CN104877913B - A kind of AMF+DSE combination microbial inoculums and its application in the preparation for promoting cucumber growth and anti-root-knot nematode is prepared - Google Patents
A kind of AMF+DSE combination microbial inoculums and its application in the preparation for promoting cucumber growth and anti-root-knot nematode is prepared Download PDFInfo
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Abstract
The invention provides a kind of AMF+DSE combination microbial inoculums and its preparing the application in promoting preparation of the cucumber growth with anti-root-knot nematode, the present invention sets different AMF+DSE combinations to carry out inoculation test to cucumber, and infection rate, the clump branch for analyzing different AMF+DSE combinations the influence to Plant Growth of Cucumber and its root-knot nematode disease such as raw rate, clump branch type.Determine that the AMF+DSE of optimal anti-Meloidogyne incognita is combined as G.m+S.a+A.l+Ph.l by result of the test, the plant height of this combined treatment, stem are thick, overground part dry weight, weight of root system and single plant yield highest and disease index is minimum, the best of breed microbial inoculum, which has, promotes the effect of cucumber growth is with improving cucumber plant anti-root-knot nematode, and the present invention provides foundation further to explore AMF and DSE interaction and mechanism, laid a good foundation.
Description
Technical field
The invention belongs to technical field of plant protection, and in particular to a kind of AMF+DSE combines microbial inoculum and its promoted preparing
Cucumber growth and the application in the preparation of anti-root-knot nematode.
Background technology
AMF (arbuscular mycorrhizal fungi, AMF) is important in terrestrial ecosystems
Member, mutualistic symbiosis body can be established with root system of plant.AMF can improve the mineral nutrition of plant, enhancing disease resistance of plant, improve and plant
Resistance of the thing in extreme environment, contribute to plant to resist the infringement of pathogen, and improve soil granular structure, promote crop
Stable high yield, high-quality and efficient.AMF infects the characteristic of plant root tip, shows that the ecological niche of the fungi is identical with the soil-borne disease original,
This just creates advantageous condition for antagonism pathogen.What is more important is formed by being extended in root outside root in soil
Huge mycelia network, and the mycorhiza networking being consequently formed plays physiological ecological function (such as water and nutrient absorbs) for it and establishes
Solid Ecology basis is determined.AMF is in the side such as agricultural sustainable development, ecological protection, environment and food security
Face also has significance.
Although the AMF researchs in China are started late, start from the end of the seventies in last century, fast development is then entered after 10 years
Stage.Into after this century, China AMF studies the development for remaining fast and stable.For example, AMF classification, resource and biology are more
130 kinds of the category of report 7 in terms of sample, including Paraglomus (Glomus), Scutellospora category (Scutellospora), no stalk capsule
Mould category (Acaulospora), Log outline (Archaeospora), Gigaspora (Gigaspora), Paraglomus
(Paraglomus), Entrophospora (Entrophospora), and constantly have new species and new records kind report.Each state all separates
To a variety of AMF species not belonged to together, Paraglomus occurrence frequency highest, especially G.clarum, G.etunicatum,
G.geosporum, G.intraradices, G.mosseae, G.sinuosum are the most extensive in distribution on global.Simultaneously in physiology
Effect and ecology etc. have carried out systematic development work；And mycorrhizal research technical elements, also establish measure AMF Inoculum potentials
New method；Propose the theory of AMF induction disease resistance of plant.At present, AMF culture presevation storehouse is had been set up in the world, is received
Collection world's AMF resources, breeding, culture presevation and exchange, the preservation that the measures such as strain carry out specification strain is provided for various countries researcher
Work.In summary, the research of AMF species diversities achieves huge progress, diversity and the Study on Resources and has obtained prosperity with protection
Country and the attention of developing country.
Dark color have colonized extensively every endogenetic fungus (dark septate endophytes, DSE) it is thin in root system of plant cortex
Intracellular or space between cells, formed dark color have every mycelia and Microsclerotia.DSE can generally promote plant growth, improve nutrient and inhale
Receive, improve disease resistance of plant, the ability of unsuitable environmental condition is resisted in enhancing.DSE and AMF can infect plant simultaneously under natural environment
Thing root system.DSE cooperates with mutual-aid relation with other edaphons, and DSE can form fine and close mycelia between Gen Nei or root with AMF
Bridge, expand root system Nutrient Absorption scope, effectively carry out mass exchange and information transmission, collectively promote the work of plant rhizosphere microorganism
It is dynamic, improve plant rhizosphere Soil Micro-environment, so as to promote growing for plant.
