CN104342394B - One plant of Siam bacillus and its application in terms of preventing and treating Fusarium graminearum - Google Patents
One plant of Siam bacillus and its application in terms of preventing and treating Fusarium graminearum Download PDFInfo
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- CN104342394B CN104342394B CN201410602407.1A CN201410602407A CN104342394B CN 104342394 B CN104342394 B CN 104342394B CN 201410602407 A CN201410602407 A CN 201410602407A CN 104342394 B CN104342394 B CN 104342394B
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
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Abstract
The invention discloses one plant of Siam bacillus and its applications in terms of preventing and treating Fusarium graminearum.One plant of Siam bacillus disclosed by the invention, deposit number are CGMCC No.9501.As the biological control material of wheat scab, new biological pesticide or biological control microbial inoculum no matter are developed, which all has a good application prospect.
Description
Technical field
The invention belongs to field of biotechnology, and in particular to a kind of Siam bacillus, the invention further relates to a kind of Siam
Bacillus and its application in terms of preventing and treating Fusarium graminearum.
Background technique
Fusarium graminearum (Fusarium graminearum) is one of important pathogen of gramineous crop, can cause wheat
Bakanae disease (Fusarium Head Blight, FHB), Fusarium graminearum are a principal element for limiting wheat yield.I
State's weather conditions are advantageous to the generation of gibberella zeaze petch of wheat and barley, and the long-term disease in In Middle And Lower Reaches of Changjiang River causes production loss 10%-
15%, the time underproduction nearly 50% of being very popular.Since the cropping systems such as global warming and straw-returning, no-tillage cultivation change
Influence, the generation area of China's gibberella zeaze petch of wheat and barley by the Yangtze river basin rapidly northwestwards, North China extend, especially 2008,
2010 and 2012, national gibberella zeaze petch of wheat and barley generation was extremely serious, caused serious production grain loss and undesirable social influence,
Seriously threaten China's grain security.
The deoxynivalenol that Fusarium graminearum generates is the main day being present in gibberellic hypha infection wheat
One of right toxin.DON has been identified as most dangerous one of naturally-occurring food contaminant, is put into international research preferentially
Position.DON can cause people's immunity degradation, anaemia, headache, abdominal pain, nausea；Activate object food refusal, growth retardation and miscarriage.DON is extensive
In the presence of and largely pollute wheat and its product, very big harm is constituted to people and animals, seriously threatens China's food safety.
Therefore, reinforcement Fusarium graminearum prevention and control research, which has become, ensures that China's grain security and the urgent of food safety are essential
It asks.
Chemical pesticide relied primarily on to gibberella zeaze petch of wheat and barley prevention and treatment for many years, but with chemical pesticide bring environmental problem,
In addition many years medication has led to the appearance of pathogen drug resistance Fusarium graminearum bacterial strain, increases difficulty of prevention and cure.Although red mould
A large amount of energy is put into terms of sick resistance quality both at home and abroad and carries out correlative study, but there is no ideal highly resistance wheat breed so far.
Biological control not only reduced environmental impact, is also less prone to develop drug resistance, the research of correlation theory and technology
It is increasingly concerned.Bacillus siamensis nutrition is simple, is widespread in nature, to the nontoxic nothing of people and animals
Evil, it is free from environmental pollution, a variety of antibiotic and enzyme can be generated, there is broad spectrum antibiotic activity and extremely strong anti-adversity ability, it is micro- compared with other
Biology is the potentiality of biological prevention and control microbial inoculum with more exploitation.
Summary of the invention
The object of the present invention is to provide one plant of Siam bacillus and its applications in terms of preventing and treating Fusarium graminearum.
One plant of Siam bacillus provided by the invention, the bacterial strain are preserved in China Microbiological bacterium on July 18th, 2014
Kind preservation administration committee common micro-organisms center, deposit number are CGMCC No.9501.
