JP2001352973A - Useful microorganism settling promoter and method for settling useful microorganism - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a new composition affording the stable settling of useful microorganisms on plant rhizosphere, to provide a method for settling such microorganisms, to provide a new composition having stable growth promotive effect of such microorganisms even in soil and having soil disease injury- controlling effect, and to provide a method for realizing the above effects. SOLUTION: This composition is designed to settle useful microorganisms it a plant rhizosphere containing microorganisms belonging to Klebsiella pneumoniae. The 2nd objective method is intended to settle useful microorganisms in a plant rhizosphere by including microorganisms belonging to Klebsiella pneumoniae together with the useful microorganisms in the plant rhizosphere. The 3rd objective plant growth promotive/soil disease injury- controlling agent comprises Pseudomonas fluorescens S272 strain (FER P-16962) and Klebsiella pneumoniae H12 strain (FERM P-17590). The 4th objective method for plant growth promotion/soil disease injury control comprises including FERM P-16962 and FERM P-17590 in a plant rhizosphere.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a composition for fixing a useful microorganism to a plant rhizosphere, and more particularly to a composition for fixing a useful microorganism to a plant rhizosphere using a microorganism belonging to Klebsiella pneumoniae. The composition of Further, the present invention relates to Klebsiella
A method of colonizing a useful microorganism using a microorganism belonging to Klebsiella pneumoniae, a method of promoting plant growth and / or controlling a soil disease using a microorganism belonging to Klebsiella pneumoniae, and a method of controlling a soil disease, and Klebsiella pneumoniae (Klebsiella)
pneumoniae)
Or a soil disease control agent.

[0002]

2. Description of the Related Art In recent years, advanced plant breeding techniques such as genetic manipulation have become possible, and attention has been paid to the development of agricultural crops having pest and disease resistance, high yield, and stress resistance. Among them, the genetic techniques of bacteria or the technique of attaching useful microorganisms to plant tissues using polysaccharides produced by microorganisms are significant in preventing plant diseases and promoting plant growth. Are also being actively conducted.

[0003] The present inventors have previously described Klebsiella
We have been studying polysaccharides that contribute to aggregation and adhesion isolated from microorganisms belonging to Pleumoniae (Klebsiella pneumoniae). The polysaccharide is a useful microorganism, Pseudomonas fluorescen.
By promoting the adhesion of microorganisms belonging to s) to plants,
It has been clarified that it has a plant growth promoting effect, and a patent application has already been filed (Japanese Patent Application No. 11-320043).

[0004] By the way, microorganisms belonging to Pseudomonas fluorescens are bacteria that exist widely in soil and are known to produce various antibacterial substances. For example, various antibacterial substances such as piorteolin, 2,4-diacetylphloroglucinol and pyrrolnitrin produced by microorganisms belonging to Pseudomonas fluorescens (Pseudomonas fluorescens) have a wide range of good antibacterial properties against bacteria, fungi and protozoa. Shows sex. Pseudomonas fluorescens (Pseud
omonas fluorescens) S272 strain (hereinafter also referred to as "S272 strain").

[0005] Klebsiella pneumoniae (Kle
bsiella pneumoniae) microorganisms, forests, soil,
It is widely found in nature as a parasite or symbiosis with water, fresh crops, cereals, and the like. Klebsiella pneumoniae H12 strain (hereinafter "H
12 strains) are considered to be able to produce viscous polysaccharides under special conditions and adhere to the plant root surface. The H
This polysaccharide produced by 12 strains is specifically called an H12 flocculant.

[0006]

However, there were some problems in the adhesive system for simultaneously treating the S272 strain and the H12 flocculant. For example, the H12 flocculant adhered to the plant rhizosphere is considered to be degraded by microorganisms existing in the plant rhizosphere, and the adhesion to the plant rhizosphere does not last long. For this reason, there was a problem in terms of persistence of the adhesive effect of the H12 flocculant. In addition, the purified H12 flocculant is hardly soluble in water,
In addition, since the prepared spray liquid has a high viscosity, inconvenience during use has been pointed out. Furthermore, purification and preparation of the H12 flocculant require a large amount of cost, which is unsuitable for mass production, and its productivity and economic efficiency have been problems. Therefore, the development of a new fixing method and a composition for fixing which solves these problems has been desired.

