CN103396954A - Biological prevention and control bacterial strain for preventing and controlling rice sheath blight, biological organic fertilizer, and preparation method of biological organic fertilizer - Google Patents
Biological prevention and control bacterial strain for preventing and controlling rice sheath blight, biological organic fertilizer, and preparation method of biological organic fertilizer Download PDFInfo
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Abstract
The invention discloses a biological prevention and control bacterial strain for preventing and controlling rice sheath blight, a biological organic fertilizer, and a preparation method of the biological organic fertilizer, which relate to biological prevention and control bacterial strains of rice and biological organic fertilizers thereof, and belong to the technology of intensive agricultural production. According to the invention, Trichodermaasperellum12, which plays an obvious role in biological prevention and control on the rice sheath blight, is obtained by separation, the biological organic fertilizer is prepared by composting the biological prevention and control bacterial strain, rice hulls, straws and poultry excrement and contains bacterial strain hypnospore of more than 0.5*10<8>cfug<-1>, the water content of the fertilizer is smaller than 15% and the organic content of the fertilizer is more than 35%. Experiments show that after the biological organic fertilizer is applied into rice fields, most of matter in the organic fertilizer can keep afloat, the hypnospore of the biological prevention and control bacterial strain, which is adsorbed on the organic fertilizer, can rapidly breed on water, aerial mycelium is formed at a water-gas junction zone, and a hypha belt is formed at the contact part of rice stalks and the water surface, so that rhizoctonia solani of the rice sheath blight on the water surface is effectively prevented from contacting the rice stalks, and the prevention and control rate of the rice sheath blight is more than 70%.
Description
Technical field
The invention belongs to microbial technology field, relate to a strain rice biological control strains separation and identify, the biological organic fertilizer that this biocontrol strains is made and the application on rice sheath blight disease controlling thereof.
Two, background technology
Paddy rice is the crop of cultivated area maximum in the world, its cultivated area reaches 1.5 hundred million hectares, account for 11% of world's cultivated area, China's rice growing area accounts for more than 20% of whole Model on Sown Areas of Farms, the main food crop of China, be also first farm crop, the population staple food more than 40% is rice.
Rice sheath blight disease is one of Main Rice Diseases, and onset area and harm loss at present occupies the rice disease first place, generally can make the paddy rice underproduction 10%-30%, in the time of serious, can reduce 50%.Except common, cause rice sheath blight disease morbidity all the more seriously, as Rice Cropping density, increase, growth cycle shortens, the popularization of the high kind of tillering of the short bar of paddy rice, the nitrogenous fertilizer usage quantity increases, in recent years because a large amount of labor forces in rural area go out, fertilizer was made in farmland to rice straw or burning becomes direct returning to farmland as the energy by former transporting, a large amount of sclerotium prolonged stay farmlands, sprout in case external environment is suitable for sclerotium, is very easy to cause the rice sheath blight disease outburst.The rice sheath blight disease pathogenic bacteria imperfect stage be dry thread Pyrenomycetes (
Rhizoctonia solani), in classification, belong to Deuteromycotina, hyphomycetes, agonomycetales, Rhizoctonia.Perfect stage be thanatephorus cucumeris(frank) donk (
Thanatephorus cucumris), belong to Basidiomycotina, Basidiomycetes, angle load Zoopagales, angle load Cordycepps, the lead fungi of dying belongs to.According to the anastomosis dry thread Pyrenomycetes, be divided into 14 and merge the group, such bacterium host range is very extensive, cause harm basal part of stem and the root of a variety of plants such as paddy and wheat, corn, cotton, fiber crops, soybean, peanut, beet, tobacco and vegetables, cause that seedling is withered, stem is withered and the damping-off such as root-rot, perhaps on basal part of stem, leaf sheath, produce the brown moire spot that is of different shades, cause the cereal plants banded sclerotial blight.This disease causes paddy rice not ear, or the not plump paddy of heading is more, and grain heavily descends.