Research report about DSE was had begun at the beginning of the last century, and it is true in mycorhiza to come from researcher to DSE understanding in early days
The mycorrhizal fungi in some non-traditional meanings found in bacterium research, is officially named through exploration after a while
DSE.DSE is widely distributed, is almost found in all habitats of the earth, but except a few DSE classification position
Clearly outer, the classification position of most of monoids is still not clear.It is still at an early stage to the understanding of this grouping fungal at present.
Cucumber (Cucumis sativus) occupies important as a kind of global important vegetable in daily life
Status.But the continuous cropping obstacle in production causes the morbidity and mortality of soil-borne disease of cucumber, especially root-knot nematode disease
Constantly rise, cause to drop in production over a large area, on cucumber cultivation influence it is very big, be in current cucumber production more distinct issues it
One.
The content of the invention
Promote cucumber growth and the system of anti-root-knot nematode the invention provides a kind of AMF+DSE combination microbial inoculums and its preparing
Application in agent, it can be used for promoting plant to grow and improve the disease-resistant of plant using combination microbial inoculum of the present invention
Property.
To achieve the above object, the present invention is achieved using following technical scheme：
The invention provides Phoma sp F2-3-10 bacterial strains, its Classification And Nomenclature is Phoma leveillei, is preserved in China
Microbiological Culture Collection administration committee common micro-organisms center, its deposit number are：CGMCC No.10482.
Present invention also offers a kind of AMF+DSE to combine microbial inoculum, and the combination microbial inoculum includes AMF microbial inoculums and DSE microbial inoculums, institute
It is microbial inoculum made of Phoma sp F2-3-10 bacterial strains that deposit number is CGMCC No.10482 to state DSE microbial inoculums.
The AMF microbial inoculums are：One or more in Glomus mosseae, global Scutellospora or light wall Acaulospora.
Invention further provides described AMF+DSE combination microbial inoculums answering in the preparation for promoting plant growth is prepared
With.
The combination microbial inoculum can improve the plant height of cucumber, overground part dry weight, underground part dry weight and single plant yield.
Invention further provides described AMF+DSE combination microbial inoculums answering in the preparation for preparing anti-root-knot nematode
With.
The root-knot nematode is Meloidogyne incognita (Meloidogyne incognita).
The combination microbial inoculum can significantly reduce the female borer population of second instar larvae number, nematode and ovum number in cucumber root.
Compared with prior art, advantages of the present invention and have the technical effect that：The present invention uses basin under the conditions of greenhouse production
Cultivating method, different AMF+DSE combinations are set to carry out inoculation test to cucumber, measure analyzes infecting for different AMF+DSE combinations
Rate, clump branch the influence to Plant Growth of Cucumber and cucumber root-knot nematode such as raw rate, clump branch type, are determined most by result of the test
The AMF+DSE of good anti-Meloidogyne incognita is combined as G.m+S.a+A.l+Ph.l, the cucumber plant height of the combined treatment, stem
Rugosity, overground part dry weight, weight of root system and single plant yield highest and disease index is minimum, respectively 130.3cm, 0.95cm,
20.7g, 2.9g, 562.9g and 15.3.Show that it is G.m+ to promote Plant Growth of Cucumber, the best of breed of anti-Meloidogyne incognita
S.a+A.l+Ph.l.The effect of this best of breed microbial inoculum has the growth for promoting cucumber and improves anti-root-knot nematode, the present invention is
The mutual effect for further exploring AMF and DSE is laid a good foundation with mechanism of action.
Embodiment
Technical scheme is further described in detail with reference to specific embodiment.
Embodiment 1
1. test material
Cucumber (Cucumis sativus, kind are " Jinyou No.35 "), by Tianjin Kerun Agricultural Science ＆ Technology Co., Ltd.
