CN101555459A - Strain SB177 for controlling strawberry replant diseases and preparation thereof - Google Patents
Strain SB177 for controlling strawberry replant diseases and preparation thereof Download PDFInfo
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Abstract
The invention discloses a strain SB177 for controlling strawberry replant diseases and a preparation thereof, belonging to the technical field of preventing and curing diseases of fruit trees. The strain SB177 and the preparation thereof are specially used for biological control of the fruit trees. The microbial inoculum also can be used for controlling strawberry overground part diseases such as powdery mildew of strawberry, gray mold, etc. The preparation prepared from the strain has the performances of long shelf life, high control effect, wide control spectrum as well as promoting and increasing production. The invention is realized by the following technical proposal: the strain SB177 for controlling strawberry replant disease is bacillus amyloliquefaciens which is the Latin literature name and is preserved in China Center for Type Culture Collection (CCTCC) in March 19th, 2009 with the preservation number of CCTCC NO: M209037.
Description
Technical field
The present invention relates to a kind of bacterial strain and preparation thereof,, belong to the fruit tree technical field of preventing and curing diseases, be exclusively used in the biological control of fruit tree more specifically to a kind of bacterial strain SB177 and preparation thereof of preventing and treating the strawberry replant disease.
Background technology
Replant disease and be meant with a kind of crop, cause that crop growth is bad, disease increases the weight of, and then cause crop yield and the serious phenomenon that descends of quality continuously in the plantation of same plot.Japan claim this class problem for avoid ground phenomenon, continuous cropping is hindered or continuous cropping obstacle, American-European countries is referred to as to replant disease (Replant disease) or replants problem (Replant problem).
For many years, the introduction of strawberry new variety, seed selection and establishment planting technology are the emphasis of my institute's (Jiangsu Hilly Ground Zhenjiang Agriculture Science Research Institute) spread application of agricultural science and technology and science service always.Become the major measure of leading city's peasant programme, obtained huge social and economic benefit.Yet facility cultivation is owing to temperature, the wet environment of continuous cropping and sealing, and pathogen is very easily bred and accumulated, and root knot nematode disease, verticillium, blight etc. are replanted disease, and foliage diseases such as Powdery Mildew and gray mold, becomes the major obstacle that the facility strawberry produces.Replant in the disease especially with verticillium at China's facility strawberry cultivating.Replant year after year breaking out of disease and the example that forces peasant household to scaffold dismantling takes place again and again by strawberry verticillium etc., seriously hinder the development of strawberry cultivating industry and increasing income of peasant.Current, how effective, science control strawberry replant disease? becoming the production practical problems that the most fervent expectation of peasant household solves, also is the problem that the headache that faces of domestic and international expert and attempt solve.
For a long time, control for the strawberry replant disease, mainly utilize halogenated hydrocarbon compound such as monobromethane soil disinfectants such as (methylbromide) in the production, and utilize chemical agent such as F-1991, prevent and treat method at the chemical agent of deciding dipped roots before planting or after field planting, irritating root.Yet the use cost height (1000 yuan/mu times) that utilizes monobromethane etc. to carry out soil disinfection is and if only if year effectively; Moreover monobromethane is hypertoxic gas, to people and animals' severe toxicity, atmospheric ozone layer is had the havoc effect, causes serious environmental to pollute.Country of Europe, the United States has forbidden in 2005 producing and using, forbids in 2015 at the latest according to " Montreal Protocol Copenhagen amendment " developing country in the global range comprehensively, and China's monobromethane is still using to some extent.Current, the domestic substitute products that on producing, still lack the chemical processing soil treatment agent of efficient, low toxicity, wide spectrum, noresidue.Except that chemical fumigant carried out soil treatment, other method of preventing and treating also had methods such as cultural control, physical control, biological control.