Germ mainly survives the winter in soil with sclerotium, also can on invalid body or on other hosts such as weeds in field, survive the winter with mycelium, and the Intolerance environment capacity is very strong.During the Second Year rice cultivation, because irrigation of paddy fields makes sclerotium separately or be mixed on the foreign material such as stalk remains to fly at the water surface, after the rice shoot plantation, sclerotium adheres on the leaf sheath of the nearly water surface of rice shoot, condition is suitable to be sprouted mycelia and invades the leaf sheath tissue, after the invasion paddy rice, germ forms aerial hyphae, can directly infect contiguous plant.The sclerotium that falls into the water in early days also can cause the rice strain and infect, the early rice sclerotium is the main source of infection water of late rice banded sclerotial blight, the sclerotium of different developmental phases all can be sprouted and causes a disease and form new sclerotium, sclerotium has repeatedly to be sprouted and the underground ability that infects, and these characteristics of sclerotium are brought very complicated problem for control.
In current agriculture production, mainly with jingganmycin, prevent and treat this disease, jingganmycin is less to the harm of environment, but because the jingganmycin life-time service has produced the jingganmycin resistant strain.Therefore, find environmental friendliness, person poultry safety, high-efficiency prevention and control method very necessary.
Utilizing the biological control microorganism to control disease in conjunction with the agriculture production measure is the effective way of instead of chemical agricultural chemicals prevention and control rice sheath blight disease, along with social development, agricultural development is nuisanceless, efficient, the low labour demand of demand all the more, therefore, in conjunction with application mode efficiently, to prevent and treat rice sheath blight disease very necessary for application high-performance bio control bacterial strain.
Up to now, there is no to find to relate to the bibliographical information of theme of the present invention.
Three, summary of the invention
Technical problem
The object of the invention is to screen the microorganism strains that a plant height effect is prevented and treated rice sheath blight disease pathogenic bacteria dry thread Pyrenomycetes, this bacterial strain suppresses the pathogenic bacterial infection paddy rice by competition for space, generation antibacterial substance and parasitization, by the specific production technique of this bacterial strain and the zymotechnique in later stage, produce bio-feritlizer, this fertilizer be conducive to biocontrol strains in the paddy growth environment survival and be conducive to remain biocontrol strain and suppress the pathogenic bacteria ability.
Technical scheme
A kind of biocontrol strains of prevention and control rice sheath blight disease, it is characterized by this bacterial strain is trichoderma asperellum 12(
Trichoderma asperellum12), on May 16th, 2013 was preserved in Chinese Typical Representative culture collection center, and the preservation address is wuchang, wuhan Luo Jiashan, and the bacterial strain preserving number is CCTCC NO:M 2013213, referred to as trichoderma asperellum CCTCC NO:M 2013213.
The Main Biological of described trichoderma asperellum CCTCC NO:M 2013213 is: on the PDA substratum, cultivated 3 days for 28 ℃, mycelia can be covered with whole culture dish, mycelial growth initial stage white, felted, by center to edge, form gradually ring-type, the ring-type that keeps to the side mycelia color is more shallow, near the centres circle, produces dense sap green conidium, the culture dish of growth mycelia gives out very heavy Coconut Juice aromatic odour, back side deep yellow; Conidiophore, to growth, is arborization, sap green, and the bottle stalk is short, and base portion attenuates, and expand centre; Conidium is spherical in shape, surface irregularity, and size is 3.5～5.0 μ m * 2.0～5.0 μ m; The ITS sequential evolution analysis shows that this bacterial strain is trichoderma asperellum
T. asperellum.
A kind of bio-organic fertilizer, is characterized in that containing 0.5 * 10
8Cfu g
-1Above-described trichoderma asperellum CCTCC NO:M 2013213 bacterial strain statospores, water content lower than 15%, organic content is greater than 35%.
The preparation method of described bio-organic fertilizer is characterized in that realizing as follows:
1) described trichoderma asperellum CCTCC NO:M 2013213 is inoculated into to the PDA nutrient solution, carry out liquid fermenting production, the condition of its fermentative production is: initial pH nature, 25 ~ 30 ℃ of culture temperature, dissolved oxygen air flow scope is 30～100%, 170rpm, fermentation later stage mycelium pellet all is broken into the mycelia fragment, the colony-forming unit of fermented liquid bacterial strain>=1 * 10
8Cfu ml
-1
2) rice husk, stalk and chicken manure be according to 50:30:20 ratio compost, and after 15 ~ 30 days, rice husk is softening, stalk rots, sterilizing 36 hours of 100 ℃ of matrix, high temperature drying, dry after water content of substrate lower than 5%, form solid fermentation matrix;
3) fermented liquid described in step (1) is inoculated into to the described solid fermentation matrix of step (2) by the inoculum size of 200 L/ tons and carries out solid fermentation, leavening temperature is 20 ~ 35 ℃, in fermenting process, turning is 2 ~ 3 times, ferments and finishes afterwards in 10 ~ 15 days, forms the solid fermentation thing.