Application of the above-mentioned bacterial strains in the plant disease caused by biological control sickle-like bacteria also belongs to protection scope of the present invention.
In above-mentioned application, sickle-like bacteria is Fusarium graminearum.
In above-mentioned application, disease is head blight or ear rot or root rot caused by sickle-like bacteria.
In above-mentioned application, plant is cereal；Cereal is wheat crops or corn；Wheat crops are specially wheat.
Application of the above-mentioned bacterial strains in the product that preparation inhibits sickle-like bacteria also belongs to protection scope of the present invention.
The present invention also provides a kind of methods of plant disease caused by prevention and treatment sickle-like bacteria, are sprayed in growing process
Mist processing；It is by spraying the bacteria suspension of above-mentioned Siam bacillus or fermentation liquid or metabolite.
In the above method, sickle-like bacteria is Fusarium graminearum.
In above-mentioned method, disease is head blight or ear rot or root rot.
In above-mentioned method, plant is cereal；Cereal is wheat crops or corn；Wheat crops are specially wheat.
The beneficial effects of the present invention are: Siam bacillus (Bacillus siamensis) provided by the invention not only exists
Fusarium graminearum can efficiently be inhibited when plate culture, and its control efficiency is up to 98.0% in greenhouse experiments.As wheat
No matter the biological control material of head blight develops new biological pesticide or biological control microbial inoculum, which all has answers well
Use prospect.
Detailed description of the invention
Fig. 1 is opposite culture experiment of the invention.
Specific embodiment
The following describes the present invention in detail with reference to the accompanying drawings and specific embodiments.
Experimental method used in following embodiments is conventional method unless otherwise specified.Institute in following embodiments
Material, reagent etc., are commercially available unless otherwise specified.
NB fluid nutrient medium: it is made of solvent and solute；Solute is peptone, beef extract and NaCl, and solvent is water；Albumen
Concentration of the peptone in NB fluid nutrient medium is 1%, and concentration of the beef extract in NB fluid nutrient medium is 0.3%, NaCl in NB liquid
Concentration in body culture medium is 0.5%, and the % is quality percent by volume (g/100ml).
NA solid medium: in NB fluid nutrient medium be added agar (ratio of agar and fluid nutrient medium be 1.5g:
100ml), NA solid medium is obtained.
PDA culture medium: a small amount of boiling 20min is added in potato 200g, and three layers of filtered through gauze take filtrate water to be settled to
20g glucose, 15g agar, 121 DEG C of high pressure sterilization 20min are added in 1L.
Sweet mung bean soup culture medium: a small amount of boiling 20min is added in mung bean 10g, and three layers of filtered through gauze take filtrate water to be settled to
1L, 121 DEG C of high pressure sterilization 20min after packing.
Phosphate buffer: it is made of solvent and solute；Solute is potassium dihydrogen phosphate and magnesium chloride, and solvent is water；Phosphoric acid
The concentration of potassium dihydrogen is 0.004%, and the concentration of magnesium chloride is 0.019%, and the % is quality percent by volume (g/
100ml)。
Fusarium graminearum (Fusarium graminearum) FG1In document " Yang Huiyong, Li Feifeng, Lu Qiongxian, Xu Jian
Macro, Shi Jianrong antagonistic strain AFR0406 is scientific to the biological activity determination Jiangsu's agriculture of fusarium graminearum and sheath blight fungus,
2006 (06) are disclosed in 142-144. ", and the public can obtain from Institute of Agricultural Product Processing, Chinese Academy of Agricultural Sc.
New 828 kind of wheat stone is in document " new 828 breeding of Sun Zhijun, Yang Xiaoli, Zhang Hui, Zhang Guanghui new variety of wheat stone
Process and high-yield culture technique modern agriculture are scientific and technological, 2011 (09), are disclosed in 96-97. ", the public can be from Scientia Agricultura Sinica
Processing of farm products research institute, institute obtains.