Facts have proved in recent years, biological control control strawberry replant disease is considered to the most effective, the most possible method of extensively promoting.It is former that microorganism is more prone to survival and grows surely in soil than on the blade face because fly up and down (1); (2) control of strawberry replant disease still lacks gratifying chemical agent; (3) cultural control (change continuous cropping be rotation system, promote nontoxic seedling, breeding resistant variety) and physical control (daylighting hyperthermia sterilization) method be to the anti-heterodyne of strawberry replant disease, and because its high labour intensity, high control cost and high environmental pollution and do not adopted by the producer.
The non-environmental pollution control method of strawberry replant disease is the focus of studying at present.Domestic research mainly concentrates on following method (1) and utilizes physical method to prevent and treat this disease, adopt the outdoor cell experimental technique as (2000) such as Yuan Huizhu, in the cultivated strawberry process, utilize soil moved in to improve the original method, artificial substratum method, sun power soil disinfection method control strawberry replant disease, prevent and treat the method amount of increase in production respectively up to 100%, 115%, 129% for 3 kinds, little with monobromethane soil disinfection difference, substantially can substitute monobromethane soil disinfection technology, but there is the big and serious environmental pollution problem of labour intensity in these class methods; (2) utilize the active bacteria formulation control, as report such as Wang Zhanwu (2002), with the field planting control strawberry replant disease of bacterium B501 of flying up and down at the strawberry rhizosphere; The red grade of horse bezoar (2004) report utilizes VAM mycorhiza bacterium (Glomus mosseae and Glomus versiforrme) that strawberry is had the short effect of giving birth to, preventing the strawberry verticillium; (3) utilize plant milk extract or microorganism secondary metabolite control strawberry verticillium, the Feng Yulong etc. (1999) that has of research report utilizes the ammonium sulfate precipitation thing of the fermentation of bacillus liquid of flying up and down to prevent and treat; Xu Shuhua etc. (2005) utilize two strains fly up and down bacterium and various plants extract for preventing and treating strawberry verticillium, and obtain first-stage success; Zhen Wenchao (2005) utilizes the medicinal plant source as soil additive control strawberry verticillium, at 3 kinds of The main pathogenic fungi Rhizoctonia of strawberry replant disease solani, Fusarium oxysporum, Verticillium dahliae has screened 125 kinds of medicinal plant decomposition object extracting solution bacteriostatic activities, and the equal amount of mixture of proof Ligusticum wallichii and kuh-seng is expected to be used for the control of strawberry replant disease; Zhao Xiujuan etc. (2007) report has certain preventive effect from the isolating ferment product of flying up and down of strawberry rhizosphere to the strawberry replant disease; Song Zhiwei etc. (2006) report reaches 76.9-84.6% with a kind of complex microorganism preparations to the field efficacy of strawberry verticillium.
Few abroad to the monographic study report of strawberry replant biological control of diseases.Vosatka etc. (1992) inoculate strawberry rhizosphere soil tool synergy do effect mutually simultaneously with AMF (ArbuscularMicorrhizal Fungi) and Pseudomonas putida.Be (2000) reports such as Vosatka equally, utilize Agrobacterium radiobacter can promote AMF at the strawberry rhizosphere colonization; Porras etc. (2007) utilize sun power soil disinfection and Trichoderma agent bonded method, draw the method for adding bacteria agent after sun high-temperature sterilization again, and 2 years preventive effects of strawberry verticillium are reached 84.5%.
At present, the biological control of strawberry replant disease research becomes focus, but can obtain better effects and develop the few of commercial preparation, utilizes the research of bacterium and active bacteria formulation thereof control strawberry replant disease then to rarely have report.
Summary of the invention
The objective of the invention is to solve above-mentioned the deficiencies in the prior art and problem, a kind of bacterial strain SB177 that prevents and treats the strawberry replant disease is provided.Be exclusively used in control strawberry replant disease such as verticillium, blight.
The present invention simultaneously also provides by microbial inoculum of the bacterial strain SB177 of control strawberry replant disease and preparation method thereof, and this microbial inoculum also can be prevented and treated strawberry over-ground part diseases such as powdery mildew of strawberry, gray mold.The preparation of being made by bacterial strain has the performance that shelf-lives is long, preventive effect is high, the control spectrum is wide and urge to give birth to volume increase.This microbial inoculum has the performance that shelf-lives is long, preventive effect is high, the control spectrum is wide and increase production.
