CN101402930A - A plant growth-promotiong rhizosphere - Google Patents
A plant growth-promotiong rhizosphere Download PDFInfo
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- CN101402930A CN101402930A CNA2008101812966A CN200810181296A CN101402930A CN 101402930 A CN101402930 A CN 101402930A CN A2008101812966 A CNA2008101812966 A CN A2008101812966A CN 200810181296 A CN200810181296 A CN 200810181296A CN 101402930 A CN101402930 A CN 101402930A
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Abstract
The invention discloses a plant growth-promoting rhizobacteria strain which is separated from dung of bos grunniens in Qinghai Province and preserved in the General Microorganisms Collection Center of China Committee for Culture Collection of Microorganisms with a preservation name of Bacillus subtilis QM34 and a preservation number of CGMCC No. 2669 on September 22, 2008; the strain QM34 has antagonistic action on 12 pathogenic bacteria of soil-born plant fungus; and the fermentation solution of the strain QM34 and the cell-free nutrient solution of the fermentation solution of the strain QM34 has obvious prevention and treatment effects on alternaria solani, significantly promotes the growth of tomato plants and can be colonized in a large quantity according to the tomato plant rhizosphere. Therefore, the rhizobacteria strain is a broad-spectrum growth-promoting rhizobacteria strain used or preventing and treating alternaria solani and most popular among people.
Description
Technical field
The present invention relates to a plant growth-promotiong rhizosphere, but wide spectrum is prevented and treated the various plants silborne fungal diseases, can grow surely in a large number at plant rhizosphere, and growth and development of plant is had obvious facilitation, belong to the agricultural microorganism technical field, also belong to plant biological Prevention Technique field.
Background technology
Since phase late 1960s, along with the secular unreasonable use of a large amount of chemical pesticides, the development and the human beings'health of rural economy in the chemical prevention serious threat of plant pest, have become the serious challenge that human society enters 21 century.
Along with science and technology development, in the face of the deterioration day by day of environment, transformation has taken place to understanding and the crop protection strategy of harmful organism in people, changes " appropriateness tolerance " into from " killing all " to harmful organism.The Ministry of Agriculture has implemented " pollution-free food action plan " in startup in 2002, and this indicates that China Agricultural Structure Adjustment has entered new developmental stage.Look forward to the development need of food safety from now on, biological control will become the important measures of the prevention and control of plant diseases, pest control, and vast potential for future development is arranged.
But biological control is seen still on the whole and is come with some shortcomings: the one, and the specimen material that is used to screen mainly concentrates on endophyte of rhizosphere soil, infected sick body tissue, susceptible plant etc. and locates, and has limited the diversity of Microbial resources; The 2nd, the biocontrol microorganisms of screening usually is at a certain or a certain class pathogenic bacteria fungistatic effect preferably to be arranged, and lacks the bacteriostatic broad spectrum; The 3rd, the variation meeting of ambient conditions directly influences the Biocontrol Mechanism and the biocontrol effect of biocontrol fungicide, i.e. preventive effect instability; The 4th, the production of biocontrol microorganisms and use technology are perfect inadequately or the like.For carry out the biological control policy of " putting prevention first; science prevention and control; improve according to law; promote healthy " comprehensively; instruct various places safety, scientific and rational use agricultural chemicals and integrated control, further promote the non-environmental pollution control of agroforestry harmful organism, preserve the ecological environment and species diversity; the structure of preventing and treating in conjunction with present China farm crop has proposed a series of new Prevention Technique.
One of new technology of biological control is exactly to separate and short living bacterium (the plant growth-promotingrhizobacteria of screening plant rhizosphere, PGPR), it can be grown at plant rhizosphere to high-density surely, have concurrently and suppress phytopathogen, rhizosphere harmful microorganism, and promote plant-growth and increase the effect of crop yield, the more important thing is and can also inducing plant produce system's resistance by some PGPR (Induced systemic resistance ISR), thereby improves the resistance against diseases of plant integral body.So further investigation and the development of PGPR will prevent and treat this difficult problem for the solution plant soil-borne diseases and represent tempting prospect.
