CN102776127B - Endophytic fungus with preventive and treatment effects on soft rot of dendrobiums - Google Patents
Endophytic fungus with preventive and treatment effects on soft rot of dendrobiums Download PDFInfo
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Abstract
The invention discloses an endophytic fungus with preventive and treatment effects on soft rot of dendrobiums. The fungus can be subjected to field planting at the root of dendrobiums, and is used for seedling raising or cultivation of dendrobiums. The endophytic fungus with preventive and treatment effects on soft rot of dendrobiums can effectively prevent the soft rod of dendrobiums from happening.
Description
Technical field
The present invention relates to technical field of bioengineering, relate in particular to the endogenetic fungal bacterial strain that 2 strains have the biological control effect to Dendrobium plant soft rot.This two fungal strain is used for growing seedlings or cultivating of Dendrobium plant, can prevents and treats the generation of Dendrobium plant soft rot effectively.
Background technology
The famous and precious Chinese medicinal materials of a lot of tradition is arranged, such as Herba Dendrobii and dendrobium stem etc. in the orchid.According to 2005 editions " records of Chinese pharmacopoeia, dendrobium stem (Dendrobium nobile LindL.), Herba Dendrobii (D.candidum Wall, ex Lind1.) or stem of Eyeshaped Dendrobium (D.fimbriatum Hook, var.oculatum Hook.) and the fresh or dry stem of allied species can do the Chinese medicine stem of noble dendrobium and use, have reinforcing stomach reg fluid, nourishing Yin and clearing heat effect, can also make eye bright by relieve sore throat mediator's body immunity function.
Having entered since 21 century, is the coming out one after another of Chinese patent medicine of main raw material with the stem of noble dendrobium along with Shiyu Yeguang Wan, MAILUONING ZHUSHEYE etc. particularly, and the consumption of the Chinese medicine stem of noble dendrobium heightens, and wild stem of noble dendrobium resource faces huge regeneration pressure.Because ecotope goes from bad to worse, and to the overexploitation of natural resources, the famous and precious kind in the Dendrobium plants such as Herba Dendrobii, Herba Dendrobii is endangered.Artificial breeding is a protection Dendrobium plant resources in conjunction with the big area artificial culture, guarantees the important channel of stem of noble dendrobium medicinal material market supply.But along with the expansion of the artificial cultivated area of various Dendrobium plants, the generation of Plant diseases also is on the rise, and this phenomenon has caused common attention.
Soft rot be a kind of in the Dendrobium plant growing recurrent disease, when susceptible from pseudobulb, water stain shape scab appears in site of pathological change, brown is to black, finally makes pseudobulb become soft shrinkage and dead color, rots rapidly.When disease was serious, soft rot can cause the big area total crop failure of raise crop, has seriously restricted the development of stem of noble dendrobium industry.Control to Dendrobium plant soft rot at present mainly contains dual mode: the one, pass through cultivation management, as control the humidity in seedbed, ventilation, the sterilization at water source, the sunlight of cultivation matrix are exposed to the sun and are handled etc., can the control of disease be played a role, but this class mode is time-consuming, operative technique lacks standard, can be handling poor, to disease to prevent and treat efficient not high; Another kind is by using agricultural chemicals in a large number, continually, though can produce the better prevention effect in a short time, cause pathogenic bacteria to produce resistance easily, and a large amount of uses of agricultural chemicals being very big to pollution and the ecological balance damage effect of environment.
In recent years, generation, the development that prevents and handle Plant diseases with the method for biological control more and more causes people's attention.The most significant advantage of these class methods is safety, environmental protection, with low cost, can reduce the use of agricultural chemicals in a large number.Therefore, the method for preventing and treating of research highly effective and safe and economical and practical Dendrobium plant soft rot just seems necessary.
Summary of the invention
The endogenetic fungus that the object of the present invention is to provide two strains that Dendrobium plant soft rot is had the biological control effect.They are applied to the artificial culture of Dendrobium plant, can reduce the sickness rate of soft rot, improve the output of the stem of noble dendrobium, alleviate the pressure of Chinese medicine stem of noble dendrobium market supply.