The present invention is achieved by the following technical solutions:
The bacterial strain SB177 of control strawberry replant disease of the present invention, this bacterial strain SB177 is a bacillus amyloliquefaciens, the Latin formal name used at school is Bacillus amyloliquefaciens, this bacterial classification on March 19th, 2009 in China's typical culture collection center preservation, preserving number is: CCTCC NO:M209037.Described bacterial classification SB177 has following feature: bacterium colony is white opaque to oyster white, surface irregularity, and colony edge is irregular.Cell is rod-short, and endogenous spore is arranged, and sporangium is expanded, ovalize, G+.Can produce amylase, lipase, proteolytic enzyme, peptase, sucrase.The phytopathogen Verticillium is belonged to (Verticillium), Fusarium (Fusarium), Rhizoctonia (Rhizoctonia), Aspergillus (Aspergillus) and phytophthora (Phytophera) strong antagonistic action is all arranged.
With the biocontrol fungicide of the bacterial strain SB177 of control strawberry replant disease of the present invention preparation, its preparation method may further comprise the steps:
(A) acquisition of bacterium liquid
Sign picking streak culture good SB177 bacterial strain list bacterium colony spot on the R2A solid medium in advance with aseptic bud, being inoculated in the volume that 5ml PDA liquid medium is housed is in the triangular flask of 20ml, shaking culture 16h under 30 ℃, 200rpm condition, spend the night, it is in the triangular flask of 1000ml that gained nutrient solution 5ml of last step all is inoculated in the volume that 400ml PDA nutrient solution is housed, shaking culture 16h under 200rpm, the 30 ℃ of conditions; The 400ml nutrient solution of gained is inoculated in the 20L fermentation culture is housed, wherein solid tolerant content is in the fermentation culture: soyflour 100g, Semen Maydis powder 100g, starch 100g, sucrose 25g, yeast powder 25g, lime carbonate 20g, MnSO41.0g, fermentation condition is set is: dissolved oxygen 100%, stirring velocity is 350rpm, 30 ℃ of leavening temperatures, fermentation time 48h, pH 7.0-7.2;
(B) microbial inoculum complete processing
The fermented liquid that ferments is processed into granular bacteria preparation as follows: earlier that ferment product and lactose is even by 1: 4 (ml/g) mixed, get mixture 1; Mixture 1, poly-ethylbenzene pyrrolidone K30, Tartaric acid and sodium bicarbonate, by 60: 5: 15: 20 weight ratios (W/W) were mixed, and got mixture 2 again; Mixture 2 is crossed 16 mesh sieve granulations on tablets press, through 60 ℃ of steam flows oven dry, the The real time measure water content is controlled between the 3-5% water content again, baking 2h, aseptic aluminium foil bag sealing protect adorn the granule of SB177 viable bacteria.Described viable bacteria total concn is preferably 1 * 10
10～1 * 10
11Cfu/g.
The invention has the beneficial effects as follows:
(1) but the granule thermophilic of gained SB177 bacterial strain of the present invention is preserved 2-3, microbial inoculum all has good preventive effect to strawberry replant disease (verticillium, blight) and strawberry leaves face disease (Powdery Mildew, gray mold).Greenhouse test is the result show, this microbial inoculum all reaches more than 90% the prevention effect of strawberry verticillium and blight, to the preventive effect of powdery mildew of strawberry and gray mold also more than 70%.
(2) but because the SB177 bacterial strain of the present invention bacterium that be safe release, so less than a series of environment negative effects that use brought because of chemical pesticide.The use of said preparation reduces the amount of application of chemical pesticide in the strawberry production greatly, and preventing and treating for the harmless boilogical of strawberry replant disease and leaf diseases provides reliable assurance, benefits modern high-efficiency agriculture.
(3) the microbial inoculum production cost is low, and the profit margin that enterprise obtains is very big.