The separation and identification of 1 bacterium of embodiment
One, the separation of bacterium
The wheat disease fringe that 10 fringe Jiangsu acquire is placed on the sterile steaming of 100ml in superclean bench by June, (one) 2012
It is shaken 15 minutes in distilled water and prepares bacteria suspension, then 80 DEG C water bath processing 10 minutes.
(2) it is coated on after bacteria suspension being carried out concentration gradient dilution with sterile distilled water on NA culture medium flat plate, 30 DEG C of items
It is cultivated 24 hours under part, bacterium colony is covered with entire plate, different with form, size, color, transparency on oese picking plate
Bacterial strain flat board scribing line purifying, point connect bacterial strain after purification and are applied to Fusarium graminearum opposite culture experiment (as shown in Figure 1).Will
To the one plant bacterium strong to Fusarium graminearum antagonistic ability be named as JS29I.
Two, it identifies
(1) according to " Berger bacterial identification manual " (the 8th edition) and " common bacteria system identification handbook " (east show pearl, Cai
Wonderful English etc. is write, Beijing: Science Press, 2001.2) it is special to carry out morphological feature, culture to bacterial strain JS29I for method described in
Property and physio-biochemical characteristics identification, concrete outcome is as follows:
The form and physio-biochemical characteristics of thallus:
In NA culture medium, 28 DEG C are cultivated 1 day, and thallus is rod-shaped, and diameter is about 1.5-2.5 μm of 0.4-0.5 μ m；Bacterium colony milk is white
Color, round, dimpling, neat in edge.
Physiological and biochemical property:
Oxidizing ferment :+；Arginine dihydrolase :-；Gelatin hydrolysis :+；Starch Hydrolysis :+；Catalase :+；Three butyric acid are sweet
Grease hydrolysis :-；Nitrate reduction :+；Aesculin hydrolysis :+；Casein hydrolysis :+；Tween80 hydrolysis :-；Indoles experiment :-；Urea
Enzyme :-；Citric acid utilizes :-；H2S is generated :-；VP experiment :+；Anaerobic growth :+；10%NaCl:+；50 DEG C :+；55 DEG C :+；
PH4.0:-.
It can use glycerol, L-arabinose, D-ribose, D- xylose, D-Glucose, D-Fructose, inositol, mannitol, mountain
Pears alcohol, Alpha-Methyl-D-Glucose glycosides, N-acetyl-glucosamine, amarogentin, arbutin, aesculin, salicin, D- cellobiose,
D-Maltose, D- lactose, D- melibiose, D- sucrose, D- melezitose, D- raffinose, starch, glycogen and D- gentiobiose produce acid.
It is poor using L- arabite acid producing ability.
(2) 16S rDNA is tested
The total DNA for extracting JS29I carries out PCR amplification using bacterial 16 S rDNA universal primer, obtains using it as template
Amplified production is recycled and is sequenced by the amplified production that length is about 1.4kb, and the sequence measured is as shown in SEQ ID No.1.
Compared according to Gen-Bank sequence homology, bacterial strain JS29I and Bacillus amyloliquefaciens RNB-
113 (GenBank accession number AB971204.1) homologys are 99%, with Bacillus amyloliquefaciens SQR9
(GenBank accession number CP006890.1) homology is 99%, and the preliminary judgement bacterium is bacillus (Bacillus
sp.)。
Based on features above, bacterial strain JS29I is accredited as Bacillus siamensis.It is recommended that classification naming be Siam
Bacillus；The bacterial strain is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms on July 18th, 2014
Center (abbreviation CGMCC 9501, address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Institute of Microorganism, Academia Sinica,
Postcode 100101), deposit number is CGMCC No.9501.
Antagonism of the embodiment 2Bacillus siamensis JS29I to Fusarium graminearum
Using opposite culture method detection Bacillus siamensisJS29I to the antagonism of Fusarium graminearum.