On the other hand, it has been found that a plant growth promoter containing the S272 strain promotes the growth of a hydroponically grown Japanese radish by simultaneously using an H12 flocculant (Japanese Patent Application No. 11-320043). However, subsequent research has revealed that the plant growth promoter has a problem that a stable effect cannot be obtained in a cultivation system via soil. Therefore, it is desired to develop a new method and a growth promoter that can achieve a stable plant growth promotion effect even in soil.

[0008] As a method for controlling diseases in soil, the use of many fungicides such as methyl bromide and PCNB has been known. However, there is a concern that these fungicides affect the atmosphere and soil, and it is desired to eliminate these fungicides. In addition, improper use may affect humans and animals, and care must be taken when handling them. Therefore, it is desired to newly provide a method and a substance for controlling a natural soil disease which is less harmful to the environment and harmless to humans and livestock.

The present invention has been made in view of the above problems, and an object of the present invention is to provide a composition for stably fixing useful microorganisms to a plant rhizosphere and a novel method showing stable fixation. Aim. Further, another object of the present invention is to provide a novel composition and a method having a stable growth promoting effect and a soil disease controlling effect even in soil.

[0010]

Means for Solving the Problems The present inventors have conducted intensive studies to solve the above-mentioned problems, and as a result, have found that useful microorganisms such as the S272 strain and Klebsiella pneumoniae (K
lebsiella pneumoniae), by simultaneously or closely treating the H12 strain, the colonization of the S272 strain in the plant rhizosphere is improved, the growth of various crops is promoted, and the control effect against soil diseases is also achieved. Was found, and the present invention was completed.

That is, the present invention relates to a composition for fixing a useful microorganism to a plant rhizosphere, which comprises a microorganism belonging to Klebsiella pneumoniae. Preferred embodiments of the composition of the present invention include the following embodiments. (1) Klebsiella pneumonie
An embodiment in which the microorganism belonging to Moniae is Klebsiella pneumoniae strain H12. (2) An embodiment in which the useful microorganism is a microorganism belonging to Pseudomonas fluorescens. (3) An embodiment in which the useful microorganism is Pseudomonas fluorescens S272 strain.

The present invention also relates to a method for establishing a useful microorganism in a plant rhizosphere, wherein a microorganism belonging to Klebsiella pneumoniae is present in the plant rhizosphere together with the useful microorganism. About the method. Preferred embodiments of the above method include the following embodiments. (1) Klebsiella pneumonie
An embodiment in which the microorganism belonging to Moniae is Klebsiella pneumoniae strain H12. (2) Useful microorganism is Pseudomonas fluorescens
An embodiment which is a microorganism belonging to (Pseudomonas fluorescens). (3) Useful microorganism is Pseudomonas fluorescens
(Pseudomonas fluorescens) An embodiment which is strain S272.

Further, the present invention provides a method for preparing Pseudomonas fluorescens S27 in the rhizosphere of a plant.
Two strains and Klebsiella pneumoniae
moniae) H12 strain, and a plant growth promoting and / or soil disease controlling agent.

Further, the present invention provides a method for preparing Pseudomonas fluorescens S27 in the rhizosphere of a plant.
Two strains and Klebsiella pneumoniae
moniae) H12 strain, and a method for promoting plant growth and / or controlling soil diseases.

[0015]

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

[Composition for fixing useful microorganisms to plant rhizosphere] The composition for fixing useful microorganisms to the plant rhizosphere of the present invention is Klebssiella pneumoniae.
iella pneumoniae). The composition of the present invention is used to establish useful microorganisms in the plant rhizosphere by using "Klebsiella pneumoniae".
Microorganisms belonging to ". This is "Klebsierra
It utilizes the property of microorganisms belonging to Pleumoniae (Klebsiella pneumoniae), which promotes the establishment of useful microorganisms in the plant rhizosphere. Therefore, in the composition of the present invention, all "Klebsiella pneumoniae" as long as they have the property of promoting the colonization of useful microorganisms in the plant rhizosphere
(Klebsiella pneumoniae).