Embodiment
Embodiment
The bacterial strain SB177 of the control strawberry replant disease of present embodiment can be extracted from March 19th, 2009 and at China's typical culture collection center preservation, preserving number be: CCTCC NO:M209037, Latin formal name used at school are Bacillus amyloliquefaciens bacterial classification, this bacterial classification has following feature: bacterium colony is white opaque to oyster white, surface irregularity, colony edge is irregular.Cell is rod-short, and endogenous spore is arranged, and sporangium is expanded, ovalize, G+.Can produce amylase, lipase, proteolytic enzyme, peptase, sucrase.The phytopathogen Verticillium is belonged to (Verticillium), Fusarium (Fusarium), Rhizoctonia (Rhizoctonia), Aspergillus (Aspergillus) and phytophthora (Phytophera) strong antagonistic action is all arranged.
The biocontrol fungicide of the bacterial strain SB177 preparation of the control strawberry replant disease of present embodiment, its preparation method may further comprise the steps:
(A) acquisition of bacterium liquid
Sign picking streak culture good SB177 bacterial strain list bacterium colony spot on the R2A solid medium in advance with aseptic bud, being inoculated in the volume that 5ml PDA liquid medium is housed is in the triangular flask of 20ml, shaking culture 16h (spending the night) under 30 ℃, 200rpm condition.To go up step gained nutrient solution (5ml), all to be inoculated in the volume that 400ml PDA nutrient solution is housed be in the triangular flask of 1000ml, shaking culture 16h under 200rpm, the 30 ℃ of conditions.The 400ml nutrient solution of gained is inoculated in the 20L fermentation culture is housed (solid tolerant content is in the nutrient solution: soyflour 100g, Semen Maydis powder 100g, starch 100g, sucrose 25g, yeast powder 25g, lime carbonate 20g MnSO41.0g), is provided with fermentation condition and is: dissolved oxygen 100%, stirring velocity is 350rpm, 30 ℃ of leavening temperatures, fermentation time 48h, pH 7.0-7.2 (acid, alkali stream add usefulness 1M HCl, 1M NaOH and regulates pH).
(B) microbial inoculum complete processing
The fermented liquid that ferments is processed into granular bacteria preparation as follows: earlier that ferment product and lactose is even by 1: 4 (V/W) mixed, get mixture 1; Mixture 1, poly-ethylbenzene pyrrolidone K30, Tartaric acid and sodium bicarbonate, by 60: 5: 15: 20 (W/W) mixed, and got mixture 2 again; Mixture 2 is crossed 16 mesh sieve granulations on tablets press, through 60 ℃ of steam flow oven dry, the The real time measure water content is controlled between the 3-5% water content, dries by the fire 2h approximately again.The sealing of aseptic aluminium foil bag protect adorn the granule of SB177 viable bacteria (the viable bacteria total concn is about 1 * 10
10～1 * 10
11Cfu/g).
The land for growing field crops application process of active bacteria formulation of the present invention is as follows:
(1) application method of control strawberry replant disease (verticillium, blight) is, is 1: 500 (W/V) in microbial inoculum and water ratio, earlier microbial inoculum and water is mixed, and irritates root (every strain 100ml) or dip in root (soaking 3min) use when the strawberry transplantation of seedlings.
(2) application method of control powdery mildew of strawberry and gray mold is, is that 1: 500 (W/V) dilution back adds 0.1% (W/V) tween 80 in microbial inoculum and water ratio, mixes back spraying control, and the mu water consumption is 40kg.
Implementation result of the present invention
(1) to the field efficiency test of strawberry verticillium and blight
Condition experimental field: this test arrangement is in fruit tree research chamber, Jiangsu Hilly Ground Zhenjiang Agriculture Science Research Institute strawberry continuous cropping (20 years) booth field, and verticillium and blight are for retransmitting.Strawberry is bright treasured in transfer on September 18 in 2007 for the examination strawberry cultivars.
