JP2021052741A - Culture material production method, plant culturing method, compost, and seedling - Google Patents

Abstract

To provide a culture material that allows bacteria of V. simplex to be cultured properly and facilitates mass production.SOLUTION: The present invention relates to a culture material for culturing bacteria of Veronaeopsis simplex. Conifer wood that is not composted is crushed with an extruder.SELECTED DRAWING: Figure 3

Description

The technique to be disclosed relates to a technique for utilizing a bacterium belonging to Veronaeopsis simplex, and relates to a method for producing a culture material, a method for cultivating a plant, a culture soil, and a seedling.

A wide variety of microorganisms such as bacteria and fungi live in the soil. It is known that many plants coexist with these microorganisms while complementing each other.

The use of such symbiotic microorganisms promotes the growth of specific plants and imparts resistance to diseases and the environment. As a typical example, VA mycorrhizal fungi are widely known.

When VA mycorrhizal fungi infect a plant, VA mycorrhizal roots are formed in the plant. The formation of VA mycorrhiza has the effects of improving the absorption of nutrients and developing resistance to diseases and the environment. Therefore, various studies have been conducted on the use of VA mycorrhizal fungi for plant cultivation (for example, Patent Document 1).

Regarding the technology to be disclosed, tomato seedlings, V.I. A technique for suppressing the absorption of radioactive cesium by inoculating a bacterium belonging to simplex (a strain belonging to Verona eopsis simplex having a receipt number of NITE AP-01933) is disclosed (Patent Document 2).

In Patent Document 2, V.I. It is disclosed that among the fungi belonging to simplex, there are those that can impart high temperature stress resistance and acid resistance to tomatoes and Chinese cabbage as well as VA mycorrhizal fungi.

The applicant has been developing woody soil suitable for agriculture, horticulture, etc., and has disclosed various techniques so far. For example, in Patent Document 3, after coarsely pulverizing coniferous wood with an extruder, the wood shards obtained by further pulverizing with a cutter mill are modified by allowing a predetermined amount of ammonium iron citrate to act on the wood fragments. , Discloses that it is possible to obtain woody soil (soil made from non-composted wood) that allows plants to grow well.

Japanese Patent No. 6315195 JP-A-2018-117589 Japanese Patent No. 6469143

As mentioned above, V.I. Among the bacteria belonging to simplex, there are those that can be expected to have an advantageous effect similar to VA mycorrhizal fungi. Therefore, the present inventors have described V.I. If the fungus belonging to simplex is put into practical use, it can be effectively used for plant cultivation. An attempt was made to mass-produce bacteria belonging to simplex.

V. Bacteria belonging to simplex are fungi. An agar medium or the like can be used for culturing only on a trial basis, but the agar medium or the like is not suitable for mass production in terms of cost.

On the other hand, mushrooms are a typical fungus. Mushrooms have already been mass-produced by fungal bed cultivation. In the fungal bed cultivation, a medium containing a coniferous "ogako (chip)" and a nutrient material such as rice bran is generally used.

Therefore, the present inventors used a culture material containing uncomposted coniferous sardines in the same manner as the medium for such fungal bed cultivation. An attempt was made to cultivate a bacterium belonging to simplex. However, V. It has been found that in the case of a bacterium belonging to simplex, it cannot be cultivated well with such a culture material. Therefore, V. As a result of studying a culture material capable of satisfactorily culturing bacteria belonging to simplex and easily mass-producing, the technology to be disclosed has been established.

That is, the main purpose of the disclosed technology is V.I. An object of the present invention is to provide a culture material capable of satisfactorily culturing a bacterium belonging to simplex and easily mass-producing.

The technology to be disclosed is V.I. It relates to a method for producing a culture material for culturing a fungus belonging to Simplex, and is characterized by including a treatment of crushing uncomposted coniferous wood with an extruder to form wood fragments.

For unknown reasons, uncomposted softwood wood is crushed with an extruder to form wood fragments with a special structure in which fibrous and granular morphologies are appropriately mixed and integrated. V. which cannot be obtained by crushing with a cutter mill. A culture material capable of satisfactorily culturing bacteria belonging to simplex can be obtained.

If it is a piece of wood, it is possible to recycle waste such as wood processing scraps. Moreover, in the case of this type of culture material, it is premised that it is inexpensive and mass-consumed due to its use, but since it is only necessary to pulverize it with an extruder, it is low in cost and easy to put into practical use.

The method for producing the culture material may further include a treatment of mixing a predetermined amount of bran with the wood fragments.

Bran is effective as a nutrient material for bacteria, and the growth of bacteria can be promoted by blending an appropriate amount. Moreover, since the bran is inexpensive, it is suitable for this kind of culture material.