There is provided, can also use other common cucumber varieties；Meloidogyne incognita (Meloidogyne incognita, Mi) is by Qingdao agriculture
Sparetime university learns Plant Protection cinemas worm laboratory and provided；DSE is Phoma sp (Phoma leveillei) F2-3-10 bacterial strains (abbreviation
Ph.l), provided by biological technology of mycorrhiza research institute of Qingdao Agricultural University；AMF is Glomus mosseae Glomus mosseae (preservations
Numbering Gm32), Glomus versiforme Glomus versiforme (deposit number Gv39), light wall Acaulospora Acaulospora
Laevis (deposit number Al57), global Scutellospora Scutellospora aurigloba (deposit number Sa97), preservation
Numbering Gm32, deposit number Gv39, deposit number Al57 and deposit number Sa97 are by Qingdao Agricultural University's mycorhiza biology skill
Art research institute provides, and the AMF in the present invention can also use other AMF bacterial strains of the prior art.
AMF spores to be stored in the root segment and its quartz sand culture matrix of clover are used as inoculum；Cucumber is potted plant
Matrix be sand than 2: 1, total nitrogen content 0.1%, available phosphorus contents 16.1mg/kg, quick-acting potassium content 102.6mg/kg, organic matter
Content 7.9g/kg.
2. experimental design
The method that present invention experiment uses greenhouse production, if 28 processing, respectively Meloidogyne incognita
(Mi)、Glomus mosseae(G.m)、Glomus versiforme(G.v)、Acaulospora laevis(A.l)、
Scutellospora aurigloba (S.a), Phoma leveillei (Ph.l), G.m+Ph.l, S.a+Ph.l, G.v+
Ph.l、A.l+Ph.l、G.m+S.a+Ph.l、G.m+G.v+Ph.l、G.m+A.l+Ph.l、G.m+S.a+G.v+Ph.l、G.m+G.v
+A.l+Ph.l、G.v+S.a+A.l+Ph.l、G.m+S.a+A.l+Ph.l、G.m+Ph.l+Mi、S.a+Ph.l+Mi、G.v+Ph.l+
Mi、A.l+Ph.l+Mi、G.m+S.a+Ph.l+Mi、G.m+G.v+Ph.l+Mi、G.m+A.l+Ph.l+Mi、G.m+S.a+G.v+
Ph.l+Mi, G.m+G.v+A.l+Ph.l+Mi, G.v+S.a+A.l+Ph.l+Mi, G.m+S.a+A.l+Ph.l+Mi and it is not inoculated with pair
According to (CK), random alignment, each processing is repeated 5 times.
The Phoma sp (Phoma leveillei) F2-3-10 bacterial strains are subjected to culture presevation, the title of depositary institution：
China Committee for Culture Collection of Microorganisms's common micro-organisms center (CGMCC)；Address：BeiChen West Road, Chaoyang District, BeiJing City 1
Number institute 3, Institute of Microorganism, Academia Sinica.Phoma leveillei preservation date：On March 31st, 2015；Preservation is compiled
Number：CGMCC No.10482.
3. inoculation and cultivation management
Matrix autoclaving (121 DEG C, 1h) loads the flowerpot (17cm × 20cm) of sterilized mistake afterwards；CK is then added and is inoculated with
The sterilizing sand of thing same volume and inoculum filtrate；Cucumber seeds first soak 10min with 0.10% mercuric chloride solution, then with clear
Water is cleaned, then with 60-70 DEG C of water seed soaking 10-15min, after Seed soaking 12h, the vernalization in 25-28 DEG C of insulating box,
Sowed after plumule exposes.Two band bud cucumber seeds are sown into flowerpot, then earthing lid kind.Each flowerpot connects prior to seeding
Kind AMF Mixed inocula 20mL, when cucumber emergence grows to 3-5 leaves, inoculation Phoma leveillei (Ph.l) bacterium solution
50ml/ basins (access mycelia block, in 25 DEG C of rotating speeds in 100r/min shaking tables in potato dextrose broth (PDB)
After shaken cultivation 14d, hypha body is collected by filtration with sterile gauze, after being rinsed for several times with aqua sterilisa, hypha body is smashed and is configured to 5
×105CFU/mL bacterium solution is standby), when cucumber seedling growth to 7~9 leaf, be inoculated with Mi ovum, inoculum concentration be about 1500 ovum/
Strain basin, using root-pouring method, is uniformly poured into around cucumber root.Need to apply fertilizer according to the upgrowth situation of the fertility of matrix and plant
Watering etc., other use unified Routine Management.