[0017] "Klebsie Klebsie
The microorganism belonging to “lla pneumoniae” is a facultative anaerobic bacterium, and its genetic research is vigorously conducted as a strain having nitrogen fixing ability under anaerobic conditions and useful as a plant nitrogen fixation. Klebsie
lla pneumoniae), for example, Klebsiella pneumoniae (Klebsiella pneumoniae)
In addition to the H12, Klebsie
lla pneumoniae) ATCC 29516.

[0018] Klebsiell
a pneumoniae) among the microorganisms belonging to Klebsiella
Klebsiella pneumoniae H12 strain is particularly useful from the viewpoint of efficiently establishing useful microorganisms in the plant rhizosphere because it has a high ability to produce polysaccharides (H12 flocculants) involved in adhesion to plant root surfaces. . In addition,
Klebsiella pneumonia
e) The H12 strain has been deposited as FERM P-17590 with the Institute of Biotechnology and Industrial Technology, Ministry of International Trade and Industry.

Klebsiell
a pneumoniae) can be produced (cultivated) by a known method. For the culture, a culture method using a general medium and a culture solution can be used. still,
Klebsiella pneumonia
When the microorganism belonging to e) uses a medium containing ethanol as a carbon source, the productivity of polysaccharide may be improved, but the method is not limited to such a method.

[0020] Klebsiell
a pneumoniae) when culturing a microorganism belonging to the medium,
The medium may contain any nutrient source that can be used by microorganisms belonging to Klebsiella. Examples of such a medium include a YMPG medium (0.3% yeast extract, 0.3% malt extract (Difco), 0.5% polypeptone (Nihon Pharmaceutical), 1% glucose, pH 7.0) and a YSE medium (0.5%).
% Yeast extract (Difco), 0.5 % soybean meal, 2% _{ethanol, 0.05% K 2 HPO 4,} 0.05% MgSO 4 · 7H 2 0, pH 7.0) In addition to various synthetic media, including, semisynthetic medium, natural A medium and the like can be mentioned.

[0021] Klebsiell
Various kinds of carbon sources and nitrogen sources can be used for the medium composition of microorganisms belonging to the genus apneumoniae). As a carbon source,
For example, ethanol, glucose, sucrose, starch syrup, molasses, dextrin, starch, glycerol, animal oil, vegetable oil and the like can be used alone or in combination. As the nitrogen source, soybean flour, wheat germ, corn stapler, cottonseed meal, meat extract, peptone, yeast extract, urea, ammonium sulfate, or the like can be used alone or in combination. If necessary, an inorganic acid salt such as sodium chloride, magnesium sulfate, calcium carbonate, phosphate, and metal salt can be added.

[0022] Klebsiell
As a method for culturing a microorganism belonging to A. pneumoniae), a method for culturing under aerobic conditions such as shaking culture by liquid culture and stirring culture is preferable. Klebsiella pneumoniae
A microorganism belonging to (Klebsiella pneumoniae) is inoculated into a nutrient medium, and cultured aerobically to grow the cells.

[0023] Klebsiell
The culture conditions for the microorganisms belonging to A. pneumoniae) can be appropriately selected so that the strain used grows well. This culture condition also depends on the characteristics of microorganisms belonging to Klebsiella pneumoniae to be used,
It can be changed appropriately according to external conditions and the like. Specifically, H
In the case of 12 strains, the pH of the medium is about 6 to 8. Usually, it is preferable to be near neutral. The culture temperature is a temperature at which the microorganism grows well. That is, the culture temperature is 15 ° C to 40 ° C, preferably 25 ° C to 35 ° C. The culturing time depends on other culturing conditions, but generally may be about 2 to 10 days, preferably,
2-5 days.

The microorganism belonging to Klebsiella pneumoniae has a level of 1 according to the microbial safety standard by the Japanese Bacteriological Journal (50, p. 1023, '95), and therefore has an appropriate level. Large-scale culture production under appropriate control can also be easily performed.

In the composition of the present invention, the microorganisms belonging to Klebsiella pneumoniae can use the cultured live bacteria as they are. Alternatively, the culture solution may be freeze-dried.
Alternatively, the microbial cells may be dried cells by microwave irradiation or the like. The lyophilized culture solution and dried cells can be used, for example, suspended in water before use.