Early blight of tomato (Tomato Early Blight) is called " ring spot " or " summer eqpidemic disease " again, throughout the growing season can generally taking place whenever and wherever possible of tomato, is to endanger one of more serious fungal disease.The host range of this cause of disease is extensive, except that the harm tomato, also can endanger solanaceous vegetable crops such as eggplant, capsicum and potato.The control of early blight of tomato evil at present mainly depends on chemical agent.Though abroad having obtained some has the antagonism bacterium of better prevention effect, but not carrying out specially with the tomato early blight bacterium is the screening operation of the antagonistic microbe of controlling object, domesticly also only be confined to simple laboratory screening work at present, antagonistic microbe is based on yeast.
Summary of the invention
Technical problem purpose of the present invention provides a strain can effectively prevent and treat the early blight of tomato evil, can effectively promote plant-growth, can grow in plant rhizosphere effectively surely, has the short endophytic bacteria bacterial strain of the plant rhizosphere antagonism broad spectrum, useful just in order to overcome above-mentioned the deficiencies in the prior art.
Technical scheme the objective of the invention is to be achieved through the following technical solutions.
The short endophytic bacteria bacterial strain of novel plant rhizosphere of the present invention is: subtilis (Bacillus subtilis) QM34, depositary institution: China Committee for Culture Collection of Microorganisms common micro-organisms center, its unit abbreviates CGMCC as, preservation day: on 09 22nd, 2008, deposit number: CGMCC No.2669.(following abbreviation B.subtilisQM34).
B.subtilis QM34 bacterial strain of the present invention is to separate first to have obtained the wide spectrum antagonistic microbe from the yak excrement of Qinghai.Seek the microflora of different geographical environments, development and utilization beneficial microorganism resource from the yak excrement of Qinghai will be a great breakthrough in the biological and ecological methods to prevent plant disease, pests, and erosion field.
The morphological specificity of B.subtilis QM34 bacterial strain of the present invention is: the QM34 bacterium colony is opaque, and there is gauffer on the surface, and shape is incised at the edge; Shaft-like, the Gram-positive of thalli morphology produces central spore, the gemma ellipse.Compare with existing withered grass gemma biocontrol strain, sporangiocyst has slightly and expands.
B.subtilis QM34 bacterial strain of the present invention all has antagonistic action to 12 kinds of soil biography property plant epiphyte cause of disease bacterium, particularly tomato early blight bacterium is had the obvious suppression effect.Therefore, be the biocontrol strain that a strain has the wide spectrum antagonism.
B.subtilis QM34 bacterial strain of the present invention has tangible prevention effect to the early blight of tomato evil; Growth to tomato plant has obvious facilitation; Can grow surely in a large number at the tomato plant rhizosphere.Therefore be the short endophytic bacteria of plant rhizosphere that a strain has potential using value.
B.subtilis QM34 bacterial strain of the present invention, it is the wide spectrum antagonism biocontrol strain of the early blight of tomato evil of seed selection first, be seed selection first be main controlling object with the early blight of tomato evil, and have multi-functional plant growth-promotiong rhizosphere bacterial strains such as short life, diseases prevention concurrently.
Description of drawings
Fig. 1 is after the dilution QM34 bacterial strain fermentation liquor of difference is handled, the variation tendency of the total genus bacillus quantity of tomato plant rhizosphere soil of 3 kinds.