2 strain endogenetic fungus provided by the invention are respectively through evaluation: bacterial strain Dhs96 is head mold (Rhizopus sp.), and deposit number is CGMCC No.3786, and bacterial strain Dfl88 is wood mould (Trichoderma sp.), and deposit number is CGMCC No.3787.This 2 fungal strain on April 28th, 2010 in China Committee for Culture Collection of Microorganisms's common micro-organisms center preservation, depositary institution address: depositary institution address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica.The preservation of 2 fungal strains and survival proof are seen Appendix.
The cultural characters of two fungal strains is respectively:
Dhs96: bacterium colony is yellowish brown on the PDA substratum, circle, and the carpet shape, the mycelia densification, back side tawny has rhizoid to form; Aerial hyphae is flourishing, and 10 days colony diameter of 25 ℃ of growths is 6.5cm; Do not produce spore on the conventional substratum.
Dfl88: bacterium colony circle on the PDA substratum, the initial stage bacteria colony white, the later stage front and back is light yellow, and mycelia is more sparse, the carpet shape; The aerial hyphae prosperity, 36 hours colony diameter 9cm of 25 ℃ of growths; Conidiophore bears from the side shoot of mycelia, and is upright, branch, and does not expand on the top, is going up living spore ball estranged, contains 4～12 in spore; Conidium is spherical to avette, light color, 2.8～4.5 * 2.2～3.9 μ m.
Above-mentioned two strain endogenetic fungal bacterial strains can be in the field planting of Dendrobium plant roots, plant is not had pathogenic, and the soft rot of the multiple Dendrobium plant of the stage of growing seedlings and field cultivation is had preventive and therapeutic effect, has excellent application value.
Embodiment
Embodiment 1
The biological control of the dendrobium stem soft rot of Dhs96
Endogenetic fungus Dhs96 mixes with the wheat bran solid substrate after the PDA liquid fermentation and culture and makes the solid vaccination agent.Collect spore after the activated cultivation of the ultimate corruption of stem of noble dendrobium soft rot nosophyte mould (Pythium ultimum), be mixed with 5 * 10
5The spore suspension of/ml mixes with the wheat bran solid substrate and to make the solid vaccination agent.As cultivation matrix, when plantation dendrobium stem seedling, the agent of Dhs96 solid vaccination is inoculated into the root of dendrobium stem seedling with bark.Potted plant seedling is put into heliogreenhouse, and temperature maintenance is at 20 ± 2 ℃.Plantation back water spray is beneficial to the growth of endogenetic fungus.Per three days later on water sprays once.Around the dendrobium stem seedling, spread the agent of pathogenic bacteria solid vaccination during respectively at 3 days behind the inoculated fungi Dhs96 and 30 days.Negative control is the dendrobium stem seedling that only inserts the wheat bran solid substrate when plantation; Positive control is to insert the wheat bran solid substrate in when plantation, and spreads the agent of pathogenic bacteria solid vaccination on every side at the dendrobium stem seedling during respectively at 3 days after the plantation and 30 days.Respectively at inserting the survival rate that pathogenic bacteria is added up seedling after two months.Two kinds of inoculations are handled, and the average survival rate of seedling reaches 78.6%, and the survival rate of seedling is 56.7% in the negative control, do not have the seedling survival in the positive control.This result shows: after infecting soft rot nosophyte, endogenetic fungus Dhs96 can improve the survival rate of dendrobium stem seedling.
