CN110169421A - Application of the Metarhizium anisopliae in preparation prevention and treatment plant root-knot nematodes drug - Google Patents
Application of the Metarhizium anisopliae in preparation prevention and treatment plant root-knot nematodes drug Download PDFInfo
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- CN110169421A CN110169421A CN201910417132.7A CN201910417132A CN110169421A CN 110169421 A CN110169421 A CN 110169421A CN 201910417132 A CN201910417132 A CN 201910417132A CN 110169421 A CN110169421 A CN 110169421A
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- A—HUMAN NECESSITIES
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- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
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Abstract
The invention belongs to technical field of plant disease biological control, and in particular to one plant of Metarhizium anisopliae is in preparation prevention and treatment plant root-knot nematodes drug and promotes the application in plant strain growth.The Metarhizium anisopliae that the present invention uses is Metarhizium anisopliae IBCCM321.93, which finds that plant root-knot nematode can be prevented and treated for the first time, and has growth-promoting functions to plant.Line eggs are after the processing of 50% fermentating liquid filtrate of the bacterium, compared with the control, the inhibiting rate of the hatching of nematode reaches 95.73%, Meloidogyne incognita second instar larvae is after the fermentating liquid filtrate of 25% concentration is handled 2 days, nematode lethality reaches 100%, with extraordinary nematode inhibitory effect, it is suitable for large-scale promotion.
Description
Technical field
The invention belongs to technical field of plant disease biological control, and in particular to Metarhizium anisopliae prevents and treats plant in preparation
Application in root-knot nematode medicine.
Technical background
Plant root-knot nematode is a kind of universal plant pathogeny line insect, can lead to plant with the root of parasitic plant
Nutrition and moisture absorption are affected, and cause plant undergrowth.In addition, the harm of root-knot nematode can also aggravate other cause of diseases
Harm of the object to plant.The various vegetables such as Curcurbitaceae, Solanaceae and beans often have root knot nematode disease, can generally cause the underproduction
20%-30% reaches 75% or more when serious.With being widely applied for the establishment planting technologies such as China's vinyl house, root knot
Causing harm for nematode gradually aggravates.The prevention and treatment of plant root-knot nematode is still based on chemical pesticide at present.However it some common kills
Line agent such as Aldicarb etc. has prevention and treatment line due to characteristics such as high poison, high residues, according to " the pesticide management rules " newly revised are a variety of
The pesticide (such as Aldicarb) of parasitosis evil ability, will be disabled in China successively.Furthermore, it has been found that nematode is to chemistry such as fosthiazates
There is drug resistance in nematode killing agent.Therefore, biological control is carried out to root knot nematode disease using beneficial microbe, it is a kind of to substitute
Safely and effectively prevention and treatment prevention and treatment nematodiasis strategy.Although having had now been found that some pairs of root-knot nematodes have biological control effect
For example pale purple purple spore bacterium of fungi etc., but different microbial action mechanism and the adaptability of environment is different from, for different
Inevitably there is certain limitation in production application demand, therefore it is particularly important to screen more more effective biocontrol microorganisms.The green deadlock of chafer
Bacterium (Metarhizium anisopliae) is a kind of important insect pathogenic fungus, and the biological control of insect is widely applied.
The invention discloses the methods prevented and treated using Metarhizium anisopliae spore preparation root-knot nematode.In the present invention
Before, Guizhou green muscardine fungus (Metarhizium guizhouense) be it is reported uniquely can parasitic root-knot nematode green deadlock
Bacterium.And the Metarhizium anisopliae in the present invention from Guizhou Metarhizium in different kinds, Guizhou green muscardine fungus can be with parasitic nematode
Ovum and second instar larvae, and the Metarhizium anisopliae in the present invention then can be with the ovum of parasitic nematode and mature female adult, and it ferments
Liquid can inhibit the hatching of line eggs and kill second instar larvae.
While Metarhizium anisopliae of the invention finds to can be used for preventing and treating plant root-knot nematode for the first time, and there is rush to plant
Raw effect.
Summary of the invention
The object of the present invention is to provide Metarhizium anisopliae IBCCM321.93 to prevent and treat plant root-knot nematodes in preparation
Application in drug.