One, the punch for being 5mm with diameter is in cultured disease fungus Fusarium graminearum (Fusarium
graminearum)FG1Colony edge, which is beaten, takes bacterium disk, is chosen bacterium disk into PDA plate center with the toothpick of sterilizing.
Two, at away from PDA plate center 25mm four chain-dotted lines inoculation same volume and concentration JS29I, to be only inoculated with standing grain
The PDA plate of paddy sickle-like bacteria bacterium disk is control group, and 28 DEG C are cultivated 4-5 days, and whether there is or not antibacterial bands to be formed for observation.Each group is done three and put down
Row.The results are shown in Table 1.Show that JS29I has very strong inhibitory effect to Fusarium graminearum.
Inhibiting effect of the JS29I to Fusarium graminearum after table 1 is cultivated 5 days
(shortest distance that antibacterial bandwidth refers to pathogen edge to antagonistic strain edge)
Inhibitory effect of the embodiment 3JS29I fermentation liquid to Fusarium graminearum
Using opposite culture method detection Bacillus siamensis JS29I to the inhibiting effect of Fusarium graminearum.
One, the punch for being 5mm with diameter is in cultured disease fungus Fusarium graminearum (Fusarium
graminearum)FG1Colony edge, which is beaten, takes bacterium disk, is chosen bacterium disk into PDA plate center with the toothpick of sterilizing.
Two, at away from PDA plate center 25mm 4 points punching inoculation same volume and concentration JS29I, to be only inoculated with standing grain
The PDA plate of paddy sickle-like bacteria bacterium disk is control group, and 28 DEG C are cultivated 4-5 days, and whether there is or not antibacterial bands to be formed for observation.Each group is done three and put down
Row.The results are shown in Table 1.Show that JS29I has very strong inhibitory effect to Fusarium graminearum.
Inhibiting effect of the JS29I to Fusarium graminearum after table 2 is cultivated 5 days
(shortest distance that antibacterial bandwidth refers to pathogen edge to antagonistic strain edge)
Greenhouse experiments of the embodiment 4Bacillus siamensis JS29I to Fusarium graminearum inhibitory effect
One, 28 DEG C of shaken overnight cultures in NB culture medium by JS29I, use phosphate buffer to be diluted to concentration as 2
×108cfu/ml。
Two, by Fusarium graminearum FG1With sweet mung bean soup culture medium shaken cultivation (28 DEG C, 5 days), bacteria suspension is obtained, using phosphorus
It is 2 × 10 that phthalate buffer, which dilutes bacterial concentration,5cfu/ml。
Three, by the JS29I and Fusarium graminearum FG after dilution1It is mixed according to the ratio of 1:1, final concentration is respectively 108cfu/
Ml and 105cfu/ml。
Four, be inoculated in the middle part of the wheat head in the flowering early stage of new 828 kind of stone, connecing thallus product is 10 μ l, as experimental group.
Simultaneously only to contain the phosphate buffer of Fusarium graminearum as positive controls, using be only inoculated with phosphate buffer as negative
Property control.Every group sets 10 repetitions, and experiment is repeated 3 times, and results are averaged.
After Fusarium graminearum bacterial suspension inoculation 16 days, the morbidity of the wheat head is observed, has the wheat head is withered to whiten or rubescent, internal seed
The wheat head that grain is not full, weight saving shows is the morbidity wheat head.
Wherein head blight index=disease incidence × disease index/100.
Greenhouse control efficiency of the 3 Bacillus siamensis JS29I of table to Fusarium graminearum
The results are shown in Table 3, and Bacillus siamensis JS29I can prevent and treat cereal reaping hook very well under greenhouse experiment
Microbial wheat scab.
Claims (10)
1. one plant of Siam bacillus, which is characterized in that the bacterial strain is preserved in Chinese microorganism strain guarantor on July 18th, 2014
Administration committee's common micro-organisms center is hidden, deposit number is CGMCC No.9501.
2. application of the bacterial strain described in claim 1 in the plant disease caused by biological control sickle-like bacteria；The sickle-like bacteria is
Fusarium graminearum.