4. data analysing method
Data analysis is carried out using 2003 editions statistical softwares of DPS 7.05 and Microsoft Excel.
5. experimental result
(1) it is inoculated with influence of the Meloidogyne incognita (Mi) to cucumber AMF and DSE infection
AMF infection rates (%)=∑ (0% × root segment+10% × root segment of number number+...+100% × root segment number) × 100%/
Total root segment number；
A type clump branches raw rate (%)=∑ (0% × root segment+10% × root segment of number number+...+100% × root segment number) ×
100%/total root segment number；
P-type clump branch raw rate (%)=∑ (0% × root segment+10% × root segment of number number+...+100% × root segment number) ×
100%/total root segment number；
P/A is that p-type clump branch the ratio that raw rate (%) raw rate (%) with A type clump branches；
DSE infection rates (%) determine according to McGonigle (1990) method.
As shown in table 1, AMF infection rates, the A type clump branch infection rates that G.m+S.a+A.l+DSE+Mi is handled after inoculation nematode are equal
It is significantly higher than other processing, DSE infection rates, the p-type clump branch infection rate of G.m+G.v+A.l+DSE+Mi processing are significantly higher than other
Processing.
Table 1 is inoculated with Meloidogyne incognita (Mi) cucumber AMF, DSE afterwards infection
Note：With after column data, different lowercase letter indication differences are notable (α=0.05) in table
(2) different AMF+DSE combine the influence to Meloidogyne incognita (Mi) Propagule number
Egg sac number on root：Often processing randomly selects the root system of 3 plants of cucumber, is cut into 1cm or so root segments, claims 1g at random,
Record egg sac number.
Single egg capsule amount containing ovum：About 10 egg capsules are put into 5% sodium hypochlorite and dissolve 2-3min, then record is each
The ovum number of egg capsule.
Experimental result is as shown in table 2, and female borer population, egg sac number and amount containing ovum are aobvious in the cucumber root of AMF+DSE combined treatments
The processing less than single inoculation nematode is write, especially on egg sac number and figureofmerit containing ovum.Meanwhile there is also difference between each processing
Different, second instar larvae number in the root of G.m+S.a+A.l+Ph.l+Mi combined treatments, female borer population, egg sac number and amount containing ovum are aobvious in root
Work is less than other combined treatments, i.e. the Meloidogyne incognita Propagule number of the combined treatment is minimum in each processing,
Show to a certain extent and suppression nematode is best.
Table 2 is inoculated with influences of the AMF+DSE to Meloidogyne incognita Propagule number
Note：With after column data, different lowercase letter indication differences are notable (α=0.05) in table
(3) different AMF+DSE combine the influence to Cucumber root-knot nematode disease
The grade scale of root-knot nematode occurrence injury, it is specific as follows using 0-6 level grade scales：
6 grades-the root quantity that forms root knot accounts for the 100% of root system sum, and root knot, which is assembled, to be formed obvious root system and expand, nothing
The system of fibrous root, the significantly rotten symptom of root system tool；
5 grades-the root quantity that forms root knot accounts for the 70%-90% of root system sum, and root knot assembles that to form obvious root system swollen
Greatly, only a small number of systems of fibrous root；
4 grades-form the 50%-70% that root knot root quantity accounts for root system sum, most of root knot agglomerates into obvious root
System expands (30%-70%)；
3 grades-formation root knot root quantity accounts for 20%-50% of root system sum or so, and part root knot agglomerates into significantly
Root system expands；
2 grades-formation root knot root quantity accounts for 10%-20% of root system sum or so, root knot median size, occurs brighter
Aobvious fibrous root group；
1 grade-form root knot root quantity accounts for root system sum less than 10%, root knot is somewhat expanded, and has some fibrous roots
The formation of group；
0 grade-do not form root knot；
0.5 grade-form root knot root quantity accounts for root system sum less than 10%, root knot is smaller, and the development of the system of fibrous root is almost
It is not affected.Below equation calculates disease index, the incidence of disease, preventive effect respectively.