Embodiment 2
The biological control of the Herba Dendrobii seedling of Dhs96 soft rot
Endogenetic fungus Dhs96 mixes with the wheat bran solid substrate after the PDA liquid fermentation and culture and makes the solid vaccination agent.Collect spore after the activated cultivation of the ultimate corruption of stem of noble dendrobium soft rot nosophyte mould (Pythium ultimum), be mixed with 1 * 10
5The spore suspension of/ml mixes with the wheat bran solid substrate and to make the solid vaccination agent.As cultivation matrix, when plantation Herba Dendrobii seedling, the agent of Dhs96 solid vaccination is inoculated into the root of Herba Dendrobii seedling with bark.Potted plant seedling is put into heliogreenhouse, and temperature maintenance is at 20 ± 2 ℃.Plantation back water spray is beneficial to the growth of endogenetic fungus.Per three days later on water sprays once.Around the Herba Dendrobii seedling, spread the agent of pathogenic bacteria solid vaccination during respectively at 3 days behind the inoculated fungi Dhs96 and 30 days.Negative control is the Herba Dendrobii seedling that only inserts the wheat bran solid substrate when plantation; Positive control is to insert the wheat bran solid substrate in when plantation, and spreads the agent of pathogenic bacteria solid vaccination on every side at the Herba Dendrobii seedling during respectively at 3 days after the plantation and 30 days.Respectively at inserting the survival rate that pathogenic bacteria is added up seedling after two months.Two kinds of inoculations are handled, and the average survival rate of seedling reaches 66.7%, and the survival rate of seedling is 43.3% in the negative control, do not have the seedling survival in the positive control.This result shows: after infecting soft rot nosophyte, endogenetic fungus Dhs96 can improve the survival rate of Herba Dendrobii seedling.
Embodiment 3
The biological control of the Dendrobium chrysotoxum seedling of Dhs96 soft rot
Endogenetic fungus Dhs96 mixes with the wheat bran solid substrate after the PDA liquid fermentation and culture and makes the solid vaccination agent.Collect spore after the activated cultivation of the ultimate corruption of stem of noble dendrobium soft rot nosophyte mould (Pythium ultimum), be mixed with 5 * 10
4The spore suspension of/ml mixes with the wheat bran solid substrate and to make the solid vaccination agent.As cultivation matrix, when plantation Dendrobium chrysotoxum seedling, the agent of Dhs96 solid vaccination is inoculated into the root of Dendrobium chrysotoxum seedling with bark.Potted plant seedling is put into heliogreenhouse, and temperature maintenance is at 20 ± 2 ℃.Plantation back water spray is beneficial to the growth of endogenetic fungus.Per three days later on water sprays once.Around the Dendrobium chrysotoxum seedling, spread the agent of pathogenic bacteria solid vaccination during respectively at 3 days behind the inoculated fungi Dhs96 and 30 days.Negative control is the Dendrobium chrysotoxum seedling that only inserts the wheat bran solid substrate when plantation; Positive control is to insert the wheat bran solid substrate in when plantation, and spreads the agent of pathogenic bacteria solid vaccination on every side at the Dendrobium chrysotoxum seedling during respectively at 3 days after transplanting and 30 days.Respectively at inserting the survival rate that pathogenic bacteria is added up seedling after two months.Two kinds of inoculations are handled, and the average survival rate of seedling reaches 59.5%, and the survival rate of seedling is 47.3% in the negative control, do not have the seedling survival in the positive control.This result shows: after infecting soft rot nosophyte, endogenetic fungus Dhs96 can improve the survival rate of Dendrobium chrysotoxum seedling.
Embodiment 4
The biological control of the Dens-valve Dendrobium seedling of Dhs96 soft rot
Endogenetic fungus Dhs96 mixes with the wheat bran solid substrate after the PDA liquid fermentation and culture and makes the solid vaccination agent.Collect spore after the activated cultivation of the ultimate corruption of stem of noble dendrobium soft rot nosophyte mould (Pythium ultimum), be mixed with 5 * 10
5The spore suspension of/ml mixes with the wheat bran solid substrate and to make the solid vaccination agent.As cultivation matrix, when plantation Dens-valve Dendrobium seedling, the agent of Dhs96 solid vaccination is inoculated into the root of Dens-valve Dendrobium seedling with bark.Potted plant seedling is put into heliogreenhouse, and temperature maintenance is at 20 ± 2 ℃.Plantation back water spray is beneficial to the growth of endogenetic fungus.Per three days later on water sprays once.Around the Dens-valve Dendrobium seedling, spread the agent of pathogenic bacteria solid vaccination during respectively at 3 days behind the inoculated fungi Dhs96 and 30 days.Negative control is the Dens-valve Dendrobium seedling that only inserts the wheat bran solid substrate when plantation; Positive control is to insert the wheat bran solid substrate in when plantation, and spreads the agent of pathogenic bacteria solid vaccination on every side at the Dens-valve Dendrobium seedling during respectively at 3 days after transplanting and 30 days.Respectively at inserting the survival rate that pathogenic bacteria is added up seedling after two months.Two kinds of inoculations are handled, and the average survival rate of seedling reaches 69.3%, and the survival rate of seedling is 46.7% in the negative control, do not have the seedling survival in the positive control.This result shows: after infecting soft rot nosophyte, endogenetic fungus Dhs96 can improve the survival rate of Dens-valve Dendrobium seedling.
