type, and furthermore, to provide a long-term suitable application period. It is in.

[0011]

Means for Solving the Problems The present inventors have made intensive studies to solve the above-mentioned problems, and as a result,
The present inventors have found a novel microorganism belonging to Bacillus popirie that has a stronger insecticidal activity than the microorganisms belonging to Bacillus popirie known to larvae of Semaphora beetle, and have completed the present invention based on this finding. Was.

That is, the present invention relates to Bacillus popirie popirie DI, which has an insecticidal activity on Semaphora kogane and Douganebuui, and has a strong insecticidal activity against Semaphora kogane.
C-2001 strain (bacillus popillia)
e var. popillae DIC-2001)
(FERM P-18250).

[0013] The present invention also provides a method for controlling Scarabaeidae insects, which comprises causing the microorganism to act on Scarabaeidae insects. The present invention also provides a control agent for Scarabaeidae insects containing the sporangia of the microorganism.

[0014]

DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will be described below in detail.

The Bacillus popirie popirie of the present invention
DIC-2001 strain (bacillus popill)
ia var. popillae DIC-200
1) is a microorganism belonging to Bacillus popirie, which is pesticidal to Semaphora kogane and Douganebuui. Bacillus popirie popirie DIC-
The 2001 strain is a strain isolated as described in Examples below from a larvae of Semaphora kogane affected by emulsification disease in Nirayama-machi, Taga-gun, Shizuoka Prefecture. The Bacillus popirie popirie DIC-2001 strain has been registered on March 9, 2001 by the Ministry of Economy, Trade and Industry, National Institute of Advanced Industrial Science and Technology, Biotechnology and Industrial Technology Research Institute, Patent Organism Depositary (1-3-1, Higashi 1-chome, Tsukuba, Ibaraki, Japan). ) Has been deposited under accession number FREM P-18250.

The Bacillus popirie popirie of the present invention
Table 1 shows the bacteriological properties of the DIC-2001 strain. As shown in the examples described later, the present microorganisms cause pathogenicity that induces emulsification disease in scarab beetles larvae, and sporanges obtained from larvae infected with emulsification disease are spores and spores having parasporal bodies. It was recognized that it belonged to Bacillus popiliae because of its formation. In addition, as shown in Examples described later, Bacillus popirie semadara strain, which is conventionally known to exhibit insecticidal activity only on Semaphora kogane and Douganebuuibu (JP-A-11-332556 FE)
RM P-18818) is a novel microorganism because it has a different insecticidal activity spectrum against Semaphora kogane, Douganebuui, Mamekogane and Himekogane, and exhibits stronger insecticidal activity against Semaedaragane and Douganebuui, especially Semadara beetle. did. Therefore, this strain was called Bacillus
It was named DIC-2001 strain.

By causing the microorganism of the present invention to act on Scarabaeidae insects, Scarabaeidae insects, especially Scutellariae squid and Semaphoridae can be controlled. Among them, it shows a particularly strong controlling activity against Scutellaria cricket. For example, the sporangia produced by the microorganism of the present invention is 150 g
~ 200%, and 200 ~ 500 for Semaphora kogane
% Insecticidal activity.

The action of the microorganism of the present invention on Scarabaeid insects is carried out by incorporating the cells of the microorganism of the present invention or sporangia, preferably sporangia, into the body of Scarabaeid insects, preferably larvae. .

Although the method of the present invention can be widely applied to insects of the order Scarabaeidae, it can be suitably applied particularly to stag beetles and Scutellariae. What can be taken into the Scarabaeid insect body may be a cell of the microorganism of the present invention or a sporangia, but a sporangium is preferred. These cells and sporangia may be obtained from larvae infected with emulsification disease or may be obtained by artificial culture.

The sporangia of the present invention can be prepared, for example, as follows. The microorganism of the present invention is incorporated into a larva of the order Scarabaeidae, preferably a larva of the scorpion beetle. Specifically, the microorganism of the present invention is sprayed on a breeding medium in which the larva is present, and taken up orally, or injected into a body fluid by injection. The larvae are preferably bred for 3 to 4 weeks, and the microorganism of the present invention is propagated in the larvae. The sporangia increased in the larval body can be obtained by, for example, incising or making a hole in the larva, collecting body fluid, and centrifuging or filtering the obtained body fluid.

In the method of the present invention, the sporangia may be used as it is, or, if necessary, water or a neutral buffer such as a phosphate buffer (pH = 7.5) or Tris-Hcl buffer. It may be washed with a liquid. In addition, spores are commonly used carriers for microbial preparations such as microbial pesticides,
The resulting microorganisms can be used as a microbial preparation by mixing with nutrient components. Further, by mixing Bacillus thuringiensis cells, spores or insecticidal components thereof with this composition, the composition may be used in combination with the microorganism of the present invention, and a synergistic effect on the scarab beetle larva is expected.