3. application according to claim 2, it is characterised in that: the disease is head blight or ear rot caused by sickle-like bacteria
Or root rot.
4. application according to claim 2 or 3, it is characterised in that: the plant is cereal；The cereal is wheat crops
Or corn.
5. application according to claim 2 or 3, it is characterised in that: the wheat crops are wheat.
6. application of the bacterial strain described in claim 1 in the product that preparation inhibits sickle-like bacteria；The sickle-like bacteria is cereal reaping hook
Bacterium.
7. a kind of method of plant disease caused by prevention and treatment sickle-like bacteria, which is characterized in that be sprayed in growing process
Processing；Described be the bacteria suspension or fermentation liquid of Siam bacillus described in claim 1 by spraying；The sickle-like bacteria is cereal sickle
Knife bacterium.
8. according to the method described in claim 7, it is characterized by: the disease is head blight or ear rot or root rot.
9. method according to claim 7 or 8, it is characterised in that: the plant is cereal；The cereal is wheat crops
Or corn.
10. according to the method described in claim 9, it is characterized by: the wheat crops are wheat.
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| CN105505834B (en) * | 2016-01-25 | 2019-01-15 | 北京市辐射中心 | Siam bacillus and its application of one plant of prevention and treatment black rot of pear and soft rot |
| CN106282067B (en) * | 2016-08-24 | 2019-10-11 | 李丽丽 | Multifunctional agricultural complex micro organism fungicide and probiotics and application |
| CN107034158B (en) * | 2017-04-27 | 2019-11-19 | 中国科学院微生物研究所 | Siam bacillus B-3 and its application |
| CN108849986B (en) * | 2018-07-09 | 2020-09-15 | 江苏师范大学 | Application of bacillus simmer in preventing and treating white rot |
| CN109294945B (en) * | 2018-09-19 | 2021-11-05 | 中国科学院成都生物研究所 | Wheat endophyte and application thereof |
| CN110964654A (en) * | 2018-09-29 | 2020-04-07 | 福建省农业科学院农业生物资源研究所 | Bacillus antagonistic to fusarium wilt and application thereof |
| CN109207404B (en) * | 2018-10-23 | 2022-02-18 | 山西农业大学 | Siam bacillus YJ15 and application thereof |
| CN109468243B (en) * | 2018-11-13 | 2021-07-30 | 河北农业大学 | Bacillus siamensis and application thereof |
| CN109362787A (en) * | 2018-12-07 | 2019-02-22 | 江苏师范大学 | Application of Siam bacillus in prevention and treatment nematode |
| CN110055186B (en) * | 2019-02-20 | 2022-05-13 | 山东省花生研究所（山东省农业科学院花生工程技术研究中心） | Microbial preparation for degrading vomitoxin and application |
| CN111560327B (en) * | 2020-04-28 | 2021-12-10 | 湖北大学 | Preparation and application of alcaligenes faecalis capable of antagonizing fusarium graminearum and efficiently degrading DON (Don) |
| CN113201478B (en) * | 2021-05-27 | 2022-07-29 | 广西民族大学 | Siamese bacillus Gxun-36 secreting iron carrier and application thereof |
| CN113462621B (en) * | 2021-09-03 | 2021-12-07 | 佛山市玉凰生态环境科技有限公司 | Bacillus siamensis capable of degrading grease and application thereof in grease-containing wastewater |
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Effective date of registration: 20220119 Address after: Room 320, floor 3, building 49, yard 2, Yuanmingyuan West Road, Haidian District, Beijing 100193 Patentee after: Beijing Zhongnong taste detection technology Co.,Ltd. Address before: 100193 No. 2 Old Summer Palace West Road, Beijing, Haidian District Patentee before: INSTITUTE OF FOOD SCIENCE AND TECHNOLOGY, CHINESE ACADEMY OF AGRICULTURAL SCIENCES |
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