The composition of the present invention can be used as long as the effect of colonizing useful microorganisms is not reduced.
(Klebsiella pneumoniae). For example, in addition to microorganisms belonging to Klebsiella pneumoniae (Klebsiella pneumoniae), a culture solution obtained by culturing the microorganisms, a composition containing a culture medium, and further optionally other solvents, carriers,
The composition may include an auxiliary agent, a diluent and the like.

The composition of the present invention is for fixing useful microorganisms to the plant rhizosphere. The useful microorganisms include, for example, antibacterial substances, antiviral substances or nutrients necessary for plant growth. Microorganisms that can be used. Such useful microorganisms include, for example, Pseudomonas fluorescens
cens), Streptomyces kasugaensis, mycorrhizal fungi that promote phosphorus metabolism, and the like.

Pseudomonas Fluorescence (Pseud
omonas fluorescens) include, for example, Pseudomonas fluorescens (Pseudomones) that produces various antibacterial substances including pioluteolin, 2,4-diacetylphloroglucinol, and pyrrolnitrin.
as fluorescens) S272 strain. The composition of the present invention is particularly useful for colonizing Pseudomonas fluorescens S272 strain as a useful microorganism in a plant rhizosphere. In addition, Pseudomonas fluorescens (Pseudomonas fluorescens)
The S272 strain has been deposited with the Ministry of International Trade and Industry at the National Institute of Advanced Industrial Science and Technology as FERM P-16962.

The "plant rhizosphere" and "method of colonization" using the composition of the present invention will be described in detail in the following description of the present invention relating to "method of fixing useful microorganisms to plant rhizosphere".

[Method of fixing useful microorganisms in plant rhizosphere] The present invention is a method of fixing useful microorganisms in plant rhizosphere. In the method of the present invention, the microorganism belonging to Klebsiella pneumoniae is present in the plant rhizosphere together with the useful microorganism in the plant rhizosphere, whereby the useful microorganism is established in the plant rhizosphere.

The plants to which the useful microorganisms are established, which are targeted by the method of the present invention, include both hydroponic plants and soil cultivated plants. For example, vegetables such as spinach, Chinese cabbage, cucumber, eggplant, perilla, cabbage, shungiku, leek, radish, sweet potato, potato,
Root crops such as onions, grains such as rice, wheat, corn, beans such as soybeans, adzuki beans and peanuts, special crops such as beets, sugar cane and asa, grapes, oranges, oysters, apples, tomatoes, melons, pears and strawberries , Fruits such as peach, banana, pineapple, coffee, houseplants such as rubber, phoenix, and benjamin; flowers such as chrysanthemum, carnation, rose, fennel, lily, and tulip.

[0032] The term "rhizosphere" of a plant in the present specification means not only a rhizome of a plant but also a tuber having a tube shape or a corm having a spherical shape.

In the method of the present invention, the useful microorganism can be established in the plant rhizosphere by allowing the microorganism belonging to Klebsiella pneumoniae to coexist with the useful microorganism. More specifically, a microorganism belonging to Klebsiella pneumoniae (Klebsiella pneumoniae) is supplied to a plant rhizosphere where a useful microorganism is present, or a useful microorganism and Klebsiella pneumoniae (a useful microorganism is present or absent in a plant rhizosphere). Klebsiell
a pneumoniae) or supplying the microorganism belonging to Klebsiella pneumoniae (Klebsiella pneumoniae) to the plant rhizosphere that supplied the microorganism belonging to, and then supplying the useful microorganism, to establish the useful microorganism in the plant rhizosphere. it can. However, it is preferable to supply the useful microorganism and the microorganism belonging to Klebsiella pneumoniae at the same time or in close proximity, since the effect obtained by colonization of the useful microorganism into the plant rhizosphere is high.