The microorganism, sporangia or controlling agent of the present invention comprises:
Lawns, orchards, and agriculture trying to control scarab beetles
1m soil on the ground^Two1 × 10 sporangia⁸From 1 ×
10 ¹⁴Pieces, preferably 1 × 10⁹From 1 × 10¹³And
Sprinkle so that it becomes. Then 1m as needed^Two
Sprinkle 1-2 L of water and / or plow the soil
You may mix. If using a suspension, add water or
Is 1 × 10⁶1 × 10 from pieces / L¹⁴Pieces
/ L, preferably 1 × 10⁹1 × 10 from pieces / L¹³Pieces/
It is preferable to use one suspended to be L.

The microorganism of the present invention, particularly the control agent containing a sporangia as an active ingredient, can be used for any kind of insects of the order Scarabaeidae, in particular, the typical and serious plant pests of the species Scarabaeidae, Semadarakogane, Himekogane and Mamekogane. Since they show the same insecticidal effect, they can be sprayed without confirming the type of Scarabaeidae insects during application, so that they are preferable as a controlling agent. For example, as shown in Example 3 below, the difference in insecticidal activity against different types of larvae with respect to a certain spore concentration is expressed as a ratio of 0.0
It tends to be within 3 and preferably within 0.03 to 0.01.

In addition, the controlling agent containing the microorganism of the present invention, particularly a sporangia as an active ingredient, exhibits a remarkably high controlling activity even after the second stage, especially against the second to third stage stag beetles. In addition, it exerts a remarkably high and excellent control activity on the other pups, such as the buoy buoy, the beetle, and the beetle. As described above, the pesticidal agent containing the microorganism of the present invention, particularly a sporangia as an active ingredient, has a long-term controlling activity, and therefore, a pesticidal agent for controlling Scarabaeidae insects having a wide suitable application period, particularly a pesticidal agent for controlling Scarabae beetle. Is useful as

[0025] In the present invention, "control of scarabidae insects" refers to control of scarabidae insects, particularly scarabidae insects, and prevention and improvement of plant insect damage by scarabidae larvae.

[0026]

EXAMPLES Hereinafter, the present invention will be described more specifically with reference to examples.

(Example 1) In April 2000, the outbreak of semaphoridae larvae occurred at Fuji Hakone Country Club located in Nirayama-machi, Tagata-gun, Shizuoka Prefecture. When the larvae were collected and kept in mulch, most of them died, and larvae whose body turned milky white were found in them. From this larva, a pathogenic strain causing an emulsifying disease was isolated by the following method.

A hole was formed in the larva that had become milky and died with a syringe needle, and a milky body fluid was collected from the blood body cavity. A precipitate fraction was collected from the body fluid by centrifugation. Microscopic observation confirmed that this fraction contained bacterial sporangia.

Distilled water was added to the precipitate fraction to adjust the sporangia concentration to 1 × 10 ⁶ / ml. (The number of sporangia was measured by a microscope using a hemocytometer.)

1 ml of the prepared solution was placed in an Eppendorf tube, heated at 70 ° C. for 20 minutes in a heat block, and then 50 microliters were applied to a plate medium having the following composition.

Medium Composition Polypeptone S 0.5% by weight (manufactured by Nippon Pharmaceutical Co., Ltd.) Yeast Extract 0.5% by weight (Oxoid) Dipotassium hydrogen phosphate 0.3% by weight (manufactured by Wako Pure Chemical Industries) Activated carbon 1% by weight (manufactured by Wako Pure Chemical Industries) Agar 2.0% by weight (manufactured by Wako Pure Chemical Industries)

The plates were cultured in an incubator at 30 ° C. under aerobic conditions for 10 days.

The colony formed on the medium was separated, and the operation of culturing was repeated in the same manner as described above to isolate a pure culture strain. Table 1 shows the results obtained by examining the bacteriological properties of the strains isolated as described above.

[0034]

[Table 1]

1): Vegetable cells were placed in a MYPGP medium at 30 ° C.
5 days culture 2): Vegetable cells are cultured in MYPGP medium field at 30 ° C. for 7 days 3): Vegetable cells are cultured in MYPGP medium field at 30 ° C. for 10 days

From the bacteriological properties shown in Table 1, it was confirmed that the strain belongs to the genus Bacillus popirie. Bergey's Manual of Determinative Bacteri
morphology, according to the mycological properties of Bacillus populiae, described in Phys.
m, a bacterium having a width of 0.5 to 0.8 m, having a motility and a growth temperature of 20 ° C. to 35 ° C., and is identical with Table 1. In addition, Bergey's Manual of Deterministic Bacteriology (Bergey's Manual
of Determinative Bacteriology) does not describe growth in the presence of 3% NaCl.

