CN103820348A - Plant growth-promoting bacteria and preparation method and application of fungicide - Google Patents
Plant growth-promoting bacteria and preparation method and application of fungicide Download PDFInfo
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- CN103820348A CN103820348A CN201210461410.7A CN201210461410A CN103820348A CN 103820348 A CN103820348 A CN 103820348A CN 201210461410 A CN201210461410 A CN 201210461410A CN 103820348 A CN103820348 A CN 103820348A
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Abstract
The invention relates to a bacillus amyloliquefaciens which can prevent and cure plant disease and promote plant growth, and a low-cost preparation method of fungicide, belonging to the technical field of microbial pesticides and microbial fertilizers. The fungicide is an efficient plant growth-promoting bacteria-bacillus amyloliquefaciens (Bacillus amyloliquefaciens) A3 bacterial strain separated and screened from aquatic plant rhizosphere soil, and is preserved in China General Microbiological Culture Collection Center, with the preservation number of CGMCC No. 6676. In the invention, bacillus amyloliquefaciens is cultured in large scale by using sweet potato starch wastewater as a fermentation culture medium to prepare the fungicide which can prevent and cure plant disease and promote plant growth. The process is low in preparation cost and simple to operate, and can also realize resource utilization of the sweet potato starch wastewater and reduce pollutant discharge. The prepared fungicide, used for preparation of tea leaves and vegetables, can not only improve the yield, but also improve the quality of tea gardens and vegetables, so that the bacillus amyloliquefaciens has wide application prospects.
Description
Technical field:
The present invention relates to the bacillus amyloliquefaciens of a plant height effect antagonism phytopathogen, Promoting plant growth, and low cost production method and using method, microbial pesticide and microbial fertilizer technical field belonged to.
Background technology:
Bacillus amyloliquefaciens (Bacillus amyloliquefaciens) possesses with it diastatic potentiality of producing and gains the name.Bacillus amyloliquefaciens is widely distributed at occurring in nature, is a kind of important biocontrol microorganisms, and it suppresses plant pathogenetic bacteria, fungi, virus, nematode by producing microbiotic, antibacterial protein or polypeptide isoreactivity material, thereby plays the effect of biological control; It is also the important plant rhizosphere growth-promoting bacterium of a class simultaneously, shows the effect that various plants is promoted to growth.For example, the report bacillus amyloliquefaciens FZB45 bacterial strains such as Idriss have phytase (phytase) activity, can stimulating plant growth (Microbiology, 2002,148:2097-2109) under the condition that lacks phosphorus.The report bacillus amyloliquefaciens Ba33 bacterial strains such as Shen Lili can promote tobacco to produce and suppress the propagation (Jilin Agriculture University's journal, 2010,32:383 ~ 386) of tobacco mosaic virus (TMV).Lee is superfine has also reported that a strain has the Bacillus amyloliquefaciens strain DC1(ACTA Scientiae Circumstantiae of algae-lysing, 31:1602-1608).
Chinese invention patent (application number: disclose a bacillus amyloliquefaciens Lx-11 bacterial strain 201010518034.1), deposit number is CGMCC No.3789, bacterial leaf streak of rice is had to good prevention effect, the substratum of cultivating this bacterial strain employing is: soyflour 1%, soybean cake powder 2%, starch 0.5%, yeast extract paste 0.2%, corn steep liquor 0.22%, sodium-chlor 0.1%, Calcium hydrogen carbonate 0.18%.Chinese invention patent (application number: disclose a bacillus amyloliquefaciens YN-1 bacterial strain 200810231406.5), deposit number is CGMCC No.2328, can effectively prevent and treat the fungal disease such as apple marssonina leaf spot, wheat hypochnus, and filter out industrial substratum with low cost, composition comprises: glucose 8～12g/L, Semen Maydis powder 10 ~ 16g/L, soybean cake powder 10 ~ 16g/L.Chinese invention patent (application number: disclose a bacillus amyloliquefaciens HN bacterial strain 200910070709.8), deposit number is CGMCC No.3261, can make nitric nitrogen, nitrite nitrogen, COD equal size in aquaculture water obviously reduce, the cultivation of this bacterial strain adopts broth culture.Chinese invention patent (application number: disclose a plants endogenetic bacillus amyloliquefaciens TF28 bacterial strain 201010264520.5), deposit number is CGMCC No.4038, it can be in plant Colonization inside plants, breeding and conduction, can produce two kinds of antimicrobial substances of antibacterial protein and lipopeptide antibiotic, there is disease prevention growth-promoting effect, the every 1000mL of its liquid nutrient medium by 10g Tryptones, 5g yeast soak powder, 10g NaCl is formulated.Chinese invention patent (application number: disclose a bacillus amyloliquefaciens EA19 bacterial strain 200910273498.8), deposit number is CCTCC M209316, Root Rot of Wheat germ, Root Rot of Wheat leaf spoting bacteria, gaeumannomyces graminis, white wheat thin,tough silk germ and rhizoctonia cerealis are all had to obvious bacteriostatic activity, its fermention medium composition: 20 grams of glucose, 8 grams of extractum carniss, 5 grams, Pidolidone sodium, MgSO
40.5 gram, 0.5 gram of KCl, KH
2pO
41 gram, FeSO
46H
20.15 milligram of O, MnSO
4h
25 milligrams of O, CuSO
45H
20.16 milligram of O, 1000 milliliters, water.Chinese invention patent (application number: disclose a bacillus amyloliquefaciens B-004 bacterial strain 201110095116.4), deposit number is CGMCC No.4557, it is adopted rear early epidemic germ to tomato and has obvious antagonistic effect, in its substratum, carbon source can be Zulkovsky starch, lotus root starch, glucose, lactose, trehalose, sucrose etc., and nitrogenous source can be peptone, extractum carnis, yeast extract paste, NH
4nO
3, NH
4cl, (NH
4)
2sO
4deng, other nutrition sources can be containing metal-salt of potassium, iron, zinc, sodium, manganese, magnesium and calcium and so on etc.
In sum, the bacterial strain of bacillus amyloliquefaciens with antagonism Plant diseases is more, and its cultural method is also different, still, has no the bacterial strain for Tea Production, also has no the report that adopts Sweet Potato Starch Processing Waste Water to cultivate bacillus amyloliquefaciens.
Summary of the invention:
The object of this invention is to provide the bacillus amyloliquefaciens of a plant height effect antagonism phytopathogen, Promoting plant growth, and a kind of method of can recycling Sweet Potato Starch Processing Waste Water producing this microbiobacterial agent is provided, and the using method of microbial inoculum.
Bacterial strain used in the present invention is bacillus amyloliquefaciens (Bacillus amyloliquefaciens) the A3 bacterial strain of being imitated Promoting plant growth, antagonism phytopathogen by contriver's separation screening plant height out from pasture and water rhizosphere soil, this bacterial strain is deposited in China Committee for Culture Collection of Microorganisms's common micro-organisms center on October 15th, 2012, deposit number is: CGMCCNo.6676, depositary institution address: Datun Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica, postcode: 100101.
The production method of bacillus amyloliquefaciens microbiobacterial agent of the present invention is as follows:
1, Sweet Potato Starch Processing Waste Water pre-treatment: regulate pH to 6.8～7.7 of waste water with aqueous sodium hydroxide solution or liming, in 110～126 ℃ of sterilizings 10～35 minutes.
2, liquid seeds preparation: to the bacillus amyloliquefaciens of inoculating slant preservation in the triangular flask of the Sweet Potato Starch Processing Waste Water that pre-treatment is good is housed, in 25～37 ℃, 120～200 revs/min of shaking culture 24～36 hours, obtain liquid seeds.
3, liquid state fermentation: to inoculate volume ratio in the Sweet Potato Starch Processing Waste Water fermentor tank that pre-treatment is good be 2%～15% liquid seeds to being equipped with, and in 25～37 ℃, logical sterile air is cultivated 15～36 hours; Cultured fermented liquid is through count plate, and viable count reaches 1.5 × 10
9more than CFU/ milliliter, gained fermented liquid can be used as liquid bacterial agent and directly uses.
4, solid fungicide preparation: to cross biological carbon (the remaining solid matter after biomass pyrolytic of 30 mesh sieves, main component is carbon) be carrier, according to fermented liquid: biological carbon is the weight ratio of 1: 1～1: 5, fermented liquid is added in biological carbon, fully mix, obtain solid-state microbial inoculum, air seasoning is to microbial inoculum water content below 30%, packing, leaves shady and cool dry place in, for subsequent use.