As a method for supplying useful microorganisms and microorganisms belonging to Klebsiella pneumoniae to the plant rhizosphere, for example, seed dressing or soaking treatment at the time of sowing, soil irrigation treatment, soil mixing treatment, transplantation of seedlings, etc. Examples of the method include root immersion treatment, transplanted seedling root treatment, planting hole treatment, and spraying treatment on plants. More specifically, in the case of hydroponic cultivation, it can be treated by adding to a nutrient solution tank in the case of submerged circulating hydroponic culture, NFT method, or hydroponic cultivation using a fixed medium. When a fixed medium is used, it can be mixed in advance. In the case of soil cultivation, methods of mixing with soil at the time of sowing, raising pots, or planting, or irrigating the soil, immersing the roots of transplanted seedlings in a microbial solution, or planting transplanted seedlings There is a method in which granulation is performed immediately before transplanting a seedling into a hole. Further, it can be sprayed, granulated and dusted on the pot cultivated seedlings and the roots of the seedlings. In addition, it can be sprayed directly on the foliage of the crop.

The number of supply of useful microorganisms and microorganisms belonging to Klebsiella pneumoniae to the plant rhizosphere is not limited, and it is possible to appropriately treat them according to the growth state and growth period of the plant. it can.

When supplying to the plant rhizosphere, the microorganisms belonging to Klebsiella pneumoniae may be used as they are, as they are, or they may be used by mixing them with starches, clays and the like. You can use it.

[Plant Growth Promoter and Soil Disease Control Agent] The plant growth promoter and soil disease control agent of the present invention are all S
272 and H12 strains. The S272 strain and the H12 strain were each FERM as described above.
It is a microorganism deposited as P-16962 and FERM P-17590. The plant growth promoting agent and soil disease controlling agent of the present invention may be the S272 strain and the H12 strain, which may be the cells themselves or a medium solution containing the cells, but may also be a freeze-dried product of the culture solution or a microorganism solution. The cells may be dried cells obtained by irradiating the cells with microwaves. The lyophilized product and the dried cells can be used as they are as plant growth promoters and soil disease control agents, but they may be used after being suspended in water.

The content ratio and amount of the S272 strain and the H12 strain in the plant growth promoting agent and the soil disease controlling agent of the present invention are not particularly limited as long as they can exert the plant growth promoting effect and / or the soil disease controlling effect. There is no limitation. In addition, it can be used after being appropriately changed depending on the kind of the target plant, the use form, the use method, the use time and the like.

The plant growth promoting agent and soil disease controlling agent of the present invention may be used in a range of not decreasing the plant growth promoting effect or soil disease controlling effect, if necessary, with commonly used conventional carriers and diluents. Can be applied in the form of, for example, powders, emulsions, wettable powders, granules and the like.

Further, the plant growth promoter and soil disease control agent of the present invention can be used in admixture with other known plant growth regulators, herbicides, fungicides, insecticides and the like.
Further, auxiliary agents such as spreading agents, emulsifiers, wetting agents, dispersants, fixing agents and disintegrants may be contained. When using these carriers, diluents, auxiliaries, and the like, it is preferable to select and use those that have the most effective plant growth promoting action and / or soil disease control action.

The plant growth promoter and soil disease control agent of the present invention can be applied to both hydroponic and soil cultivated plants. Examples of such plants are the same as those mentioned in the composition for colonizing useful microorganisms in the aforementioned plant rhizosphere. Further, the soil disease controlling agent of the present invention is mainly a soil-borne plant disease Fusarium spp., Pycium spp., Trichoderma spp., Rhizopus spp., Aphanomyces spp., Rhizoctonia spp., Verticillium spp. It is effective against diseases caused by bacteria such as Agrobacterium, Erbinia, and Streptomyces, which are a kind of fungal and plant pathogenic bacteria such as T. elabiopsis. And, for example, Pseudomonas solanacearum (Pseudomonas solanacearum) caused by infestation of tobacco blight or solanaceous plant blight caused by parasitism, legumes, cucurbitaceae, okra family plants, etc., black roots generated by Teielaviopsis basicola (Thielaviopsis basicola) Disease, spinach wilt fungus (Fusarium o
xysporum f. sp. Spinaciae, caused by spinach wilt, Verticilli darrier (Verticilli)
um dahliae), wilt disease of various crops, Agrobacterium tumefaciens
ens) against many diseases such as rose head carcinoma.