Embodiment
The separation screening test of embodiment 1QM34 bacterial strain
Separate from the yak excrement of Qinghai through the gradient dilution method, obtain a plant growth-promotiong rhizosphere, B.subtilis QM34 bacterial strain through the screening of flat board face-off growth method.Specific implementation process comprises: claim Qinghai yak excrement sample 10.0g, grind, join under aseptic condition and shake in the 90ml sterilized water evenly, get its supernatant liquor and be diluted to suitable (10
-5, 10
-6, 10
-7) concentration gradient, gradient liquid 100 ℃ of heating in water bath 10 minutes, is inoculated in beef-protein medium with conventional dilution method coating, cultivated 1 day for 37 ℃, select the apparent in view bacterium colony of bacterial colony morphological differences, repeat streak inoculation on corresponding agar plate, purifying obtains single bacterium colony; Adopt the dull and stereotyped face-off of PDA growth method primary dcreening operation: with 12 kinds of pathogenic fungies is indicator, connect pathogenic bacteria in dull and stereotyped central point, when treating that pathogenic bacteria grows into the about 1-1.5cm of colony diameter, the bacterial strain of primary dcreening operation will be inserted, put and connect on 6 angle points at an inoculation point anomaly plate central authorities 2.5cm place, and to connect the primary dcreening operation bacterial strain be contrast not put, place 28 ℃ of incubators to cultivate the growing state of observed and recorded pathogenic fungi 3-5 days.Selecting has stronger relatively inhibiting bacterial strain to enter multiple sieve to the pathogenic bacteria growth.The same primary dcreening operation of concrete operations of multiple sieve, be respectively indicator with 12 kinds of pathogenic bacterias, antagonism is dull and stereotyped to be cultivated after 5 days or 10 days, measure related data respectively, and record R value (R=pathogenic bacteria and distance/antagonism bacterium center, antimicrobial mycelia short of money edge are to the distance at pathogenic bacteria mycelia edge), according to the size of R value, screening obtains having the wide spectrum antagonistic action, good antimicrobial effect, the persistent B.subtilis QM34 of antagonistic action bacterial strain (seeing Table 1), 3 observed values of every processing, test repeats 2 times.The QM34 bacterium colony is opaque, and there is gauffer on the surface, and shape is incised at the edge; Shaft-like, the Gram-positive of thalli morphology, generation central spore, sporangiocyst have expands the gemma ellipse.
Table 1B.subtilis QM34 bacterial strain passes the antagonistic action of plant epiphyte germ to 12 kinds of soil
R
*: pathogenic bacteria and distance/antagonism bacterium center, antimicrobial mycelia short of money edge are to the distance at pathogenic bacteria mycelia edge
Embodiment 2QM34 bacterial strain is to the potted plant efficiency test of early blight of tomato
Test prevents and treats early blight of tomato with the fermented liquid and the sterile liquid of QM34 bacterial strain respectively, and each is prevented and treated liquid and respectively establishes 2 processing, promptly in the day before yesterday of inoculating tomato early blight pathogenic bacteria with prevent and treat the inoculation (being designated as forward and backward respectively) of liquid one day after respectively.With the inoculation sterilized water is contrast.All are handled and implement in three tomato varieties simultaneously.Whole test is established 2 repetitions, random alignment.
B.subtilis QM34 bacterial strain is inoculated in the YPF-Ca liquid nutrient medium, and 35 ℃, shake-flask culture 4d, concentration is about 10
9Cfu/ml dilutes 10 times of fermented liquids that are the QM34 bacterial strain; With centrifugal 20 minutes of the fermented liquid 10000rpm of QM34 bacterial strain, remove precipitation, supernatant liquor with millipore filtration (0.22um) filtration sterilization, is QM34 bacterial strain sterile liquid after diluting 3 times.Tomato variety is respectively: No. 1, and bright red cooperation 903, Shanghai tomato institute; No. 2, the big people 601 of haw king, big people's research of agricultural science institute; No. 3, the bright red guarantor's hat of Qin Feng (disease-resistant), Lintong District, Xi'an Qin Feng Vegetable Research Institute.Three kinds of tomato seeds percentage of germination are all greater than 85%.
Tomato early blight bacterium (Alternaria solani) is after cultivating a week on the PDA substratum, inject 4 ℃ of cold water in the ware, strike off aerial hyphae, uviolizing 2min, 25 ℃ of following constant temperature culture 2d, mix with the Cha Shi liquid medium of early blight bacterium then, be pathogenic bacteria liquid, pathogenic bacteria liquid spore count is 10
6Cfu/ml.