It is another object of the present invention to provide Metarhizium anisopliae IBCCM321.93 in preparation while to prevent and treat plant
Application in the preparation of root knot nematode disease and promotion plant growth.
In order to achieve the above object, the present invention takes following technical measures:
Application of the Metarhizium anisopliae IBCCM321.93 in preparation prevention and treatment plant root-knot nematodes drug, including with gold
Tortoise green muscardine fungus IBCCM321.93 spore or its fermentation liquid are the drug that effective component is prepared into prevention and treatment root knot nematode disease.
As long as in addition, by Metarhizium anisopliae IBCCM321.93 spore or its fermentation liquid be used to prevent and treat root knot nematode disease or/
With promotion plant growth, protection scope of the present invention is belonged to.
Compared with prior art, the invention has the following advantages that
1. present invention firstly discovers that Metarhizium anisopliae IBCCM321.93 can with the ovum of parasitic nematode and mature female adult,
And its fermentation liquid can inhibit the hatching of line eggs and kill second instar larvae, therefore can be used for inhibiting the numerous of plant root-knot nematode
It grows.
2. can be used as growth-promoting present invention firstly discovers that Metarhizium anisopliae IBCCM321.93 has growth-promoting functions to plant
Agent uses.
Detailed description of the invention
Fig. 1 is that Metarhizium anisopliae bacterial strain IBCCM321.93 illustrates the parasitics of root-knot nematode worm's ovum and mature female adult
Figure；
Wherein: A and B is respectively to handle 5 days as the line eggs of control and mature female adult using clear water；C and D is respectively golden
Line eggs and mature female adult after tortoise muscardine spore is 5 days parasitic.
Fig. 2 is the fermentation liquid of Metarhizium anisopliae on nematode egg hatching and on the active influence schematic diagram of two age nematodes；
Wherein: A is that the ovum of nematode can normally ovum be incubated in Cha Shi fluid nutrient medium, occurs a large amount of second instar larvae after 8 days；
Scheming B is with after ovum 8 days of fermentation liquor treatment nematode of 95% Metarhizium anisopliae, and the hatching of ovum is suppressed；C is with nematode
Second instar larvae still large number of viable after 2 days in Cha Shi fluid nutrient medium；D is the second instar larvae of nematode green with 95% chafer
In the fermentation liquid of stiff bacterium after 2 days, nematode mortality, polypide is in stiff shape.
Fig. 3 is prevention and treatment schematic diagram of the Metarhizium anisopliae to tomato root-knot eelworm disease；
Wherein: after A is prevents and treats tomato root-knot eelworm disease with the Metarhizium anisopliae spore preparation of various concentration, tomato
Plant strain growth situation；After B is prevents and treats tomato root-knot eelworm disease with the Metarhizium anisopliae spore preparation of various concentration, tomato
Root growth situation, arrow are directed toward the root knot of root；C is to prevent and treat tomato root with the Metarhizium anisopliae spore preparation of various concentration
After Root-knot evil, the plant height and root long of tomato.
Fig. 4 is Metarhizium anisopliae to tomato growth facilitation schematic diagram；
Wherein: A be various concentration Metarhizium anisopliae spore preparation handle tomato after, the aerial part of tomato plant and
Extent of the root system；B is the aerial part and root of tomato plant after the Metarhizium anisopliae spore preparation of various concentration handles tomato
The fresh weight of system.
Specific embodiment
Technical solution of the present invention is if not otherwise specified the conventional scheme of this field, the reagent or material,
If not otherwise specified, commercial channel is derived from.Nematode used in the embodiment of the present invention is Meloidogyne incognita.
Embodiment 1:
Parasitics of the Metarhizium anisopliae spore to root-knot nematode:
Metarhizium anisopliae IBCCM321.93 (bacterial strain deposit number is CGMCC No.0853) is in PDA culture medium in 28
DEG C culture 2 days, then with the aseptic water washing containing 0.05% tween and spore is collected, and be filtered to remove in spore with lens wiping paper
Mycelia.The preparation of PDA carries out according to a conventional method, and every 200g potato cuts fritter and is cooked, and filtrate is collected by filtration and the Portugal 20g is added
Sugared, moisturizing to total volume is that 1000ml is PDB culture medium；Then it is dispensed, is added 1.5g's in every 100ml culture medium
Agar is prepared into PDA culture medium after high-temp steam sterilizing.