[Method of Promoting Plant Growth, Method of Controlling Soil Disease]
The method for promoting plant growth and the method for controlling soil diseases according to the present invention comprise:
272 strain and H12 strain coexist in the plant rhizosphere. The S272 strain and the H12 strain are microorganisms deposited as FERM P-16962 and FERM P-17590, respectively, as described above. The S272 strain and the H12 strain may be any of the cells themselves, a medium solution containing the cells, a freeze-dried product of the culture solution, a dried cell, etc. Is also good.

The S272 strain and the H12 strain are, for example,
Supplying the H12 strain to the plant rhizosphere where the S272 strain is present, supplying the S272 and H12 strains to the plant rhizosphere, or to the plant rhizosphere supplied with the H12 strain, then to S2
By supplying 72 strains, they can coexist in the plant rhizosphere. However, it is preferable to supply the S272 strain and the H12 strain at the same time or close to each other, since a high plant growth promotion and / or soil disease control effect can be obtained. Alternatively, the S272 strain and the H12 strain may be supplied in advance to soil or the like where seeds are grown, and at the time of rooting, the S272 strain and the H12 strain are already present, and soil diseases may be controlled in advance. As a method for supplying the S272 strain and the H12 strain to the plant rhizosphere, for example, irrigation treatment, soaking treatment, seed dressing and soaking treatment at the time of sowing, soil irrigation treatment, soil mixing treatment, root soaking treatment of transplanted seedling, transplantation Examples of the method include seedling treatment, planting hole treatment, and spraying treatment on plants. More specifically, in the case of hydroponic cultivation, it can be treated by adding it to a nutrient solution tank in a method such as a hydroponics circulating hydroponic culture, an NFT method, or a hydroponic culture using a fixed medium. When a fixed medium is used, it can be mixed with the medium in advance. In the case of soil cultivation, methods of mixing with soil at the time of sowing, raising pots, or planting, or irrigating the soil, immersing the roots of the transplanted seedlings in a microbial solution, and planting holes for transplanted seedlings There is a method in which granulation treatment is performed immediately before transplanting a seedling into a plant. Further, it can be sprayed, granulated and dusted on the pot cultivated seedlings and the roots of the seedlings. In addition, it can be sprayed directly on the foliage of the crop.

The S272 strain and H1 in the method of the present invention
The abundance ratio of the two strains can be appropriately determined according to the growth environment of a hydroponic system or a soil cultivation system, the type of a target plant, and the like.

[0045]

The present invention will be described in more detail with reference to the following preferred embodiments.

<Example 1> Plant growth promotion effect in a hydroponic system A cotton wool was spread on a 6-well microtiter plate, and 10
After moistening with 10 ml of water, 10 seeds of Japanese radish were sowed in each section. In order to improve the experimental accuracy, six more sections under the same conditions were prepared. Cultivated while keeping the humidity under 28 ° C. light irradiation, and at the time of seedling germination on the second day, a culture solution of Klebsiella pneumoniae H12 strain or
(Pseudomonas fluorescens) S272 culture (both bacteria are YSE medium (0.5% yeast extract (Difco), 0.5%
Soy flour, 2% ethanol, 0.05% K ₂ HPO ₄ , 0.05% MgSO ₄
(7H ₂ O, pH 7.0) at 28 ° C. for 2 days) on the second to third days. Prepare a 1% solution of H12 flocculant and dilute it to a concentration of 10 mg / l.
Added at day. CaCl ₂ has a concentration of 300 mg / l,
Added at day. The average elongation (mm) of the stem on day 5 after sowing was measured. The average value was calculated by averaging the values obtained from the other eight samples, excluding the maximum and minimum ones among the ten particles, and the values measured for the six samples under the same conditions. Sowing 5
Table 1 shows the stem elongation on the day.

[0047]

[Table 1]

[0048] As a result, stem elongation was clearly accelerated when both bacteria were treated simultaneously or in proximity to each other, compared to when each strain was treated alone. Klebsiella pneumoniae
H12 bacteria belonging to (Klebsiella pneumoniae) and Pseudomonas fluorescens
S272 bacteria belonging to Pseudomonas fluorescens (Pseudomonas
It is considered that the same growth promoting effect as in the case of treating the S272 bacterium belonging to S.