When growing to 5-6 sheet leaf for the examination tomato seedling, the beginning inoculation test.Smear and root-pouring method inoculating tomato early blight bacterium with acupuncture simultaneously, inoculate 2 times; After smearing inoculation QM34 bacterial strain fermentation liquor and QM34 bacterial strain sterile liquid, irritate the every strain of root inoculation 10ml/ again.
Potted plant control test draws (seeing Table 2), and each is handled all certain prevention effect, and exists significant difference.Three kinds all are obviously to be better than inoculation afterwards prior to the prevention effect of inoculating the QM34 bacterial strain fermentation liquor before the pathogenic bacteria; The test-results of No. 3 kinds is shown, also is prior to better before the inoculation pathogenic bacteria with the prevention effect of QM34 bacterial strain sterile liquid; For No. 3 tomatoes, the prevention effect of direct inoculation QM34 fermented liquid is better than the prevention effect of inoculation sterile liquid.
Embodiment 3 bacterial strain QM34 are to the promotion test of tomato plant strain growth
With extension rate is that 1,10,20 times QM34 fermented liquid is inoculated three kinds of tomato plants respectively, is contrast to connect sterilized water, begins to measure the number of blade and plant height the same day from inoculation, every 7 days record one secondary data.Each 2 strain of the plant of the average plant height of the every treatment group of picked at random after 28 days, investigate its blade size, root development, spacing of fins from, physical signs indexs such as diameter stem (seeing Table 3).
Get the method for root: plant and seedling alms bowl body are soaked in the water together, changed water then and rinse well, obtain the sample and the photograph of root; Root is put into a transparent pallet, and inject the dark water of 3～5mm, directly measure and to write down total root long; Root is placed on the 1h that completes in 105 ℃ the baking oven and is placed on baking 48h under 55 ℃ of constant temperature, the cooling back is taken out and is calculated weight of root system.
After the QM34 of different extension rates fermentation liquor treatment, the average number of blade of finding three kinds of tomato varieties is different with the changing conditions of average plant height, the average number of blade and plant height all increase in time prolongation, the average number of blade and average plant height all increase (removing No. 2 tomatoes), plant are not produced restraining effect (seeing Table 3).To No. 1, No. 2 and No. 3 optimum QM34 fermented liquid of tomato variety extension rates is respectively 20,10 and 1 times.In entire test, No. 2 tomato varieties all are better than No. 1 and No. 3 kinds on the speed of the size of percentage of germination, germination and growing way, so adding QM34 fermented liquid rear blade number still in increase, and plant height increases slower with respect to contrast, this is just illustrating that the QM34 fermented liquid can promote that plant effectively grows, and avoids plant the excessive growth phenomenon to occur.After the concentration multiple of finding three kinds of QM34 fermented liquids is in addition handled plant, blade size, blade gap distance, characteristic index such as diameter stem with contrast between in various degree increase (seeing Table 3) is all arranged.Greater than other treatment group, and diameter stem is also thin than other treatment group from obviously for the 10 times of diluents in No. 1 tomato variety and the spacing of fins of No. 2 tomato control groups, and the basic stem of other treatment group is lignifying all, and the stem color has also become sorrel; The blade area of 20 times of diluent treatment group in No. 1 tomato variety and root length overall are obviously greater than other treatment group, though as seen the variation of average plant height is little, but other characteristic indexs such as its lobe numbers increase, and this state is unusual ideal on seedling management, is real strong sprout.This explanation QM34 bacterial strain fermentation liquor all has certain promoter action to the growth of three kinds of tomato varieties.
Table 2QM34 bacterial strain is to the protection effect of early blight of tomato
*Identical letter representation does not have significant difference (p＞0.05), and different alphabets is shown with significant difference (p＜0.05).