4) the described solid fermentation thing of step (3) is inoculated into to pig manure or the cow dung compost of becoming thoroughly decomposed according to volume ratio 20% ~ 30%, and after-ripening 5 ~ 8 days, in After-ripening, turning is 3 ~ 5 times, finally under temperature is no more than the condition of 35 ℃, the water content of biological organic fertilizer is evaporated to below 15%, packing is dispatched from the factory and is the bio-organic fertilizer of preventing and treating rice sheath blight disease.
Wherein step (1) PDA nutrient solution used compound method is, take preparation 1L substratum as example:
After with 200 g potatoes, peeling, be cut into small pieces to be put in water and boil, after boiling, boil 30 min, in filtrate, add after filtering 20 g common sucrose, be settled to 1000 ml, pH value nature, 121 ℃ of sterilizing 20 min.
Wherein the pig manure after used the becoming thoroughly decomposed of step (4) or cow dung compost are germination index more than 98%, quality of organic matter ratio content >=35%, and the organonitrogen quality is 1.2% ~ 2% than content, water content mass ratio 20% ~ 30%.
Described bio-organic fertilizer can be used for preventing and treating rice sheath blight disease.
Beneficial effect
A kind of biological organic fertilizer and production method thereof that can the biological control rice sheath blight disease of the present invention, utilize biocontrol strain fermentation agricultural solid residue to produce biocontrol strain nourishing body and asexual spore, with organic fertilizer, mix the conidial biological organic fertilizer that the nourishing body of the overwhelming majority is converted to dormancy again, its product is compared and is had following advantage with product in the market:
1) rice husk in this fertilizer product and rice straw powder etc. can float over the rice field water surface chronically, the mycelia or the asexual spore that are adsorbed on the biocontrol strain on rice husk and straw powder can form a large amount of aerial hyphaes, effectively suppress dry thread Pyrenomycetes sclerotium in water and infect Culm of Rice, it suppresses rice sheath blight disease pathogenic bacteria effect highly significant, bio-feritlizer can provide the nutrition of biological and ecological methods to prevent plant disease, pests, and erosion thalline simultaneously, for bacterial strain, grows continuously and healthily.
2) this product can effectively avoid simple application biocontrol strain to cause thalline to sink to rice field water surface bottom, thereby suppresses the growth of biocontrol strain, also can avoid biocontrol strain to drop on the plant surface and by ultraviolet ray, high temperature, agricultural chemicals etc., is killed or weaken activity.Test-results shows that the biological control effect of the rice sheath blight disease of this product reaches more than 70%.
3) owing to being biological bacterial strain preparation, the series of problems that brings less than the use because of chemical pesticide fully, be conducive to agriculture nuisanceless production and agricultural sustainable development.
Four, accompanying drawing explanation
Fig. 1 biocontrol strains electromicroscopic photograph, biocontrol strains asexual spore and conidiophore. a asexual spore, b conidiophore;
Fig. 2 biocontrol strains suppresses the effect of rice sheath blight disease pathogenic bacteria, i.e. the restraining effect of biocontrol strains to the rice sheath blight disease pathogenic bacteria, the wherein competitive inhibition of A biocontrol strains, the Spurious suppression effect of B biocontrol strains;
The sickness rate of Fig. 3 different treatment rice sheath blight disease, between different letter representations processing, there were significant differences (
p0.05).
Five, embodiment
Embodiment 1
(1) separation of bacterial strain and evaluation
Gather the samples such as the poor soil of air permeability, beach aleuritic texture soil, diving mud, cryopreservation, adopt the Ma Dingshi substratum to separate fungi, adopts the face-off culture of isolated to the inhibited bacterial strain of rice sheath blight disease pathogenic bacteria dry thread Pyrenomycetes.Biocontrol strains can suppress fully at 72 hours the growth (Fig. 2) of dry thread Pyrenomycetes.