Test treatment process: when the strawberry transplantation of seedlings, dip in root and handle.That is, (viable bacteria content is about 5.16 * 10 with finished product SBl77 microbial inoculum
10Cfu/g) after the dilution 500 times (W/V), when the strawberry transfer, soak root and handle 3min, blank replaces with clear water, contrast biological and ecological methods to prevent plant disease, pests, and erosion medicament is handled 3min for " EM mycorhiza " 500 times of liquid soak root, 300 seedlings of every processing divide 3 to repeat the sub-district random alignment, 50d investigation incidence calculates diseased plant rate and preventive effect after the transfer.
The result: greenhouse test result (table 1) shows that the SBl77 granule is respectively 91.3% and 91.9% to the prevention effect of strawberry verticillium and blight, is significantly higher than " EM mycorhiza " prevention effect to strawberry verticillium (46.4%) and blight (23.0%).
Table 1SBl77 microbial inoculum is to the field controling test result of strawberry verticillium and blight
Annotate: different capitalizations are represented significance level of difference P=0.01.
(2) to the field efficiency test of powdery mildew of strawberry and gray mold
Condition experimental field: this test arrangement is in knob, Jiangsu Zhenjiang institute of agricultural sciences fruit tree research chamber strawberry continuous cropping (20 years) booth field.Strawberry is Feng Xiang in transfer on September 30 in 2007 for the examination strawberry cultivars.
Test treatment process: behind strawberry transplantation of seedlings 45d,, prevent and treat altogether 4 times every spraying control in 15 days 1 time.Institute's with medicament and concentration are: (viable bacteria content is about 5.16 * 10 to the SBl77 microbial inoculum
L0Cfu/g) 500 times of liquid, 12.5% flutriafol 90ml/ mu (contrast chemical agent), blank is not is not prevented and treated.Sub-district area 20m
2, 3 repeat every processing.Last 1 control finishes back 15d investigation incidence at interval, calculates disease index and preventive effect.
The result: greenhouse test result (table 2) shows that the SBl77 particle is respectively 77.62% and 77.89% to the prevention effect of powdery mildew of strawberry and gray mold, and is suitable to the prevention effect of Powdery Mildew (81.05%) and gray mold (82.92%) with the chemical agent flutriafol.
Table 2SB177 microbial inoculum is to the field controling test result of powdery mildew of strawberry and gray mold
Annotate: different capitalizations are represented significance level of difference P=0.01.
Claims (5)
1, a kind of bacterial strain SB177 that prevents and treats the strawberry replant disease, this bacterial strain SB177 is a bacillus amyloliquefaciens, the Latin formal name used at school is Bacillus amyloliquefaciens, this bacterial classification on March 19th, 2009 in China's typical culture collection center preservation, preserving number is: CCTCC NO:M209037.
2, the application of the bacterial strain SB177 of the described control strawberry replant of claim 1 disease aspect control strawberry replant disease.
3, a kind of biocontrol fungicide for preparing with the bacterial strain SB177 of the described control strawberry replant of claim 1 disease.
4, the preparation method of biocontrol fungicide according to claim 3 is characterized in that may further comprise the steps:
(A) acquisition of bacterium liquid
Sign picking streak culture good SB177 bacterial strain list bacterium colony spot on the R2A solid medium in advance with aseptic bud, being inoculated in the volume that 5ml PDA liquid medium is housed is in the triangular flask of 20ml, shaking culture 16h under 30 ℃, 200rpm condition, spend the night, it is in the triangular flask of 1000ml that gained nutrient solution 5ml of last step all is inoculated in the volume that 400ml PDA nutrient solution is housed, shaking culture 16h under 200rpm, the 30 ℃ of conditions; The 400ml nutrient solution of gained is inoculated in the 20L fermentation culture is housed, wherein solid tolerant content is in the fermentation culture: soyflour 100g, Semen Maydis powder 100g, starch 100g, sucrose 25g, yeast powder 25g, lime carbonate 20g, MnSO41.0g, fermentation condition is set is: dissolved oxygen 100%, stirring velocity is 350rpm, 30 ℃ of leavening temperatures, fermentation time 48h, pH 7.0-7.2;
(B) microbial inoculum complete processing
The fermented liquid that ferments is processed into granular bacteria preparation as follows: earlier that ferment product and lactose is even by 1: 4 mixed, get mixture 1; Mixture 1, poly-ethylbenzene pyrrolidone K30, Tartaric acid and sodium bicarbonate, by 60: 5: 15: 20 weight ratios were mixed, and got mixture 2 again; Mixture 2 is crossed 16 mesh sieve granulations on tablets press, through 60 ℃ of steam flows oven dry, the The real time measure water content is controlled between the 3-5% water content again, baking 2h, aseptic aluminium foil bag sealing protect adorn the granule of SB177 viable bacteria.