On the other hand, as shown in Example 3, the above strain exhibits an insecticidal effect on the larvae of Scarabaeidae insects such as Scutellaria bufo and Semaphoridae, and it is a novel microorganism having a higher insecticidal activity than conventionally known Bacillus sporiae. Bacillus popirie popirie DIC-200
One strain was named.

(Example 2) Production of sporangia of Bacillus popirie popirie DIC-2001 strain 20 g of humus was placed in a plastic cup having a diameter of 6 cm, and spores collected from larvae infected with emulsification disease were 1 ×.
About 10 ⁷ were sprayed. The second instar larvae of Douganebuui were placed in a cup and raised at 25 ° C. When the body of the larva became milky white, the body surface was sterilized with a 70% ethyl alcohol solution, a hole was made with an injection needle, and the body fluid was collected. 10 from the body fluid
Spores were collected by centrifugation at 00 rpm for 5 minutes.
The spores were washed twice with distilled water and adjusted to 1 × 10 ⁹ / ml to obtain a spore fraction.

In the same manner as described above, a sporangia fraction of Bacillus popirie semada was obtained.

Example 3 Insecticidal activity of microorganisms of the present invention against larvae of Scarabaeidae Insect humus was placed in 60 plastic cups having a diameter of 6 cm.
Then, the spore fraction prepared in Example 2 was sprayed so that the number of spores of Bacillus populus sp. DIC-2001 strain was 2 × 10 ⁸ . 15 larvae of the same larvae in 15 out of 60 cups
Five of the second-stage larvae of the buddha buoy and five of the second larvae of the beetle and one of the second-stage larvae of the beetle were placed in each of the 15 larvae, and bred at 20 ° C. for 40 days, and the number of dead animals was examined. In a similar manner, cups sprayed with sporangia of Bacillus popirie semara strain were prepared and tested to determine the number of dead larvae.

The results are shown in Table 2. The microorganism Bacillus popirie popirie DIC-2001 strain of the present invention had a higher mortality rate against Semaphora kogane and Douganebuui than the Bacillus popirie semadara strain, which is a known microorganism.

[0042]

[Table 2]

It has been found that the microorganism of the present invention exhibits an insecticidal activity of 500% against Semaphora kogane and 165% against D. persicae compared to conventionally known microorganisms. It was found that the microorganism of the present invention stably exhibited the same level of insecticidal activity to Semaphora kogane, Douganebuui, Mamekogane and Himekogane regardless of the type. In other words, the highest insecticidal activity (40%) against the Japanese moss
%) And the lowest insecticidal activity (3
The ratio of the difference (3%) (7%) was 0.175.

In addition, it showed an excellent control effect on the third-instar larvae of Scutellaria cricket.

EFFECTS OF THE INVENTION The microorganism of the present invention induces a lethal emulsification disease on a scarab beetle larva. Therefore, the microorganism of the present invention controls a scarab beetle larva by causing fungi, particularly a sporangia, to act on the larva. Fruit trees, agricultural and horticultural plants, etc. can be protected from the damage of the pests. Especially this microorganism is Bacillus
As a microorganism belonging to Popiriae, it also has an insecticidal activity against larvae of Semaphora kogane and Douganebuibu, which are important pests in Japan, and its use effect is extremely large. In addition, there is almost no toxicity to the natural environment and the human body, and the control method of the present invention is an excellent control method that also contributes to global environmental protection. In addition, it is possible to provide an insecticide exhibiting the same degree of insecticidal effect non-selectively, regardless of the type, to Scarabaeidae insects, particularly to the typical and serious plant pests Douganebuui, Semadarakogane, Himekogane, and Mamekogane. .

[Brief description of the drawings]

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 shows the Bacillus popirie popirie DIC-2001 strain of the present invention (Example 3).
piliae var. popillae DIC-
2001) (FERM P-18250) and Bacillus
It is a graph which shows the comparison of the insecticidal activity with the Popiliae semara strain (FERM P-16818).

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Claims (5)

[Claims] 1. Bacillus popirie popirie DIC-2001, which exhibits insecticidal activity on Semaphora kogane and Douganebui buoy and exhibits strong insecticidal activity against Semaphora kogane.
Strain (bacillus popilliae var.
popillae DIC-2001) (FERM P-
18250). 2. A method for controlling Scarabaeidae insects, comprising causing the microorganism according to claim 1 to act on Scarabaeidae insects. 3. A method for controlling Scarabaeidae insects, which comprises causing the sporangia of the microorganism according to claim 1 to act on Scarabaeidae insects. 4. The control method according to claim 2, wherein the insects of the family Scarabaeidae are the insects of the family Scarabaeidae or the insects of the species Scarabaeidae. 5. An agent for controlling scarabid insects, comprising the sporangium of the microorganism according to claim 1.