The ovum of nematode sterilizes 2 minutes in 1% sodium hypochlorite, is then hatched in 25 DEG C to obtain second instar larvae.Line
The second instar larvae of worm and mature female adult then carry out surface sterilization with 0.5% sodium hypochlorite.
Glassine paper is cut into 1cm2Fritter, be laid in after sterilizing diameter 9cm water agar preparation plate on, Mei Gepei
Feeding ware can tile about 10 pieces of glassine papers.The spore of the Metarhizium anisopliae of 10 μ l various concentrations is dripped on every piece of glassine paper.With 10 μ
The water droplet of l is on glassine paper as control.After 28 DEG C of culture 12h, be inoculated on glassine paper 50 nematodes ovum or 50 two
Instar larvae or 50 mature female adults.Green muscardine fungus is observed to the parasitic ability of nematode, the results showed that green muscardine fungus has the ovum of nematode
There is parasitic ability, and as the extension parasitism efficiency of time is continuously increased (Fig. 1).Nematode after counting different number of days hatches feelings
Condition shows ovum of the Metarhizium anisopliae by parasitic nematode, has good inhibitory effect to the hatching of nematode, wherein 1 × 108
The spore of a/ml can achieve 70% or more to the inhibiting rate of nematode egg hatching after handling 8 days.Only a small number of second instar larvaes are golden
Tortoise green muscardine fungus is parasitic, but has Spore adhesion around the polypide of most of nematodes, and the stiff mobility decline of polypide.This
Outside, Metarhizium anisopliae can be with the mature female adult (Fig. 1) of parasitic nematode, and parasitic rate is greater than 60%, so as to cause the breeding of nematode
Ability decline, can effectively reduce source of infection again.
Inhibiting rate of the 1 Metarhizium anisopliae spore of table to nematode egg hatching
Embodiment 2:
Poisoning of the Metarhizium anisopliae fermentating liquid filtrate to root-knot nematode:
In czapek's medium (sodium nitrate 3g, dipotassium hydrogen phosphate 1g, magnesium sulfate 0.5g, potassium chloride 0.5g, the sulfuric acid of 100ml
Ferrous 0.01g, sucrose 30g, distilled water 1000ml, pH6-6.5) in inoculation 2 pieces of mycelia block of Metarhizium anisopliae diameter 7mm,
Then in 160rpm, 28 DEG C are centrifuged off mycelium after shaken cultivation 2 weeks, while with 0.22 μm of diameter of membrane filtration with abundant
Remove fungal tissue.The fermentating liquid filtrate of green muscardine fungus is diluted with czapek's medium, until the final concentration of fermentation liquid is respectively
95%, 50%, 25% and 10% (v/v).The ovum of nematode or second instar larvae are carried out surface sterilization to be placed in water, and adjust to
Contain 100 ovum or 100 second instar larvaes in every 50 μ l solution.Take the line eggs of 50 μ l or second instar larvae be easily positioned at containing
In 24 porocyte culture plates of 950 μ l various concentration ferment filtrates.Ferment filtrate is substituted as control using czapek's medium.It will be thin
Born of the same parents' culture plate is placed in 25 DEG C, and observes influence of the fermentation liquid to nematode of various concentration.
As a result, it has been found that the fermentating liquid filtrate of Metarhizium anisopliae has apparent inhibition to the hatching of the ovum of nematode, and inhibit
Effect increases (Fig. 2) with the increase of ferment filtrate concentration.Wherein 50% fermentating liquid filtrate and Cha Shi Liquid Culture base phase
Than the inhibiting rate of the hatching of nematode reaches 95.73% (table 2) in control.
In addition, the fermentating liquid filtrate of Metarhizium anisopliae has stronger poisoning property (table 3) to the second instar larvae of nematode.?
After handling 2 days in the fermentating liquid filtrate of 25% concentration, nematode lethality reaches 100%.