Disease index=∑ (each sick series × at different levels strain number) × 100/ (highest disease series × strain number always investigated)
The incidence of disease (%)=[disease plant number/investigation total strain number] × 100%
Relative control effect (%)=[(disease index of single inoculation root-knot nematode-inoculation AMF+DSE+Mi disease index)/
The disease index of single inoculation root-knot nematode] × 100%
Experimental result is as shown in table 3, is inoculated with the Cucumber root-knot nematode disease state of an illness of S.a+Ph.l and G.v+Ph.l combined treatments
Index reaches 66.7, is not significantly different with compareing, while preventive effect is relatively low；And the incidence of disease of G.m+S.a+A.l+Ph.l combinations is only
Have 40%, substantially less than control and other AMF+DSE combinations, and disease index is minimum, preventive effect highest, with compareing or its elsewhere
Manage significant difference, show G.m+S.a+A.l+Ph.l combined treatments compare other AMF+DSE combination to a certain extent can be more
Significantly mitigate Meloidogyne incognita to endanger to caused by cucumber.
Table 3 is inoculated with influences of the AMF and DSE to Cucumber root-knot nematode disease
Note：With after column data, different lowercase letter indication differences are notable (α=0.05) in table
(4) different AMF+DSE combinations and influence of the Meloidogyne incognita (Mi) to cucumber growth
Experimental result is as shown in table 4：Single inoculation AMF or DSE every growth indexes difference are simultaneously little, but substantially lower than
The processing of AMF+DSE combinations is inoculated with, wherein, the plant height of G.m+S.a+A.l+Ph.l combined treatments, stem are thick, ground/bottom dry weight
It is notable in other single inoculation processing differences with single plant yield highest.Meanwhile after inoculation Meloidogyne incognita (Mi), AMF+DSE
Combination is in the indexs such as yield, plant height, underground part dry weight and fresh weight of plant seedlings, and majority processing is significantly higher than CK and is only inoculated with Mi processing, especially
It is that plant height, overground part and the underground part dry weight of G.m+S.a+A.l+Ph.l+Mi combined treatments and single plant yield will be higher than it
The combined treatment of its wiring worm.Experimental result shows, for same combination, is inoculated with the cucumber yield of nematode well below not being inoculated with
Plant.Therefore, experiment shows after Meloidogyne incognita is inoculated with, and AMF+DSE combinations have certain rush to Plant Growth of Cucumber
Enter effect, also there is certain inhibitory action to root-knot nematode.
Table 4 is inoculated with the influence of AMF, DSE and root-knot nematode (Mi) to Plant Growth of Cucumber
Note：With after column data, different lowercase letter indication differences are notable (α=0.05) in table
AMF can suppress the growth, breeding and harm of soil-borne pathogen and nematode to some extent, improve the disease-resistant of plant
Property, DSE is to colonize extensively in the soil fungi in plant root cells or space between cells, it is possible to increase the resistance of host, enhancing
The resistance against diseases of host.Second instar larvae number, the female borer population of nematode and ovum in the cucumber root of different AMF+DSE combined treatments in the present invention
Number is substantially lower than the Meloidogyne incognita brood body number of Mi processing (table 2), wherein G.m+S.a+A.l+Ph.l+Mi combined treatments
Amount is minimum among each processing.
Individually inoculation AMF or DSE processing significantly improves the increment and ground, underground part dry weight of cucumber in the present invention,
Each index of wherein different AMF+DSE combined treatments is significantly higher than single inoculation AMF or DSE processing, and AMF and DSE can promote
Host plant grows.Test result indicates that the yield of G.m+S.a+A.l+Ph.l combined treatments, plant height, dry weight index
It is significantly higher than other combined treatments (table 4).This is due to that AMF and DSE can interact and collectively promote growing for plant.
