JPH0875A - Artificial mass breeding of 'tochukaso' fungus - Google Patents

Abstract

PURPOSE:To provide a process for the artificial mass-cultivation of fungi of TOCHUKASO (fungi parasitic on the larva of lepidopterous insect, etc.) by controlling the growth of a host and that of the fungi. CONSTITUTION:Fungi of TOCHUKASO artificially inoculated on a host insect are grown at a low temperature near the critical growth temperature of the host insect until the formation of a sclerotium and then grown at the optimum growth temperature of the fungi to get fruit bodies.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for artificially breeding Cordyceps sinensis such as Pleurotus cornucopiae, and more particularly to a mass breeding method using dormant pupae of Lepidoptera insects such as Spodoptera litura, dead pupae of Bombyx mori, and larvae.

[0002]

2. Description of the Related Art Cordyceps sinensis, which is said to have about 400 species in the world, has traditionally killed the host by the spores of the fungus parasitizing a living insect, forming a sclerotia in the body of the host insect, and matures. It has been thought to form an entity (mushroom) outside the host's body.
Since Cordyceps sinensis has long been used as a long-lived, tonic and tonic drug in China, it has been reported to have a suppressive effect on cancer cells in recent studies in Japan. However, artificial breeding of Cordyceps sinensis is extremely difficult, and the number that can be collected in the natural world is extremely small. Attempts have been made to artificially raise the animals, but none have reached the point of forming fruiting bodies. Until now, the environmental conditions under which fruit bodies are formed have not been elucidated. Furthermore, the mass breeding method for use in research and industry is completely untouched.

[0003]

[Problems to be Solved by the Invention] In order to investigate the medicinal components of Cordyceps sinensis and to obtain a large amount of the medicinal components, large-scale artificial rearing of Cordyceps fungus is indispensable. For that, a large number of host is necessary. further,
A stable supply is needed. The silkworm pupa after silk thread production are produced in large quantities, but now they are partially used for fish feed and fertilizer, and the rest are discarded. Bombyx mori after producing silk thread is killed by heat. If this dead pupa can be used as a host to breed Cordyceps sinensis, a large amount of Cordyceps sinensis can be raised. By the way, conventionally, it has been considered that Cordyceps sinensis is capable of developing only in living insects. Under what conditions can a dead pupa be used as a host? Clarification of this point is essential.

On the other hand, it is also possible to use pupa dormant insects such as Spodoptera litura and silkworm larvae as a host. These insects can be mass-produced by artificial feed. By the way, it takes about two months from the invasion of Cordyceps sinensis such as Pleurotus cornucopiae into the host to form the primordia of the fruiting body. In the pupa and larvae of living insects, the development of the host progresses during this period and the adults emerge, and the Cordyceps sinensis fungi cannot form fruit bodies. How to control host growth and Cordyceps sinensis growth? The present invention has solved these problems.

[0005]

[Means for Solving the Problems] In order to solve the problem, it is essential to clarify the conditions under which the caterpillar fungus grows, but the host insects and other fungi do not grow. In other words, it is to clarify the temperature and environmental conditions in which the caterpillar fungus grows but the host insect does not. Since the dead pupae are dead, the host does not develop. It is also essential to clarify the environmental conditions that do not spoil the dead pupae and also prevent the development of other fungi and bacteria and promote the growth of Cordyceps sinensis.

[0006] Many studies have been conducted on the growth limit temperature of insects, and although it varies depending on the species, it is 10 ° C to 12 ° C.
The degree of growth is zero. Inoculate the potato agar medium with mycelium of Cordyceps fungus (Pleurotus cornucopiae and Pleurotus cornucopiae),
0 ° C, 5 ° C, 7.5 ° C, 10 ° C, 15 ° C, 20 ° C, 25
Culturing was carried out at 30 ° C. As a result, the optimum culture temperature is 15
℃ to 20 ℃ was found, 0 ℃, 25 ℃, 30
It became clear that it hardly grew at ℃. In particular, hyphae do not grow at high temperatures of 25 ° C or higher.

However, it was revealed that the hyphae of Cordyceps sinensis grow sufficiently even at low temperatures of 5 ° C, 7.5 ° C and 10 ° C. The host insects do not develop at this temperature. It is clear that fruiting bodies are formed by keeping the host inoculated with Cordyceps sinensis in this temperature range for a certain period of time and then returning the temperature to 15 ° C to 20 ° C. became. It was found that dead pupae can also be used as a host by aseptically culturing during this period. It was also clarified that the dormant pupae remain dormant at 10 ° C to 20 ° C, and it is possible to form fruiting bodies during this period.

[0008]

By using the above-mentioned means, it is possible to carry out large-scale artificial breeding of Cordyceps sinensis, mainly from dead pupae, dormant pupae and surviving larvae and pupae. In the case of dead pupae, by aseptically inoculating Cordyceps sinensis and keeping it at 7 ° C to 10 ° C for about 2 months, the sclerotium of Cordyceps sinensis is formed without the development of other bacteria. Then, by breeding at 20 ° C., fruit bodies are formed after about 1 month. That is, a fruiting body of Cordyceps sinensis is obtained in 3 months. When using dormant pupae, inoculate and breed the Cordyceps sinensis in a state that is as sterile as possible. Temperature from 10 ℃ to 20
Keep at ℃ and stay dormant. As a result, sclerotia are formed in about 1 month, and fruiting bodies are formed in the following 1 month. That is, a fruiting body of Cordyceps sinensis is obtained in 2 months. When a non-diapause pupa or larva of Bombyx mori is used as a host, after inoculating Cordyceps sinensis, the silkworm is bred for about 1 month at a growth limit temperature of about 10 ° C, and then raised at 15 ° C to 20 ° C. After several weeks, sclerotia are formed, and in the following month, fruit bodies are formed.