Above-mentioned Ma Dingshi substratum compound method is (take preparation 1L substratum as example): peptone 5g, glucose 10g, KH
2PO
41g, MgSO
47H
2O 0.5g, agar 20g, water 1000ml, pH nature, 1% rose-bengal aqueous solution 3.3ml, 115 ℃ sterilizing 30 minutes, face in every 100ml substratum of used time and add 1% streptomycin solution 0.3ml.
Biocontrol strains belong to trichoderma asperellum (
T. asperellum), Main Biological is: on the PDA substratum, cultivated 3 days for 28 ℃, mycelia can be covered with whole culture dish, mycelial growth initial stage white, felted ，You center to edge forms ring-type gradually, the ring-type that keeps to the side mycelia color is more shallow, on near the centres circle, produce dense sap green conidium, the culture dish of growth mycelia gives out very heavy Coconut Juice aromatic odour, back side deep yellow; Conidiophore, to growth, is arborization, sap green, and the bottle stalk is short, and base portion attenuates, and expand centre; Conidium is spherical in shape, surface irregularity, size is 3.5～5.0 μ m * 2.0～5.0 μ m(Fig. 1); The ITS sequential evolution analysis show this bacterial strain be trichoderma asperellum (
T. asperellum).This biocontrol strain is in preservation on May 16 in 2013 and Chinese Typical Representative culture collection center, and the preservation address is wuchang, wuhan Luo Jiashan, and deposit number CCTCC NO:M 2013213 is called for short trichoderma asperellum CCTCC NO:M 2013213.
(2) microbial inoculum production
1) trichoderma asperellum CCTCC NO:M 2013213 is inoculated in the PDA nutrient solution, carry out liquid fermenting production, the condition of its fermentative production is: initial pH nature, 25 ℃ of culture temperature, dissolved oxygen: the air flow scope is 30～100%, 170rpm, the fermentation middle and later periods forms mycelia fragment, the colony-forming unit of fermented liquid bacterial strain>=1 * 10
8Cfu ml
-1
PDA nutrient solution compound method used is (take preparation 1L substratum as example): after with 200 g potatoes, peeling, be cut into small pieces to be put in water and boil, after boiling, boil 30 min, with after filtered through gauze, filtrate adds 20 g common sucrose, be settled to 1000 ml, the pH value transfers to 6.5 ~ 7.0,121 ℃ of sterilizing 20 min.
2) rice husk, Straw and pig manure be according to 50:30:20 ratio compost, and after 15 ~ 25 days, composting substrate starts cooling, sterilizing 36 hours of 100 ℃ of matrix, high temperature drying, dry after water content of substrate lower than 5%;
3) by the fermented liquid microbial inoculum by the inoculum size of 200 L/ tons be inoculated into oven dry, stable, sterile material carries out solid fermentation, leavening temperature is 20 ~ 35 ℃, in fermenting process, turning is 2 ~ 3 times, ferment and finishes afterwards in 10 ~ 15 days, formation solid fermentation thing.
(3) biological organic fertilizer production
The fermented substrate that will contain trichoderma asperellum CCTCC NO:M 2013213 is inoculated into pig manure or the cow dung compost of becoming thoroughly decomposed according to volume ratio 20 ~ 30%, and after-ripening 5 ~ 8 days, in After-ripening, turning is 3 ~ 5 times, finally under temperature is no more than the condition of 35 ℃, the water content of biological organic fertilizer is evaporated to below 15%, packing is dispatched from the factory and is the bio-organic fertilizer of preventing and treating rice sheath blight disease.
Described bio-organic fertilizer can be used for preventing and treating rice sheath blight disease.
(4) Biological Control Test
1. rice varieties " military fortune round-grained rice 24 ".Process as follows:
T
1: contrast; T
2: the inoculation pathogenic bacteria; T
3: inoculation pathogenic bacteria+biological organic fertilizer
5 repetitions, every repetition 5kg soil/basin are adopted in every processing.
T
1Process and do not inoculate pathogenic bacteria and do not use biological organic fertilizer
T
2200/kg of every basin inoculation dry thread Pyrenomycetes sclerotium soil, after rice transplanting, sclerotium is sprinkled upon on the water surface uniformly;
T
3Every basin is used 200/kg of dry thread Pyrenomycetes sclerotium soil, and after rice transplanting, sclerotium is sprinkled upon on the water surface uniformly, before rice transplanting, uses biological organic fertilizer 10g/kg soil.