The strain is intended to include at least one strain having a receipt number of NITE AP-01933, a strain having MAFF number 240802, and a strain having a CBS number of "CBS 588.66". Good.

For these strains, V.I. It has been identified as a bacterium belonging to simplex and is available through a given institution.

The base material of the culture material is wood fragments. Therefore, by utilizing the characteristics, the method for producing the culture material can be directly utilized for the method for cultivating plants. That is, a process of forming wood fragments by crushing coniferous wood with an extruder, and V.I. A culture treatment in which a bacterium belonging to simplex is inoculated and cultured may be carried out, and an infection treatment may be carried out in which the wood fragments subjected to the culture treatment are brought into contact with the roots of the plant.

By doing so, effects such as promotion of plant growth and imparting advantageous resistance can be obtained.

Specific examples of utilizing the characteristics of the culture material include potting soil and seedlings for plant cultivation. That is, the culture material produced by the above-mentioned production method is contained in the culture soil for plant cultivation in a state where the bacteria are cultivated, or the culture material produced by the above-mentioned production method and in which the bacteria are cultivated. May be attached to the roots of seedlings.

According to the disclosed technology, V.I. Since a culture material in which bacteria belonging to simplex can be cultivated well and mass production is easy can be obtained, it can be effectively used for plant cultivation.

It is a photograph of the main sample after the culture test. (A) is a comparative example, and (b) is an example. It is a figure for demonstrating the method of measuring the elongation rate of hyphae. It is a graph which summarized the measurement result of the hyphal elongation rate. It is a flow chart which shows the main processing of the plant cultivation method using a culture material. (A) and (b) are specific examples utilizing the shape characteristics of the culture material. It is a graph which summarized the measurement result of the hyphal elongation rate in other strains (K45 strain, CBS strain). The measurement results of the Y34 strain shown in FIG. 3 are also shown.

Hereinafter, embodiments of the disclosed technology will be described in detail with reference to the drawings. However, the following description is essentially merely an example and does not limit the present invention, its application or its use.

<Culture material>
As mentioned above, the inventors of the present invention described V.I. It has been found that the fungi belonging to simplex cannot be cultivated well in a medium based on uncomposted coniferous sardines.

On the other hand, if the applicant modifies the wood fragments formed by crushing uncomposted coniferous wood with ammonium ferric citrate, the plant will be good without removing the growth inhibitor by washing with water. It has been found that it will be able to grow in. The present inventors have focused on this technique, and have described V.I. It was examined whether it could be used for culturing bacteria belonging to simplex.

As a result, for unknown reasons, uncomposted softwood wood is crushed with an extruder to form wood fragments with a special structure in which fibrous and granular morphologies are appropriately mixed and integrated. Therefore, even if the reforming treatment is not performed, V.I. It has been found that a culture material capable of satisfactorily culturing bacteria belonging to simplex can be obtained. The specific test contents will be described.

(Bacteria belonging to V. simplex)
V. There are a plurality of strains of bacteria belonging to simplex. Among the available strains, for example, a strain having a receipt number of NITE AP-01933 (Y34 strain), a strain having a strain name of K45 (K45 strain), and a CBS number of "CBS 588.66" are used. There is a strain having (CBS588.66 strain, hereinafter also referred to as CBS strain).

The K45 strain is registered in the National Institute of Agrobiological Sciences with MAFF number 240802, and can be obtained through the Gene Bank. The CBS 588.66 strain is registered with the Dutch culture collection institution (CBS-KNAW culture collection) under the CBS number (CBS 588.66) and can be obtained through the institution.

(Preparation of bacteria)
A strain of Y34 strain was used in the test (K45 strain and CBS strain will be described later). Cornmeal agar: 8.5 g, malt extract: 10 g, yeast extract: 1.0 g dissolved in 1 L of water, sterilized (121 ° C, 20 minutes)
After that, 20 ml each was dispensed into a sterilized petri dish to prepare an agar medium. The agar medium was inoculated with the bacteria, and each strain was cultured in a culture chamber under appropriate temperature conditions for 2 weeks.

(Making wood fragments)
Sugi scraps were crushed (ground) with an extruder, and the crushed material (wood shards) was subjected to a test and used as an example. On the other hand, sugi scraps were cut and crushed using a cutter mill, sieved with a mesh of 4 mm, and crushed wood having a size of 4 mm or less that passed through the mesh was subjected to a test and used as a comparative example. The definition of the mesh size here refers to the equivalent of the nominal opening of the test sieve specified in JIS Z 8801-1.

(Preparation of culture material)
A culture material was prepared by adding a predetermined amount of nutrient material (bran) and water to each sample of Examples and Comparative Examples. Specifically, rice bran and bran (wheat bran) were mixed in a predetermined ratio with the wood fragments of each sample, and a predetermined amount of water was added.