Influence of the Metarhizium anisopliae fermentating liquid filtrate of 2 various concentration of table to nematode egg hatching
Influence of the Metarhizium anisopliae fermentating liquid filtrate of 3 various concentration of table to nematode second instar larvae
Embodiment 3: Metarhizium anisopliae spore preparation mitigates nematode to the parasitic rate of tomato
Metarhizium anisopliae spore preparation can be prepared using two different modes: one is trained with PDA solid plate
Metarhizium anisopliae is supported after 15 days, spore is collected with the aseptic water washing containing 0.05% tween, is then filtered, obtained with lens wiping paper
Obtain Metarhizium anisopliae spore liquid.Another kind is every 100ml PDB inoculation of medium 1 × 108Metarhizium anisopliae spore, and
28 DEG C shaken cultivation 21 days, be then centrifuged for remove supernatant liquid collect spore, by spore with sterile water dilute obtain chafer
The spore preparation of green muscardine fungus.
It is cultivated in sterile Nutrition Soil tomato 2 weeks, the spore liquid of Metarhizium anisopliae is then applied in tomato root week
Spore concentration setting 1 × 10 in the soil enclosed, in every 100 grams of soil6、1×107With 1 × 108Three kinds of different processing, simultaneously
It is inoculated with the ovum of 1000 nematodes.To be only inoculated with line eggs, without being inoculated with the tomato of Metarhizium anisopliae spore as control.
The result shows that treated that tomato root combined symptoms shape is substantially reduced for Metarhizium anisopliae.Wherein concentration is 1 × 108Spore
After subprocessing, the line eggs number in each reduces about 59%, and second instar larvae number reduces by about 61% (table in every 100g soil
4).Furthermore height, root long and the fresh weight of Metarhizium anisopliae treated tomato plant are obviously higher than pair for being only inoculated with nematode
According to group (Fig. 3).Therefore, the spore preparation of Metarhizium anisopliae has good prevention and control to act on crop root knot nematode disease.
Control efficiency of the 4 Metarhizium anisopliae spore preparation of table to tomato root-knot eelworm
Embodiment 4:
Metarhizium anisopliae IBCCM321.93 is promoting the application in plant growth:
Metarhizium anisopliae is collected after PDA solid plate culture 15 days with the aseptic water washing containing 0.05% tween
Spore.Then it is filtered with lens wiping paper, collects the spore of Metarhizium anisopliae.After tomato seedling 7 days, growing state basic one is selected
The healthy plant of cause is transplanted.After tomato is transplanted 7 days, the spore of green muscardine fungus is inoculated in the soil around tomato root, often
About 5 × 10 are inoculated in 100g soil8Or 1 × 109Spore.Using the tomato of sterile water process as control.In inoculation green muscardine fungus 4 weeks
Compare the growing state (Fig. 4) of tomato afterwards.Compared with the control, 5 × 108Spore processing after, the plant height of tomato aerial part increases
Add about 32.1%, fresh weight increase about 44.4%；Root long increase about 34.2%, root increases about 68% again.With the increasing of spore concentration
Growth-promoting effect is added to enhance, 1 × 109Spore processing after tomato aerial part plant height increase about 72%, about 2 times of fresh weight increase；
Root long increase about 51.4%, root increases about 94.4% again.Therefore, root system and ground of the spore liquid of Metarhizium anisopliae to tomato
Growth has significant facilitation, can be used as growth-promoting agent use.
Claims (4)
1. application of the Metarhizium anisopliae IBCCM321.93 in preparation prevention and treatment plant root-knot nematodes drug.
2. Metarhizium anisopliae IBCCM321.93 fermentating liquid filtrate inhibits plant root-knot nematode larva or plant roots tie lines in preparation
Application in the drug of egg hatch.
3. Metarhizium anisopliae IBCCM321.93 spore inhibits plant root-knot nematode female adult or plant root-knot nematode ovum to incubate in preparation
Application in the drug of change.
4. Metarhizium anisopliae IBCCM321.93 is preparing while preventing and treating plant root-knot nematodes and promoting the preparation of plant growth
In application.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN112493256A (en) * | 2020-12-03 | 2021-03-16 | 重庆谷百奥生物研究院有限公司 | Metarhizium anisopliae preparation for promoting tomato growth and preventing and treating tomato anthracnose |
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| CN112493256A (en) * | 2020-12-03 | 2021-03-16 | 重庆谷百奥生物研究院有限公司 | Metarhizium anisopliae preparation for promoting tomato growth and preventing and treating tomato anthracnose |
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Application publication date: 20190827 |