After rice transplanting, 5 days start to record incidence, and after 10 days, the biological organic fertilizer that applies rice terrace forms aerial hyphae in water-area, gas boundary, at Culm of Rice and water surface contact position, form the mycelia band, after 20 days, and T
2Process blade and scab clearly, T occur
1And T
3Process blade and scab do not occur.
(5) field plot trial
In Nanjing Jiangning District land for growing field crops, use biological organic fertilizer in 2012 and carry out the test of biological control rice sheath blight disease.
Area 50 square meters in residential quarter arrange test and process: 1, contrast, and application of organic fertilizers or biological organic fertilizer, do not spray jingganmycin; 2, use common fertilizer, after the leveling of field, before rice transplanting, use amount of application 50kg/ mu; 3, use biological organic fertilizer, after the leveling of field, before rice transplanting, use amount of application 50kg/ mu; 4, jingganmycin is processed, and every mu sprays 20% wettable powder 50g.Rice varieties adopts " military fortune round-grained rice 24 ".
After rice transplanting, start regular image data, statistics sickness rate, the difference of calculating different treatment intercurrent disease rate.Morbidity statistics is according to internationally accepted regulations: 0 grade, and blade health; 1 grade, 1% ~ 25% plant leaf morbidity; 2 grades, plant leaf 26% ~ 50% morbidity; 3 grades, plant leaf 51% ~ 75% morbidity; 4, plant 76% falls ill with blade;
Sickness rate=[Σ (morbidity grade * plant quantity)/(total plant quantity * highest ranking)] * 100%
Control rate=[(contrast sickness rate-biological organic fertilizer is processed sickness rate)/contrast sickness rate] * 100%
Calculate CIR before rice harves, test-results (Fig. 3) shows, control treatment and the sickness rate of using the banded sclerotial blight that common fertilizer processes be up to 10.11% ~ 11.05%, and there is no significant difference between two processing (
p0.05); Use biological organic fertilizer and spray jingganmycin process the sickness rate of banded sclerotial blight be between 2.88% ~ 3.03%, two processing, do not have significant difference (
p0.05); Use biological organic fertilizer process and spray jingganmycin process the sickness rate of rice sheath blight disease be markedly inferior to contrast and fertilizer processing (
p0.05).With respect to control treatment, biological organic fertilizer is 70% to the control rate of rice sheath blight disease.Biological organic fertilizer can suppress the generation of rice sheath blight disease significantly.
Claims (6)
1. the biocontrol strains of a prevention and control rice sheath blight disease, it is characterized by this bacterial strain is trichoderma asperellum 12(
Trichoderma asperellum12), on May 16th, 2013 was preserved in Chinese Typical Representative culture collection center, and the preservation address is wuchang, wuhan Luo Jiashan, and the bacterial strain preserving number is CCTCC NO:M 2013213, referred to as trichoderma asperellum CCTCC NO:M 2013213.
2. a kind of biocontrol strains of prevention and control rice sheath blight disease according to claim 1, the Main Biological that it is characterized in that described trichoderma asperellum CCTCC NO:M 2013213 is: on the PDA substratum, cultivated 3 days for 28 ℃, mycelia can be covered with whole culture dish, mycelial growth initial stage white, felted, by center to edge, form gradually ring-type, the ring-type that keeps to the side mycelia color is more shallow, on near the centres circle, produce dense sap green conidium, the culture dish of growth mycelia gives out very heavy Coconut Juice aromatic odour, back side deep yellow; Conidiophore, to growth, is arborization, sap green, and the bottle stalk is short, and base portion attenuates, and expand centre; Conidium is spherical in shape, surface irregularity, and size is 3.5～5.0 μ m * 2.0～5.0 μ m; The ITS sequential evolution analysis shows that this bacterial strain is trichoderma asperellum
T. asperellum.
3. a bio-organic fertilizer, is characterized in that containing 0.5 * 10
8Cfu g
-1The described trichoderma asperellum CCTCC of above claim 1 or 2 NO:M 2013213 bacterial strain statospores, water content lower than 15%, organic content is greater than 35%.