Embodiment 5
The biological control of the Dendrobium chrysanthum seedling of Dhs96 soft rot
Endogenetic fungus Dhs96 mixes with the wheat bran solid substrate after the PDA liquid fermentation and culture and makes the solid vaccination agent.Collect spore after the activated cultivation of the ultimate corruption of stem of noble dendrobium soft rot nosophyte mould (Pythium ultimum), be mixed with 1 * 10
6The spore suspension of/ml mixes with the wheat bran solid substrate and to make the solid vaccination agent.As cultivation matrix, when plantation Dendrobium chrysanthum seedling, the agent of Dhs96 solid vaccination is inoculated into the root of Dendrobium chrysanthum seedling with bark.Potted plant seedling is put into heliogreenhouse, and temperature maintenance is at 20 ± 2 ℃.Plantation back water spray is beneficial to the growth of endogenetic fungus.Per three days later on water sprays once.Around the Dendrobium chrysanthum seedling, spread the agent of pathogenic bacteria solid vaccination during respectively at 3 days behind the inoculated fungi Dhs96 and 30 days.Negative control is the Dendrobium chrysanthum seedling that only inserts the wheat bran solid substrate when plantation; Positive control is to insert the wheat bran solid substrate in when plantation, and spreads the agent of pathogenic bacteria solid vaccination on every side at the Dendrobium chrysanthum seedling during respectively at 3 days after transplanting and 30 days.Respectively at inserting the survival rate that pathogenic bacteria is added up seedling after two months.Two kinds of inoculations are handled, and the average survival rate of seedling reaches 58.7%, and the survival rate of seedling is 40.7% in the negative control, do not have the seedling survival in the positive control.This result shows: after infecting soft rot nosophyte, endogenetic fungus Dhs96 can improve the survival rate of Dendrobium chrysanthum seedling.
Embodiment 6
The biological control of the dendrobium stem seedling of Dfl88 soft rot
Endogenetic fungus Dfl88 makes 1 * 10 after the PDA liquid fermentation and culture
7The spore suspension of/ml is mixed and made into the solid vaccination agent with the wheat bran solid substrate.Collect spore after the activated cultivation of the ultimate corruption of stem of noble dendrobium soft rot nosophyte mould (Pythium ultimum), be mixed with 5 * 10
5The spore suspension of/ml mixes with the wheat bran solid substrate, makes the solid vaccination agent.As cultivation matrix, when plantation dendrobium stem seedling, the agent of Dfl88 solid vaccination is inoculated into the root of dendrobium stem seedling with bark.Potted plant seedling is put into heliogreenhouse, and temperature maintenance is at 20 ± 2 ℃.Plantation back water spray is beneficial to the growth of endogenetic fungus.Per three days later on water sprays once.Around the dendrobium stem seedling, spread the agent of pathogenic bacteria solid vaccination during respectively at 3 days behind the inoculated fungi Dfl88 and 30 days.Negative control is the dendrobium stem seedling that only inserts the wheat bran solid substrate when plantation; Positive control is to insert the wheat bran solid substrate in when plantation, and spreads the agent of pathogenic bacteria solid vaccination on every side at the dendrobium stem seedling during respectively at 3 days after transplanting and 30 days.Respectively at inserting the survival rate that pathogenic bacteria is added up seedling after two months.Two kinds of inoculations are handled, and the average survival rate of seedling reaches 73.3%, and the survival rate of seedling is 56.7% in the negative control, do not have the seedling survival in the positive control.This result shows: after infecting soft rot nosophyte, endogenetic fungus Dfl88 can improve the survival rate of dendrobium stem seedling.