Example 2 Effect of Promoting Plant Growth in Soil System The seedling raising soil is a mixture of red volcanic soil and black volcanic soil in a volume ratio of 1: 1 and a 10% volume ratio of peat moss. The soil used was a well-mixed soil and chemical fertilizer. This soil was filled in a pot having a diameter of 6 cm, and cucumber seeds (variety; Sharp One) were sown. Klebsiella pneumoniae (Klebsiella pnoniae) prepared in the same manner as in Example 1.
eumoniae) H12 strain (10 ⁷ cfu / ml) and Pseudomonas fluorescens (Pseudomonas f.
luorescens) S272 strain (10 ⁷ cfu / ml)
Was irrigated at a volume of 10 ml / pot. In addition, Klebsiella pneumoniae (Klebsiella pneumoniae) H12
H12 flocculant was added to soil at a concentration of 10 instead of strain treatment.
Mixing treatment was performed so as to be 0 ppm and 1000 ppm.
The germination rate 7 days after sowing, the seedling establishment rate 21 days after sowing, and the above-ground weight were examined. The test was performed with 10 pots and 5 repetitions per section, and the average value was shown. Table 2 shows the results.

[0050]

[Table 2]

As a result, both the germination rate and the above-ground weight of the mixed treatment of both bacteria were higher than those of the untreated group, and a growth promoting effect was recognized. In addition, since seedling withering was observed after emergence,
As a result of examining the number of remaining seedlings and examining the seedling establishment rate ((number of seedlings / number of seeds) × 100), the establishment rate of seedlings was higher than that of no treatment.

<Example 3> Control effect on eggplant wilt disease Eggplant (cultivar: Senryo No. 2) was sowed in a 98-well seedling cell seedling box filled with sterilized volcanic black soil and cultivated for 3 weeks. Cell seedlings were prepared. The cell seedlings were raised in pots having a diameter of 9 cm filled with soil prepared in the same manner as in Example 2, and cultured in the same manner as in Example 1 to obtain Klebsiella pneumoniae H12 strain and Pseudomonas fluorescens ( Pseudomonas fluorescen
s) 10 ml of the bacterial solution of the S272 strain (10 ⁶ cfu / ml)
1 / pot volume was irrigated. After cultivation in a greenhouse for 3 weeks, the plants were planted in a container (9 liters of soil) filled with soil contaminated with eggplant wilt fungus (Verticillium dahliae). Disease investigation was performed 50 days after planting. Investigation criteria are index 0 when no wilt disease symptoms are observed, and index 1 when the wilt leaf of the top leaf is 1/3 or less of the plant height.
The index of 1/3 to 2/3 was set to index 2, and the index of 3 was set when it was found to be 2/3 or more. The disease severity is Σ (index ×
The number was calculated by multiplying the (number of concerned strains) / 3 × the number of investigated strains by 100. Further, the control value was calculated by a formula of (1− (untreated disease rate−untreated disease rate) / untreated disease rate) × 100, and shown in Table 3. In addition, Klebsiella
Instead of the bacterial solution treatment of Klebsiella pneumoniae H12 strain, the concentration of H12 flocculant in soil was 1%.
000 ppm. Table 3 shows the results.

[0053]

[Table 3]

[0054] Klebsiell
apneumoniae) H12 strain and Pseudomonas fluorescens S272 strain clearly suppressed the onset of disease and showed a high effect.