[0009]

EFFECTS OF THE INVENTION A comparison with a fruiting body inoculated with DNA (deoxyribonucleic acid) extracted from a fruiting body of Cordyceps sinensis, which has been artificially bred according to the present invention, that is, those of natural pupae and hanatake mushrooms, was made by comparison with the DNA fingerprinting method. , Found to be the same species. In other words, it became possible to artificially breed a large amount of the same species as the Cordyceps sinensis in nature indoors.

The medicinal effect of Cordyceps sinensis has been handed down for a long time, and it is currently sold in the market as an extremely expensive product. Since the breeding method of the present invention has enabled large-scale breeding of Cordyceps sinensis, it is possible to clarify its medicinal component. Further, when the medicinal components are analyzed, it is possible to produce a large amount of Cordyceps by the breeding method of the present invention, and it is possible to provide a large number of Cordyceps on the market.

It is also possible to rear a pupa inoculated with Aspergillus niger and observe the state in which fruit bodies of Aspergillus niger are formed.
It also plays a very effective role in education and research. That is, it is possible to utilize the caterpillar fungus artificially reared by the present invention and use it as a science teaching material for understanding the mystery of nature.

Furthermore, dead pupae of Bombyx mori have been conventionally treated as wastes after the production of silk thread, and are discarded as well as used as fertilizers and feeds. By using these industrial wastes, effective use of resources can be planned.

[0013]

EXAMPLE An experimental example of the present invention will be described below. [Experimental Example 1] First, a fruit body of Cordyceps sinensis collected from the field is crushed in a mortar to prepare a suspension of ascospores. For the ascospore concentration of the suspension, the number of spores in 1 ml was adjusted to 125,000 using a Toma cytometer. The suspension was inoculated with dead host pupae of silkworm, dead pupa of Spodoptera frugiperda, and live pupae of Bombyx mori. A total of 50 pupae were used in the experiment. Prior to inoculation, all experimental equipment, breeding containers, spaces, etc. were sterilized, and the host was washed with sterile water.

After inoculation, each pupa was placed in a constant temperature dark room at 7.5 ° C for 2 days.
It was bred for a month and promoted the growth of Cordyceps sinensis. Then 2
The cells were transferred to a thermostatic chamber with a light-dark cycle of 12:12 hours at 0 ° C., and the growth of Cordyceps sinensis was promoted. As a result, after 2 weeks, fruit body primordia were formed in 48 of 50 pupae used in the experiment. Of these, 24 individuals formed fruiting bodies, ie, Cordyceps sinensis mushrooms. Fruit bodies were formed in 24 of the 50 pupae, and it was possible to inoculate and breed Cordyceps sinensis with a high probability of about 50%.

[Experimental Example 2] Next, the result of immersing 100 dormant pupae of Spodoptera frugiperda in the ascospore suspension will be described below. 20 pupae were inoculated in the same manner as in Experimental Example 1 above.
The animals were bred for 1 month in a thermostatic chamber at a temperature of 12 ° C for 12:12 hours. As a result, fruit body primordia were formed in 92 of 100 pupae. 20 if dormant pupae are used as host
It became possible to breed at ℃, the growth of Cordyceps sinensis was further promoted, and fruit bodies could be obtained in about one and a half months.

Further, the pupae of live Spodoptera frugiperda were immersed in hot water to cause death, and it was investigated whether or not the dead pupae could be used as a host. In this case, 50 individuals were used in the experiment. After inoculation with spores by the immersion method, Cordyceps sinensis was bred for 2 months in a total dark constant temperature chamber at 7.5 ° C. Then, at 20 ° C., a light-dark cycle 12:
It was transferred to a temperature-controlled room for 12 hours and further promoted the growth of Cordyceps sinensis. As a result, after 2 weeks, fruit body primordia were formed in 43 of 50 pupae used in the experiment. Similar results were obtained when silkworm pupae were killed by treating them with boiling water.

Furthermore, when Bombyx mori pupa is used as a host, since the pupa is larger than the mosquito pupa, a large fruiting body of Aspergillus niger is obtained.

As described above, it has been confirmed by the gene analysis as described above that the Cordyceps sinensis obtained by artificial rearing is of the same species as Pleurotus cornucopiae and Pleurotus cornucopiae collected in the field. It will be possible to breed artificially large amounts of Cordyceps sinensis, which has been extremely difficult for a long time, and it will be extremely important to contribute to the future development of the pharmaceutical industry.

Claims (5)

[Claims] 1. A fruiting body is obtained by growing Cordyceps sinensis artificially inoculated into a host insect at a low temperature around the growth limit temperature of the host insect and after breeding the sclerotium, breeding at a suitable growth temperature. Artificial breeding method for Cordyceps sinensis. 2. A larva of a lepidopteran insect such as Bombyx mori or Spodoptera litura is used as the host.
A method for artificially breeding Cordyceps sinensis as described. 3. The artificial breeding method for Aspergillus oryzae of claim 1, which is characterized in that pupae of Lepidoptera insects such as Bombyx mori and Spodoptera litura are used as the host. 4. The artificial breeding method for Aspergillus oryzae according to claim 1, characterized in that dead pupae of Lepidoptera insects such as Bombyx mori are mainly used to obtain fruiting bodies of Aspergillus oryzae such as Pleurotus cornucopiae. 5. The artificial breeding method for Aspergillus oryzae according to claim 1, wherein a dormant pupa of Lepidoptera insects such as Spodoptera litura is used as the host to obtain fruit bodies of Aspergillus oryzae such as Pleurotus cornucopiae.