5, the preparation method of biocontrol fungicide according to claim 4 is characterized in that described viable bacteria total concn is 1 * 10
10～1 * 10
11Cfu/g.
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| CN102604864A (en) * | 2012-03-09 | 2012-07-25 | 陕西加伦多作物科学有限公司 | Microbial bactericide and application thereof |
| CN102876603A (en) * | 2012-09-23 | 2013-01-16 | 江苏丘陵地区镇江农业科学研究所 | Bacillus amyloliquefaciens Mg116 and application thereof |
| CN103004886A (en) * | 2012-12-11 | 2013-04-03 | 广东海洋大学 | Marine bacterium preparation for preventing banana vascular wilt |
| CN103253991A (en) * | 2013-05-23 | 2013-08-21 | 江苏丘陵地区镇江农业科学研究所 | Microbial organic fertilizer specially used for strawberries, as well as preparation method and application thereof |
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| CN104012649A (en) * | 2014-06-02 | 2014-09-03 | 镇江绿健农业科技开发有限公司 | Strawberry preservative containing strains SB177 and preparing method of strawberry preservative |
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| CN105439657A (en) * | 2015-12-17 | 2016-03-30 | 山西省农业科学院生物技术研究中心 | Preparation method for strawberry dedicated anti-continuous cropping biological organic fertilizer |
| CN105532735A (en) * | 2015-11-21 | 2016-05-04 | 安徽省农业科学院植物保护与农产品质量安全研究所 | Bactericidal and insecticidal composition containing Bacillus amyloliquefaciens and matrine |
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| CN103004886A (en) * | 2012-12-11 | 2013-04-03 | 广东海洋大学 | Marine bacterium preparation for preventing banana vascular wilt |
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| CN103651582A (en) * | 2013-09-17 | 2014-03-26 | 周剑平 | Broad-spectrum composite microbial fungicide and preparation method thereof |
| CN103651582B (en) * | 2013-09-17 | 2016-03-02 | 周剑平 | Broad-spectrum composite microbial fungicide and preparation method thereof |
| CN104012649A (en) * | 2014-06-02 | 2014-09-03 | 镇江绿健农业科技开发有限公司 | Strawberry preservative containing strains SB177 and preparing method of strawberry preservative |
| CN104012649B (en) * | 2014-06-02 | 2016-04-13 | 镇江绿健农业科技开发有限公司 | A kind of strawberry preservative containing bacterial strain SB177 and preparation method thereof |
| CN104388335B (en) * | 2014-10-24 | 2017-04-12 | 吉林农业大学 | Bacillus amyloliquefacien FS6 and application thereof |
| CN104388335A (en) * | 2014-10-24 | 2015-03-04 | 吉林农业大学 | Bacillus amyloliquefacien FS6 and application thereof |
| CN104560837B (en) * | 2015-02-02 | 2017-10-03 | 青岛蔚蓝生物集团有限公司 | A kind of bacillus amyloliquefaciens and its application |
| CN104988100A (en) * | 2015-07-29 | 2015-10-21 | 江苏丘陵地区镇江农业科学研究所 | Bacillus amyloliquefaciens CD-17 strain and application thereof |
| CN105532735A (en) * | 2015-11-21 | 2016-05-04 | 安徽省农业科学院植物保护与农产品质量安全研究所 | Bactericidal and insecticidal composition containing Bacillus amyloliquefaciens and matrine |
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| CN107858294A (en) * | 2017-11-16 | 2018-03-30 | 陕西师范大学 | A kind of multi-functional bacterial strain for preventing and treating disease in replanting |
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