<Example 4> Control effect on spinach wilt The contaminated soil of spinach wilt was previously treated with spinach wilt (Fusarium oxysporum f. Sp. Spinacia).
The spores obtained in e) were inoculated into soil and cultured for 7 days. Klebsiellapneumoniae H12 strain and Pseudomonas fluorescens S according to the method of Example 1
A 10 ⁷ cfu / ml bacterial solution of the 272 strain was prepared. Also, Klebsiella pneumoniae
For the H12 strain, a bacterial solution of 10 ⁹ cfu / ml was also prepared according to Example 1. A pot having a diameter of 9 cm was filled with contaminated soil of spinach wilt and 10 seeds (variety: Pareto) were sown per pot. After seeding, Klebsiella pneumoniae H12 strain and Pseudomonas fluorescens (Pseudomonas f.
luorescens) S272 strain 10 ⁷ cfu / ml bacterial solution
ml / pot treatment. In addition, Klebsiella Nyuumoniae (Klebsiella pneumoniae) 10 ⁹ cfu of H12 strain
Spinach seeds are immersed in / ml bacterial solution, and then sown,
Pseudomonas fluorescens
escens) 10 ml of 10 ⁷ cfu / ml bacterial solution of S272 strain
/ Pot treatment. Disease investigation was performed 31 days after sowing.
Investigations showed that the index was 0 for healthy seedlings, index 1 for cotyledons withering, index 2 for cotyledons and true leaves withering, or that the entire seedlings were withering or after germination but before germination The seedling that died was designated as index 3. Disease incidence and control value were determined in the same manner as in Example 3. In addition, Klebsiella
Instead of the bacterial solution treatment of Klebsiella pneumoniae H12 strain, the concentration of H12 flocculant in soil was 1%.
000 ppm. The test was repeated 10 times in 1 pot per section. As a result, as shown in Table 4, Klebsiella pneumoniae H
Twelve strains obtained the same effect as when the seeds were treated with irrigation to the soil.

[0056]

[Table 4]

[0057]

According to the composition and method of the present invention for promoting the establishment of useful microorganisms in the plant rhizosphere, the microorganism belonging to Klebsiella pneumoniae can be co-existed with the useful microorganisms in the plant rhizosphere. Of useful microorganisms can be improved, and as a result, plant growth and the like can be promoted.

Further, according to the composition and method for promoting the colonization of useful microorganisms in plant rhizospheres of the present invention, Klebsiella pnumoniae (Klebsiella pnuniae) cultured under simple culture conditions is used.
eumoniae). Therefore, costs required for purification and preparation can be reduced, and mass production can be economically achieved.

Further, according to the composition and method of the present invention for promoting the establishment of useful microorganisms in a plant rhizosphere, not only a hydroponic system but also a cultivation system via soil, such as spinach, lettuce, eggplant and cucumber. It has the advantage that it can promote the growth of various agricultural crops, including others.

Further, according to the agent and method for promoting plant growth and controlling soil diseases according to the present invention, it is possible to provide a good control action against soil diseases caused by filamentous fungi such as Fusarium and Verticillium. it can.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) C12R 1:22) C12R 1:22) (C12N 1/20 (C12N 1/20 E C12R 1:39) C12R 1:39) F term (reference) 4B065 AA29X AA43X AC14 CA47 CA60 4H011 AA01 AB03 BA01 BB21 BC03 BC18 BC21 BC22 DA13 DD04 DF04 DG05

Claims (10)

[Claims] 1. Klebsie
lla pneumoniae), a composition for establishing useful microorganisms in the plant rhizosphere. 2. The Klebsiella pneumoniae (Kle
bsiella pneumoniae)
Klebsiella pneumoniae strain H12 (F
ERM P-17590). 3. The composition according to claim 1, wherein the useful microorganism is a microorganism belonging to Pseudomonas fluorescens. 4. The composition according to claim 1, wherein the useful microorganism is Pseudomonas fluorescens S272 strain (FERM P-16962). 5. A method for establishing a useful microorganism in a plant rhizosphere, wherein a microorganism belonging to Klebsiella pneumoniae is present in the plant rhizosphere together with the useful microorganism. 6. The Klebsiella pneumoniae (Kle
bsiella pneumoniae)
Klebsiella pneumoniae strain H12 (F
ERM P-17590). 7. The method according to claim 5, wherein the useful microorganism is a microorganism belonging to Pseudomonas fluorescens. 8. The method according to claim 5, wherein the useful microorganism is Pseudomonas fluorescens S272 strain (FERM P-16962). 9. Pseudomonas fluorescens (Pse
udomonas fluorescens) strain S272 (FERM P-1)
6962) and Klebsiell
a pneumoniae) H12 strain (FERM P-17590)
And a plant growth promoting and / or soil disease controlling agent comprising: 10. Pseudomonas fluorescens strain S272 (FE) in a rhizosphere of a plant.
RM P-16962) and Klebsiella pneumoniae strain H12 (FERM P-
17590). A method for promoting plant growth and / or controlling soil diseases.