The using method of bacillus amyloliquefaciens microbiobacterial agent of the present invention is as follows:
1, fermented liquid or solid fungicide being diluted to viable count with irrigation water is 5 × 10
6cFU/ milliliter～5 × 10
7cFU/ milliliter, mixes, and applies near soil crop root according to the consumption ditch of every mu of 1～3 cubic metre of diluent;
2, fermented liquid or solid fungicide are added in fertilizer, mix, making the bacillus amyloliquefaciens viable count in fertilizer is 5 × 10
6cFU/ gram～1 × 10
8cFU/ gram, in being manured into soil as base manure according to the consumption of 1～3 ton every mu.
Embodiment:
Can promote absorption and the utilization of crop to nutrient by bacillus amyloliquefaciens provided by the invention, and can effectively prevent and treat multiple fungi plant disease, the microbiobacterial agent that technical scheme provided by the invention is produced is applied to agriculture production and can reduces the usage quantity of chemical fertilizer and agricultural chemicals, thereby improves farm environment quality; And production cost is very low, and high-concentration waste water is got utilization, reduce pollutant emission.Embodiment is as follows:
Embodiment 1: Sweet Potato Starch Processing Waste Water is produced bacillus amyloliquefaciens microbiobacterial agent
Sweet Potato Starch Processing Waste Water pre-treatment: by liming adjusting wastewater pH to 7.5, in 115 ℃ of sterilizings 20 minutes.
Liquid seeds preparation: to the bacillus amyloliquefaciens of inoculating slant preservation in the triangular flask of the Sweet Potato Starch Processing Waste Water that pre-treatment is good is housed, in 32 ℃, 150 revs/min of shaking culture 26 hours, obtain liquid seeds.
Liquid state fermentation: inoculate 5%(volume ratio to being equipped with in the Sweet Potato Starch Processing Waste Water fermentor tank that pre-treatment is good) liquid seeds, in 32 ℃, logical sterile air is cultivated 20 hours; Cultured fermented liquid is through count plate, and viable count reaches 5.5 × 10
9cFU/ milliliter.
Microbial inoculum preparation: take the biological carbon of crossing 30 mesh sieves as carrier, according to fermented liquid: the weight ratio that biological carbon is 1: 2, fermented liquid is added in biological carbon, fully mix, obtain solid-state microbial inoculum, air seasoning is to microbial inoculum water content below 30%, packing, leaves shady and cool dry place in.
Embodiment 2: bacillus amyloliquefaciens microbiobacterial agent is applied to organic tea and produces
Select tea tree growing way 2 mu of left and right experimental field uniformly at organic tea garden, other tea trees management maintenances of experiment tea tree and base, are just applied fertilizer different always.Experimental field be divided into 5, respectively corresponding 5 processing, between each processing, have the isolation strip that is provided with 2 meters of left and right, by microbial inoculum, with tap water dilution or directly by fermented liquid dilution use, 5 processing are respectively clear water contrast (CK), starch wastewater contrast (FCK), high density (2.5 × 10
7cFU/ milliliter) ditch spread (H), middle concentration (1.8 × 10
7cFU/ milliliter) ditch spread (M), lower concentration (1.2 × 10
7cFU/ milliliter) ditch spread (L), use according to the amount of the microbial inoculum diluents of 3 liters every square metre.Each processing used twice microbial inoculum in an identical manner in the winter time, the tealeaves of gathering spring, result shows: the contrast spring tea output of using wastewater dilution liquid contrasts without significant difference with clear water, and use the spring tea output of microbial inoculum significantly to increase, wherein, the spring tea output of using middle concentration and high density microbial inoculum is suitable, all more than 65%, uses the spring tea output of lower concentration microbial inoculum also than clear water contrast volume increase 30% left and right than clear water contrast volume increase.And use aminoacids content in the tealeaves of microbial inoculum and be significantly higher than clear water contrast.