The physical signs of table 3 each tomato plant after the QM34 bacterial strain fermentation liquor is handled
Embodiment 4 bacterial strain QM34 decide to grow test the tomato plant rhizosphere
With the inoculation sterilized water is contrast, to the rhizosphere soil of 3 tomato varieties through the QM34 fermented liquid of inoculation different concns, has carried out the detection of dynamic (see figure 1) of total genus bacillus quantity with plate dilution method.Sampling in per 7 days is once tested and is repeated 3 times.The number change that test finds respectively to handle the total genus bacillus of tomato plant rhizosphere soil obviously increases (2.8 * 10 along with the prolongation of time is all increasing than the quantity of not irritating the root inoculation
6Cfu/g) (p＜0.01) illustrates that the QM34 bacterial strain has very strong competitive power, can grow surely in the rhizosphere soil of plant and breeding in a large number.
Claims (6)
1, a plant growth-promotiong rhizosphere, depositary institution: China Committee for Culture Collection of Microorganisms common micro-organisms center, its unit abbreviates CGMCC as, preservation name: subtilis (Bacillus subtilis) QM34, preservation day: on 09 22nd, 2008, deposit number: CGMCC No.2669.
2, a plant growth-promotiong rhizosphere bacterial strain according to claim 1 is characterized in that, separates obtaining from the yak excrement of Qinghai.The QM34 bacterium colony is opaque, and there is gauffer on the surface, and shape is incised at the edge; Shaft-like, the Gram-positive of thalli morphology, generation central spore, sporangiocyst have slightly expands the gemma ellipse.
According to claim 1 and 2 described plant growth-promotiong rhizosphere bacterial strains, it is characterized in that 3,12 kinds of soil biography property plant epiphyte cause of disease bacterium are all had antagonistic action, particularly tomato early blight bacterium is had the obvious suppression effect.
4, a plant growth-promotiong rhizosphere bacterial strain according to claim 3 is characterized in that the early blight of tomato evil is had tangible prevention effect.
5, a plant growth-promotiong rhizosphere bacterial strain according to claim 3 is characterized in that, the growth of tomato plant is had obvious facilitation.
6, a plant growth-promotiong rhizosphere bacterial strain according to claim 3 is characterized in that, can grow surely in a large number at the tomato plant rhizosphere.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102409014A (en) * | 2011-11-23 | 2012-04-11 | 山东省林业科学研究院 | Winter jujube rhizosphere growth promoting bacillus subtilis and application thereof |
| CN112195108A (en) * | 2020-11-09 | 2021-01-08 | 广西壮族自治区农业科学院微生物研究所 | Cladosporium cladosporioides strain MS2 and application thereof |
| CN116396889A (en) * | 2023-01-13 | 2023-07-07 | 南京农业大学 | Iron-producing carrier golden fungus for preventing and controlling soil-borne bacterial wilt and promoting tomato growth and application thereof |
-
2008
- 2008-11-15 CN CN2008101812966A patent/CN101402930B/en not_active Expired - Fee Related
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102409014A (en) * | 2011-11-23 | 2012-04-11 | 山东省林业科学研究院 | Winter jujube rhizosphere growth promoting bacillus subtilis and application thereof |
| CN112195108A (en) * | 2020-11-09 | 2021-01-08 | 广西壮族自治区农业科学院微生物研究所 | Cladosporium cladosporioides strain MS2 and application thereof |
| CN112195108B (en) * | 2020-11-09 | 2023-03-14 | 广西壮族自治区农业科学院 | Cladosporium cladosporioides strain MS2 and application thereof |
| CN116396889A (en) * | 2023-01-13 | 2023-07-07 | 南京农业大学 | Iron-producing carrier golden fungus for preventing and controlling soil-borne bacterial wilt and promoting tomato growth and application thereof |
| CN116396889B (en) * | 2023-01-13 | 2024-01-09 | 南京农业大学 | Iron-producing carrier golden fungus for preventing and controlling soil-borne bacterial wilt and promoting tomato growth and application thereof |
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