4. the preparation method of a bio-organic fertilizer claimed in claim 3 is characterized in that realizing as follows:
1) trichoderma asperellum CCTCC NO:M 2013213 claimed in claim 1 is inoculated into to the PDA nutrient solution, carry out liquid fermenting production, the condition of its fermentative production is: initial pH nature, 25 ~ 30 ℃ of culture temperature, dissolved oxygen air flow scope is 30～100%, 170rpm, fermentation later stage mycelium pellet all is broken into the mycelia fragment, the colony-forming unit of fermented liquid bacterial strain>=1 * 10
8Cfu ml
-1
2) rice husk, stalk and chicken manure be according to 50:30:20 ratio compost, and after 15 ~ 30 days, rice husk is softening, stalk rots, sterilizing 36 hours of 100 ℃ of matrix, high temperature drying, dry after water content of substrate lower than 5%, form solid fermentation matrix;
3) fermented liquid described in step (1) is inoculated into to the described solid fermentation matrix of step (2) by the inoculum size of 200 L/ tons and carries out solid fermentation, leavening temperature is 20 ~ 35 ℃, and in fermenting process, turning is 2 ~ 3 times, ferments and finishes afterwards in 10 ~ 15 days; Form the solid fermentation thing;
4) step (3) solid fermentation thing is inoculated into to pig manure or the cow dung compost of becoming thoroughly decomposed according to volume ratio 20% ~ 30%, and after-ripening 5 ~ 8 days, in After-ripening, turning is 3 ~ 5 times, finally under temperature is no more than the condition of 35 ℃, the water content of biological organic fertilizer is evaporated to below 15%, packing is dispatched from the factory and is the bio-organic fertilizer of preventing and treating rice sheath blight disease.
5. the preparation method of bio-organic fertilizer according to claim 4, is characterized in that wherein step (1) PDA nutrient solution used compound method is, take preparation 1L substratum as example:
After with 200 g potatoes, peeling, be cut into small pieces to be put in water and boil, after boiling, boil 30 min, in filtrate, add after filtering 20 g common sucrose, be settled to 1000 ml, pH nature, 121 ℃ of sterilizing 20 min.
6. the preparation method of according to claim 4 or 5 described bio-organic fertilizers, it is characterized in that, wherein the pig manure after used the becoming thoroughly decomposed of step (4) or cow dung compost are that germination index is more than 98%, quality of organic matter ratio content >=35%, the organonitrogen quality is 1.2% ~ 2% than content, water content mass ratio 20% ~ 30%.
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| CN104877927A (en) * | 2015-03-18 | 2015-09-02 | 河海大学 | Liquid preparation produced by utilization of livestock and poultry breeding wastewater and application in controlling and eliminating continuous cropping obstacles |
| CN104877927B (en) * | 2015-03-18 | 2017-11-03 | 河海大学 | A kind of liquid preparation of utilization livestock breeding wastewater production and its application in continuous cropping obstacle is prevented and kill off |
| CN108002885A (en) * | 2016-11-01 | 2018-05-08 | 上海交通大学 | A kind of Trichoderma-jinggangmeisu granule and preparation method thereof |
| CN107058126A (en) * | 2017-03-28 | 2017-08-18 | 东北林业大学 | One plant of trichoderma asperellum and its application |
| CN107400682A (en) * | 2017-09-25 | 2017-11-28 | 安徽省中日农业环保科技有限公司 | A kind of biological control method of rice sheath blight disease |
| CN107624794A (en) * | 2017-09-25 | 2018-01-26 | 安徽省中日农业环保科技有限公司 | A kind of microorganism Control Technology of rice sheath blight disease |
| CN107624795A (en) * | 2017-09-25 | 2018-01-26 | 安徽省中日农业环保科技有限公司 | A kind of prevention and controls of rice sheath blight disease |
| US11920126B2 (en) | 2018-03-28 | 2024-03-05 | Ecovative Design Llc | Bio-manufacturing process |
| CN109160861A (en) * | 2018-10-31 | 2019-01-08 | 周口师范学院 | A kind of simple trichoderma-straw bacterial manure preparation method |
| CN110484464A (en) * | 2019-07-31 | 2019-11-22 | 南京宁粮生物肥料有限公司 | It is a kind of for preventing and treating the microorganism formulation and organic fertilizer of the withered line disease of rice |
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