Embodiment 3: bacillus amyloliquefaciens microbiobacterial agent is used for preventing cucumber samping off
Cucumber samping off is the disease being caused by fungal infection.Gather the soil in the warmhouse booth of planting for many years cucumber, mix, pack in flowerpot, 2 groups of flowerpots are set, every group of 10 basins, in the greenhouse of 15 ℃～20 ℃, keep higher levels of humidity and lower intensity of illumination.The 1st group of direct sowing cucumber seed of flowerpot, 15, every basin, in contrast.In the 2nd multicast kind cucumber seeds forward direction soil, admix weight ratio and be 0.2% bacillus amyloliquefaciens microbiobacterial agent, then 15 seeds of every basin sowing.Statistics sum after coming up, seedling grows after 1 true leaf, statistics lodging seedling sum before 2 true leaves.Result shows, contrasts seedling lodging rate and exceedes 40%, and add the experimental group of microbial inoculum, and seedling lodging rate is lower than 5%.
Claims (3)
1. a strain has the bacillus amyloliquefaciens (Bacillus amyloliquefaciens) of controlling plant diseases and Promoting plant growth, it is characterized in that: this bacillus amyloliquefaciens is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, deposit number: CGMCC No.6676.
2. bacillus amyloliquefaciens claimed in claim 1 is produced the method for microbiobacterial agent, comprises the steps:
(1) Sweet Potato Starch Processing Waste Water pre-treatment: regulate pH to 6.8～7.7 of waste water with aqueous sodium hydroxide solution or liming, in 110～126 ℃ of sterilizings 10～35 minutes;
(2) liquid seeds preparation: to the bacillus amyloliquefaciens of inoculating slant preservation in the triangular flask of the Sweet Potato Starch Processing Waste Water that pre-treatment is good is housed, in 25～37 ℃, 120～200 revs/min of shaking culture 24～36 hours, obtain liquid seeds;
(3) liquid state fermentation: to inoculate volume ratio in the Sweet Potato Starch Processing Waste Water fermentor tank that pre-treatment is good be 2%～15% liquid seeds to being equipped with, and in 25～37 ℃, logical sterile air is cultivated 15～36 hours; Cultured fermented liquid is through count plate, and viable count reaches 1.5 × 10
9more than CFU/ milliliter, gained fermented liquid can be used as liquid bacterial agent and directly uses;
(4) solid fungicide preparation: to cross biological carbon (the remaining solid matter after biomass pyrolytic of 30 mesh sieves, main component is carbon) be carrier, according to fermented liquid: biological carbon is the weight ratio of 1: 1～1: 5, fermented liquid is added in biological carbon, fully mix, obtain solid-state microbial inoculum, air seasoning is to microbial inoculum water content below 30%, packing, leaves shady and cool dry place in.
3. the using method of bacillus amyloliquefaciens microbiobacterial agent claimed in claim 2, comprising:
(1) fermented liquid or solid fungicide being diluted to viable count with irrigation water is 5 × 10
6cFU/ milliliter～5 × 10
7cFU/ milliliter, mixes, and applies near soil crop root according to the consumption ditch of every mu of 1～3 cubic metre of diluent;
(2) fermented liquid or solid fungicide are added in fertilizer, mix, making the bacillus amyloliquefaciens viable count in fertilizer is 5 × 10
6cFU/ gram～1 × 10
8cFU/ gram, in being manured into soil as base manure according to the consumption of 1～3 ton every mu.