Specifically, the sample: rice bran: bran was mixed so as to have a ratio of 3: 1: 1 (volume ratio). By adding water to these mixtures, the water content was adjusted so that the water content was 215% based on the absolute dry weight. Each sample (prepared culture material) thus obtained was sterilized in an autoclave (121 ° C. for 40 minutes), and then 40 ml each was sterilized in a sterilized petri dish and subjected to a test.

(Inoculation and culture of bacteria)
From the colonies formed by culturing on the agar medium, a bacterial mass and an agar medium having a diameter of about 5 mm were cut out with a patterned needle and placed in the center of the surface of each sample to inoculate the bacteria. Each sample inoculated with the bacterium was cultured in a culture chamber under a temperature condition of 30 ° C. for 15 days.

FIG. 1 shows the state after culturing. FIG. 1 (a) shows a comparative example, and FIG. 1 (b) shows an embodiment. By culturing, substantially circular colonies are formed on the surface of each sample.

(Measurement of hyphal elongation rate, results)
As shown in FIG. 2, after culturing, two reference lines were set on the lid of the sterile petri dish of each sample so as to be orthogonal to each other on the substantially center of the colony. Based on these reference lines, the diameter of the colony was measured as shown by an arrow in FIG.

From the initial value (diameter of the bacterial mass at the time of inoculation) and the culture period (15 days), the average elongation rate of hyphae (mm / day) per day for each sample was calculated. The result is shown in FIG.

In the comparative example, the average elongation rate of the hyphae was 0.83 (mm / day), whereas in the example, it was 1.50 (mm / day), which was clearly good by pulverization with an extruder. It was confirmed that it could be cultivated.

The reason is not clear, but the extruder adds a grinding action that the cutter mill does not have. Therefore, as a result of the progress of pulverization at the cellular level, V.I. It seems that the increase in the nutrient source components of the fungi belonging to simplex has an effect.

From this test result, it is only necessary to crush the uncomposted softwood wood with an extruder to obtain V.I. It is possible to obtain a culture material capable of satisfactorily culturing bacteria belonging to simplex.

According to the culture material to which the disclosed technology is applied, it is possible to recycle waste such as wood processing scraps. Moreover, in the case of a culture material, it is premised that it is inexpensive and mass-consumed due to its use, but in the disclosed technology, it is only necessary to pulverize it with an extruder, so that the cost is low and it is easy to put it into practical use.

<Cultivation of plants using culture materials>
If the culture material to which the disclosed technique is applied is used, V. Since bacteria belonging to simplex can be cultivated in large quantities at low cost, mass production becomes possible. Mass-produced V. If the fungus belonging to simplex is used for cultivation of agricultural crops and horticultural plants in the same manner as VA mycorrhizal fungi, advantageous effects such as improvement in yield can be expected.

FIG. 4 shows an example of a method of cultivating a plant using a culture material to which the disclosed technique is applied. FIG. 4 shows the basic processing flow of the cultivation method, and the cultivation method is composed of pulverization treatment, mixing treatment, culture treatment, and infection treatment.

The crushing process is a process of crushing coniferous wood with an extruder to form wood fragments. The mixing treatment is a treatment in which bran is mixed with wood fragments as a nutritional material at a predetermined ratio. In the culture treatment, the wood shards mixed with bran were mixed with V. This is a process of inoculating and culturing a bacterium belonging to simplex. These processes may be scaled up according to the specifications based on the test method described above.

In addition, after the culture treatment, V. An extraction process for extracting bacteria belonging to simplex may be performed. For example, a predetermined amount of water is added, stirred and mixed to isolate the fungus from the wood shards. The mixed solution is filtered to prepare a bacterial solution (crude bacterial solution). The crude bacterial solution may be further concentrated by centrifugation or the like to prepare the concentrated bacterial solution.

Infection processing is performed by V.I. This is a process in which a fungus belonging to simplex is brought into contact with the roots of a plant to infect it. Specific examples of the infection treatment include, for example, cultivated wood fragments, that is, V.I. State bacterium belonging to simplex is sufficiently increased wood debris (e.g., wood debris containing 10 ³ or more bacteria per timber debris 1g) and by contacting the roots of plants, may be performed infection process.

That is, in the case of this culture material, since the base material is wood fragments, there is an advantage that it can be used as a cultivation material as it is. Therefore, as shown in FIG. 5 (a), V.I. The culture material 1 in which the fungus belonging to simplex is cultured can be used alone or after being mixed with other cultivation materials and packed in a predetermined bag 2 as a culture soil 3 for plant cultivation. ..