Embodiment 7
The biological control of the Herba Dendrobii seedling of Dfl88 soft rot
Endogenetic fungus Dfl88 makes 5 * 10 after the PDA liquid fermentation and culture
6The spore suspension of/ml is mixed and made into the solid vaccination agent with the wheat bran solid substrate.Collect spore after the activated cultivation of the ultimate corruption of stem of noble dendrobium soft rot nosophyte mould (Pythium ultimum), be mixed with 1 * 10
5The spore suspension of/ml evenly mixes with the wheat bran solid substrate, makes the solid vaccination agent.As cultivation matrix, when plantation Herba Dendrobii seedling, the agent of Dfl88 solid vaccination is inoculated into the root of Herba Dendrobii seedling with bark.Potted plant seedling is put into heliogreenhouse, and temperature maintenance is at 20 ± 2 ℃.Plantation back water spray is beneficial to the growth of endogenetic fungus.Per three days later on water sprays once.Around the Herba Dendrobii seedling, spread the agent of pathogenic bacteria solid vaccination during respectively at 3 days behind the inoculated fungi Dfl88 and 30 days.Negative control is the Herba Dendrobii seedling that only inserts the wheat bran solid substrate when plantation; Positive control is to insert the wheat bran solid substrate in when plantation, and spreads the agent of pathogenic bacteria solid vaccination on every side at the Herba Dendrobii seedling during respectively at 3 days after transplanting and 30 days.Respectively at inserting the survival rate that pathogenic bacteria is added up seedling after two months.Two kinds of inoculations are handled, and the average survival rate of seedling reaches 56.7% (46.7%), and the survival rate of seedling is 43.3% in the negative control, do not have the seedling survival in the positive control.This result shows: after infecting soft rot nosophyte, endogenetic fungus Dfl88 can improve the survival rate of Herba Dendrobii seedling.
Embodiment 8
The biological control of the Dendrobium chrysotoxum seedling of Dfl88 soft rot
Endogenetic fungus Dfl88 makes 5 * 10 after the PDA liquid fermentation and culture
5The spore suspension of/ml is mixed and made into the solid vaccination agent with the wheat bran solid substrate.Collect spore after the activated cultivation of the ultimate corruption of stem of noble dendrobium soft rot nosophyte mould (Pythium ultimum), be mixed with 5 * 10
4The spore suspension of/ml evenly mixes with the wheat bran solid substrate, makes the solid vaccination agent.As cultivation matrix, when plantation Dendrobium chrysotoxum seedling, the agent of Dfl88 solid vaccination is inoculated into the root of Dendrobium chrysotoxum seedling with bark.Potted plant seedling is put into heliogreenhouse, and temperature maintenance is at 20 ± 2 ℃.Plantation back water spray is beneficial to the growth of endogenetic fungus.Per three days later on water sprays once.Around the Dendrobium chrysotoxum seedling, spread the agent of pathogenic bacteria solid vaccination during respectively at 3 days behind the inoculated fungi Dfl88 and 30 days.Negative control is the Dendrobium chrysotoxum seedling that only inserts the wheat bran solid substrate when plantation; Positive control is to insert the wheat bran solid substrate in when plantation, and spreads the agent of pathogenic bacteria solid vaccination on every side at the Dendrobium chrysotoxum seedling during respectively at 3 days after transplanting and 30 days.Respectively at inserting the survival rate that pathogenic bacteria is added up seedling after two months.Two kinds of inoculations are handled, and the average survival rate of seedling reaches 62.3%, and the survival rate of seedling is 47.3% in the negative control, do not have the seedling survival in the positive control.This result shows: after infecting soft rot nosophyte, endogenetic fungus Dfl88 can improve the survival rate of Dendrobium chrysotoxum seedling.