After different AMF+DSE combined treatments are inoculated with the present invention, S.a+Ph.l+Mi and G.v+Ph.l+Mi combined treatments
The incidence of disease is with compareing no significant difference, and the incidence of disease of G.m+S.a+A.l+Ph.l+Mi combinations is minimum (40%), is substantially less than pair
According to the processing combined with other AMF+DSE, and disease index is minimum, preventive effect highest, reaches the level of signifiance with compareing.Meanwhile
Experimental result shows that the p-type clump branch of G.m+S.a+A.l+Ph.l+Mi combinations raw quantity and is significantly higher than other combinations, shows p-type
Clump branch possesses the effect of stronger induction disease resistance of plant.
The research generally to Plant Growth of Cucumber and anti-root-knot nematode is mainly that a kind of fungi (such as AMF) exists in the prior art
To growth indexes such as plant height, yield, dry weight and fresh weight of plant seedlings, and in the measure of nematode morbidity index, and (such as AMF+ are combined to two kinds of fungies
DSE is combined) influence to root-knot nematode rarely has research.Therefore, the present invention determines different AMF+DSE on the basis of conventional research
Combined treatment is to Plant Growth of Cucumber and the effect of anti-root-knot nematode.By result of the test as can be seen that G.m+S.a+A.l+
Ph.l combinations significantly improve cucumber growth, reduce the nematode incidence of disease and reduce nematode population in root, defend Meloidogyne incognita
Best results.
The above embodiments are merely illustrative of the technical solutions of the present invention, rather than is limited；Although with reference to foregoing reality
Example is applied the present invention is described in detail, for the person of ordinary skill of the art, still can be to foregoing implementation
Technical scheme described in example is modified, or carries out equivalent substitution to which part technical characteristic；And these are changed or replaced
Change, the essence of appropriate technical solution is departed from the spirit and scope of claimed technical solution of the invention.
Claims (6)
1. Phoma sp F2-3-10 bacterial strains, it is characterised in that：Its Classification And Nomenclature isPhoma leveillei, it is preserved in the micro- life of China
Thing culture presevation administration committee common micro-organisms center, its deposit number are：CGMCC No.10482.
2. a kind of AMF+DSE combines microbial inoculum, it is characterised in that the combination microbial inoculum includes AMF microbial inoculums and DSE microbial inoculums, the DSE
Microbial inoculum is microbial inoculum made of Phoma sp F2-3-10 bacterial strains that deposit number is CGMCC No.10482, and the AMF microbial inoculums are：Rub
Western sacculus is mould, the one or more in global Scutellospora or light wall Acaulospora.
3. application of the AMF+DSE combination microbial inoculums in the preparation for promoting plant growth is prepared described in claim 2.
4. application according to claim 3, it is characterised in that：The combination microbial inoculum can improve the plant height of cucumber, overground part
Dry weight, underground part dry weight and single plant yield.
5. application of the AMF+DSE combination microbial inoculums in the preparation for preparing anti-root-knot nematode described in claim 2, its feature exist
In：The root-knot nematode is Meloidogyne incognitaMeloidogyne incognita 。
6. application according to claim 5, it is characterised in that：The combination microbial inoculum can significantly reduce two ages children in cucumber root
The female borer population of borer population, nematode and ovum number.
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| CN109874808B (en) * | 2019-01-25 | 2021-02-02 | 青岛农业大学 | Application of arbuscular mycorrhizal fungi and/or plant symbiotic actinomycetes in preparation of biocontrol preparation for hot pepper and solanaceae vegetables |
| CN110205248B (en) * | 2019-06-12 | 2021-04-27 | 中国矿业大学(北京) | Method for promoting plant growth by jointly inoculating AM and DSE fungi and microbial agent used by method |
| CN110972811B (en) * | 2019-12-23 | 2021-10-12 | 云南农业大学 | Method for establishing DSE (DSE-activated carbon), AMF (Amf) and plant symbiotic system and application of DSE-AMF and plant symbiotic system |
| CN111235037B (en) * | 2020-01-21 | 2021-12-28 | 广西壮族自治区农业科学院微生物研究所 | AMF + DSE combined microbial inoculum and application thereof in promoting ginger growth and resisting bacterial wilt |
| CN115067153A (en) * | 2022-05-17 | 2022-09-20 | 西安科技大学 | Method for enhancing tensile damage resistance of root system and shear resistance of root-soil complex by combining DSE and AMF |
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