In this case, the potting soil can be sprinkled or mixed with the soil in which the target plant is cultivated to indirectly contact the roots. It is more preferable to mix the potting soil with the soil before cultivation or spray it on the cultivation site of the soil because the roots are easily infected.

In addition, the prepared V.I. The bacterial solution belonging to simplex may be sprayed on the soil in which the target plant is cultivated, or the bacterial solution may be added to the water at the time of watering. When the fungus solution soaks into the soil, the fungus comes into contact with the roots of the plant and infects it.

In addition, for seedlings (young plants before transplantation), V.I. Infection with a bacterium belonging to simplex is effective because it can impart advantageous properties for stabilizing growth in the early stage of cultivation. In general, seedlings are often raised using pots or nursery boxes. Therefore, as shown in FIG. 5 (b), pot 4 (or nursery box) is used to raise seedlings. By filling the culture material 1 in which the fungus belonging to simplex is cultured and planting the seedling 5 there, the culture material 1 is attached to the root of the seedling 5. Before planting, the roots of the seedlings were changed to V. The fungus may be attached to the roots of the seedlings by performing a treatment of immersing in a bacterial solution belonging to simplex.

As another example, the fungus may be infected by planting the seedling 5 in the potting soil to which the fungus solution is added. Further, when the fungus is infected with the culture material 1 or the potting soil 3, the infection may be carried out by sowing seeds or cutting trees instead of seedlings. Further, according to the concept of so-called coated seeds, the seeds may be sprayed with a bacterial solution before sowing, or the seeds may be coated with a material containing endophyte.

<Tests with other strains>
In the above-described embodiment, the content of the test using the Y34 strain has been described. Here, the contents of the test using each of the K45 strain and the CBS strain will be described.

Appropriate culture conditions vary from strain to strain. Therefore, the test conditions are partially changed from the above-described embodiment. First, in the K45 and CBS strains, the proportion of nutrients is halved compared to the Y34 strain (40% to 20%). In the Y34 strain, the sample: rice bran: bran was mixed so as to have a ratio of 3: 1: 1 (volume ratio), but in the K45 strain and the CBS strain, the sample: rice bran: bran was 4: 0.5: 0.5. It was mixed so as to have (volume ratio).

In the CBS strain, the water content was also changed. Specifically, in the Y34 strain, the water content was adjusted so that the water content was 215% of the absolute dry weight, and in the K45 strain, the water content was adjusted in the same manner, but in the CBS strain, the water content was adjusted to the absolute dry weight. The water content was adjusted so as to be 195%.

The culture temperature was also changed. For the Y34 strain, the culture temperature was set to 30 ° C., but for the K45 strain and the CBS strain, the culture temperature was set to 20 ° C. Other test conditions are the same as in the above-described embodiment.

FIG. 6 shows the test results using each of the K45 strain and the CBS strain. The test results (see FIG. 3) with the above-mentioned Y34 strain are also shown.

As shown in FIG. 6, in each of the K45 strain and the CBS strain, as in the Y34 strain, the average elongation rate of hyphae was increased in the examples as compared with the comparative examples. Therefore, it can be seen that in each of the K45 strain and the CBS strain, crushing the uncomposted coniferous wood with an extruder enables clearly good culture.

As described above, since the same effect is observed in a plurality of strains, even in uncomposted coniferous wood, V.I. Good culture of bacteria belonging to simplex can be expected.

1 V. Culture material in which bacteria belonging to simplex are cultivated 2 bags 3 culture soil 4 pots 5 seedlings

Claims (6)

A method for producing a culture material for culturing a bacterium belonging to Veronaeopsis simplex.
A method for producing a culture material, which comprises a process of crushing uncomposted softwood wood with an extruder to form wood fragments. In the method for producing a culture material according to claim 1,
A method for producing a culture material, further comprising a treatment of mixing a predetermined amount of bran with the wood fragments. In the method for producing a culture material according to claim 1 or 2.
A culture medium in which the bacterium contains at least one strain having a receipt number of NITE AP-01933, a strain having MAFF number 240802, and a strain having a CBS number of "CBS 588.66". Manufacturing method. It ’s a method of growing plants,
The process of forming wood fragments by crushing softwood wood with an extruder, and
A culture treatment in which the wood fragments are inoculated with bacteria belonging to Veronaeopsis simplex and cultured.
And
A method for cultivating a plant, wherein an infection treatment is performed in which the cultivated wood fragments are brought into contact with the roots of the plant. Potting soil for plant cultivation
A potting soil for plant cultivation, which comprises the culture material produced by the production method according to any one of claims 1 to 3 in a state in which the bacteria are cultivated. Seedlings
A seedling in which the culture material produced by the production method according to any one of claims 1 to 3 and in which the bacterium is cultured is attached to the root.

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