Embodiment 9
The biological control of the Dens-valve Dendrobium seedling of Dfl88 soft rot
Endogenetic fungus Dfl88 makes 1 * 10 after the PDA liquid fermentation and culture
7The spore suspension of/ml is mixed and made into the solid vaccination agent with the wheat bran solid substrate.Collect spore after the activated cultivation of the ultimate corruption of stem of noble dendrobium soft rot nosophyte mould (Pythium ultimum), be mixed with 5 * 10
5The spore suspension of/ml evenly mixes with the wheat bran solid substrate, makes the solid vaccination agent.As cultivation matrix, when plantation Dens-valve Dendrobium seedling, the agent of Dfl88 solid vaccination is inoculated into the root of Dens-valve Dendrobium seedling with bark.Potted plant seedling is put into heliogreenhouse, and temperature maintenance is at 20 ± 2 ℃.Plantation back water spray is beneficial to the growth of endogenetic fungus.Per three days later on water sprays once.Around the Dens-valve Dendrobium seedling, spread the agent of pathogenic bacteria solid vaccination during respectively at 3 days behind the inoculated fungi Dfl88 and 30 days.Negative control is the Dens-valve Dendrobium seedling that only inserts the wheat bran solid substrate when plantation; Positive control is to insert the wheat bran solid substrate in when plantation, and spreads the agent of pathogenic bacteria solid vaccination on every side at the Dens-valve Dendrobium seedling during respectively at 3 days after transplanting and 30 days.Respectively at inserting the survival rate that pathogenic bacteria is added up seedling after two months.Two kinds of inoculations are handled, and the average survival rate of seedling reaches 60.3%, and the survival rate of seedling is 46.7% in the negative control, do not have the seedling survival in the positive control.This result shows: after infecting soft rot nosophyte, endogenetic fungus Dfl88 can improve the survival rate of Dens-valve Dendrobium seedling.
Embodiment 10
The biological control of the Dendrobium chrysanthum seedling of Dfl88 soft rot
Endogenetic fungus Dfl88 makes 5 * 10 after the PDA liquid fermentation and culture
7The spore suspension of/ml is mixed and made into the solid vaccination agent with the wheat bran solid substrate.Collect spore after the activated cultivation of the ultimate corruption of stem of noble dendrobium soft rot nosophyte mould (Pythium ultimum), be mixed with 1 * 10
6The spore suspension of/ml evenly mixes with the wheat bran solid substrate, makes the solid vaccination agent.As cultivation matrix, when plantation Dendrobium chrysanthum seedling, the agent of Dfl88 solid vaccination is inoculated into the root of Dendrobium chrysanthum seedling with bark.Potted plant seedling is put into heliogreenhouse, and temperature maintenance is at 20 ± 2 ℃.Plantation back water spray is beneficial to the growth of endogenetic fungus.Per three days later on water sprays once.Around the Dendrobium chrysanthum seedling, spread the agent of pathogenic bacteria solid vaccination during respectively at 3 days behind the inoculated fungi Dfl88 and 30 days.Negative control is the Dendrobium chrysanthum seedling that only inserts the wheat bran solid substrate when plantation; Positive control is to insert the wheat bran solid substrate in when plantation, and spreads the agent of pathogenic bacteria solid vaccination on every side at the Dendrobium chrysanthum seedling during respectively at 3 days after transplanting and 30 days.Respectively at inserting the survival rate that pathogenic bacteria is added up seedling after two months.Two kinds of inoculations are handled, and the average survival rate of seedling reaches 53.3%, and the survival rate of seedling is 40.7% in the negative control, do not have the seedling survival in the positive control.This result shows: after infecting soft rot nosophyte, endogenetic fungus Dfl88 can improve the survival rate of Dendrobium chrysanthum seedling.
Claims (2)
1. the 1 strain endogenetic fungal bacterial strain that the orchid family Dendrobium plant soft rot is had preventive and therapeutic effect, it is the Rhizopus fungi Dhs96 (Rhizopus sp.) of CGMCC No.3786 for deposit number.
2. 1 strain endogenetic fungal bacterial strain as claimed in claim 1 is in the purposes of preventing and treating aspect the soft rot, it is characterized in that seedling dish, flowerpot or the seedbed that can be used for Herba Dendrobii and dendrobium stem grow seedlings, and also can be used for the field production of Herba Dendrobii and dendrobium stem.
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| CN104521469A (en) * | 2014-12-03 | 2015-04-22 | 柳州市泓吉农业科技有限公司 | Dendrobium soft rot controlling methods |
| CN104593273A (en) * | 2015-01-28 | 2015-05-06 | 上海中医药大学 | Dendrobium officinale endophytic fungi strain NT43J06 and application thereof |
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