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|---|---|---|---|---|
| CN104671416A (en) * | 2015-02-15 | 2015-06-03 | 中国科学院生态环境研究中心 | Device for restoring eutrophic water |
| CN104962492A (en) * | 2015-06-18 | 2015-10-07 | 中国农业大学 | Bacillus amyloliquefaciens, method for preparing solid inoculant thereof and application of solid inoculant |
| CN105176821A (en) * | 2015-09-17 | 2015-12-23 | 商洛学院 | Preparation method of medicinal plant balloonflower growth-promoting microbial inoculant |
| CN105505819A (en) * | 2015-12-19 | 2016-04-20 | 佛山市艳晖生物科技有限公司 | Method for producing amylolytic spore bacterial fertilizer through rice soaking and washing wastewater in sugar industry |
| CN106007950A (en) * | 2016-05-23 | 2016-10-12 | 四川省兰月科技有限公司 | Application of bacillus LY152 to preparing microbial fertilizers |
| CN106167777A (en) * | 2016-08-05 | 2016-11-30 | 四川农业大学 | A kind of cultural method of the bacillus amyloliquefaciens producing Substance |
| CN106434418A (en) * | 2016-08-05 | 2017-02-22 | 四川农业大学 | Culture medium for bacillus amyloliquefaciens culture medium for producing bacteriostatic active substances |
| CN107018995A (en) * | 2017-04-27 | 2017-08-08 | 四川省农业科学院植物保护研究所 | A kind of method that use biocontrol bacterial strain improves organic tea yield |
| CN107034152A (en) * | 2016-10-11 | 2017-08-11 | 天津科技大学 | A kind of method that use sweet potato starch waste liquid prepares azotate pollution soil remediation microbial inoculum N4 |
| CN108558486A (en) * | 2018-01-09 | 2018-09-21 | 中国科学院生态环境研究中心 | A kind of microbial manure and its method using potato starch wastewater preparation |
| CN109020644A (en) * | 2018-09-05 | 2018-12-18 | 河北大学 | Utilize the method and application of Pig raising wastewater fermenting and producing microbial manure |
| CN109456091A (en) * | 2018-09-05 | 2019-03-12 | 河北大学 | Utilize the method and application of cowboying wastewater fermentation production microbial manure |
| CN110117556A (en) * | 2019-03-27 | 2019-08-13 | 海南大学 | A kind of efficient biocontrol microorganisms HN-2 and zymotechnique for producing lipopeptid substance |
| CN110257366A (en) * | 2019-07-05 | 2019-09-20 | 中国农业科学院烟草研究所(中国烟草总公司青州烟草研究所) | It is a kind of based on the bacillus subtilis of charcoal to the control method of tobacco diseases |
| CN114940635A (en) * | 2022-06-08 | 2022-08-26 | 中资蓝天生态环境科技有限公司 | Method for reducing emission of greenhouse gas by organic solid waste semipermeable membrane fermentation and preparing microbial fertilizer |
| CN115125159A (en) * | 2022-04-24 | 2022-09-30 | 沈阳农业大学 | Bacillus amyloliquefaciens and microbial inoculum thereof, and preparation method and application of microbial inoculum |
| CN115777417A (en) * | 2022-12-27 | 2023-03-14 | 中国烟草总公司四川省公司 | Biological control method for preventing and treating tobacco climate spots |
| CN117467577A (en) * | 2022-10-16 | 2024-01-30 | 青岛科技大学 | Sweet potato paste fermentation product and application thereof |
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| CN104671416A (en) * | 2015-02-15 | 2015-06-03 | 中国科学院生态环境研究中心 | Device for restoring eutrophic water |
| CN104962492A (en) * | 2015-06-18 | 2015-10-07 | 中国农业大学 | Bacillus amyloliquefaciens, method for preparing solid inoculant thereof and application of solid inoculant |
| CN105176821A (en) * | 2015-09-17 | 2015-12-23 | 商洛学院 | Preparation method of medicinal plant balloonflower growth-promoting microbial inoculant |
| CN105505819A (en) * | 2015-12-19 | 2016-04-20 | 佛山市艳晖生物科技有限公司 | Method for producing amylolytic spore bacterial fertilizer through rice soaking and washing wastewater in sugar industry |
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| CN110117556A (en) * | 2019-03-27 | 2019-08-13 | 海南大学 | A kind of efficient biocontrol microorganisms HN-2 and zymotechnique for producing lipopeptid substance |
| CN110117556B (en) * | 2019-03-27 | 2022-05-13 | 海南大学 | Biocontrol bacterium HN-2 for producing lipopeptide substances and fermentation process |
| CN110257366A (en) * | 2019-07-05 | 2019-09-20 | 中国农业科学院烟草研究所(中国烟草总公司青州烟草研究所) | It is a kind of based on the bacillus subtilis of charcoal to the control method of tobacco diseases |
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| CN115777417A (en) * | 2022-12-27 | 2023-03-14 | 中国烟草总公司四川省公司 | Biological control method for preventing and treating tobacco climate spots |
| CN115777417B (en) * | 2022-12-27 | 2023-11-21 | 中国烟草总公司四川省公司 | Biological control method for controlling tobacco climate spots |
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