CN112795506A - Bacillus belgii capable of preventing and treating root rot of salvia miltiorrhiza and application thereof - Google Patents
Bacillus belgii capable of preventing and treating root rot of salvia miltiorrhiza and application thereof Download PDFInfo
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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
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
- A01N63/22—Bacillus
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Abstract
The Bacillus beiLeisi capable of producing root rot of red sage root is Bacillus velezensis Bv1-4, which has been preserved in China Center for Type Culture Collection (CCTCC) with the preservation address: the preservation number of the registration book is CCTCC M2020926, and the preservation date is 2020, 12 months and 21 days. The Bacillus belgii has a good inhibiting effect on root rot pathogenic bacteria of salvia miltiorrhiza, has a control effect on root rot diseases of salvia miltiorrhiza of more than 25%, can effectively reduce the incidence of root rot diseases of salvia miltiorrhiza, and has an effect superior to that of the existing common chemical pesticide. The Bacillus belgii screened by the invention can not generate the common problems of chemical pesticides such as drug resistance, pesticide residue, environmental pollution and the like after being used, is healthy and environment-friendly to people and livestock, and meets the requirements of people on ecological agriculture.
Description
Technical Field
The invention relates to Bacillus belgii capable of preventing and treating root rot of salvia miltiorrhiza and application thereof, belonging to the technical field of microorganisms.
Background
Salvia miltiorrhiza Salvia milithiohizaBge is a perennial herb of Labiatae, is taken as a medicine by drying roots and rhizomes, and is one of the first-choice medicines for treating cardiovascular and cerebrovascular diseases. In recent years, the planting of salvia has become an important measure for the poverty elimination and the countryside revivification in poverty-stricken areas in western Henan province. Root rot of salvia miltiorrhiza can cause the yield reduction of salvia miltiorrhiza by 50-60%, and destructive loss can be caused when the disease is serious.
The root rot of salvia miltiorrhiza is the most important disease in salvia miltiorrhiza production, pathogenic bacteria mainly comprise Fusarium proliferatum (Fusarium proliferatum), Fusarium solani (F.solani), Fusarium oxysporum (F.oxysporum) and Alternaria tenuissima (Alternaria tenuissima) of Fusarium, the disease can occur in the whole growth period of the salvia miltiorrhiza, the root of the salvia miltiorrhiza is mainly damaged, and field symptoms are represented as plant rigidity, purple red at the edge of a leaf, yellow after-treatment, and finally withered and shed; the underground part of fibrous roots are firstly attacked to become brown and rotten, the main roots are blackened and rotten due to the spread, and the wood fibers are finally browned and become dry and rotten, so that the yield and the quality of the salvia miltiorrhiza are reduced.
In order to ensure the yield of the salvia miltiorrhiza, most medicinal material growers widely use chemical fertilizers and chemical agents in the planting process of the salvia miltiorrhiza, so that the quality of the salvia miltiorrhiza is reduced to a certain extent, fertilizer damage and chemical damage can also occur under severe conditions, even chemical agent residue exceeds the standard, and meanwhile, the ecological environment of a planting area is polluted.
The utilization of beneficial antagonistic bacteria for biological prevention and soil control is a research hotspot which is concerned at present. At present, the biological mode of microorganisms is mainly relied on to control plant diseases, including bacteria, fungi, actinomycetes and the like. Biological control has been paid attention by students in the aspect of controlling root rot diseases of salvia miltiorrhiza, biocontrol strain screening of pathogenic bacteria F.solani of the root rot disease of salvia miltiorrhiza and microbial inoculum and application thereof have been widely researched, and it has been disclosed that biocontrol strains mainly comprise bacillus subtilis, bacillus amyloliquefaciens, trichoderma and the like, and have less effective biocontrol strains on pathogenic bacteria F.proliferum, F.oxysporum and Alternaria spp.
Disclosure of Invention
Aiming at the defects of the prior art, the invention aims to provide the Bacillus beijerinckii capable of preventing and treating the root rot of the salvia miltiorrhiza, and the screening method and the application of the strain.
The technical scheme adopted by the invention for solving the technical problems is as follows:
the Bacillus beiLeisi capable of producing root rot of red sage root is Bacillus velezensis Bv1-4, which has been preserved in China Center for Type Culture Collection (CCTCC) with the preservation address: the preservation number of the registered book is CCTCC M2020926, and the preservation date is 2020, 12 and 21 days.
The Bacillus beilesensis capable of producing the root rot disease of the salvia miltiorrhiza comprises the following screening methods: collecting salvia miltiorrhiza rhizosphere soil in a salvia miltiorrhiza field, weighing 1g of soil, adding 10ml of sterile water, fully oscillating to obtain a soil suspension, shaking the soil suspension uniformly, diluting, coating 0.2ml of diluent on an LB solid plate, culturing, separating and purifying after bacterial colonies grow out, screening out a strain with an inhibiting effect on salvia miltiorrhiza root rot by using a plate antagonism test, and identifying to obtain bacillus beilesiensis Bv 1-4.
The Bacillus beilesiensis capable of preventing and treating root rot of red sage root is applied to prevention and treatment of root rot of red sage root.
The Bacillus beilesiensis capable of producing the bacterial agent for preventing and treating the root rot of the salvia miltiorrhiza bunge can be applied to preparation of the bacterial agent for preventing and treating the root rot of the salvia miltiorrhiza bunge.
The preparation method of the microbial inoculum comprises the following steps: inoculating Bacillus belgii Bv1-4 into LB culture medium, performing shake culture at 28 deg.C and 180rpm for 12h to prepare seed bacteria, inoculating seed bacteria into LB culture medium at an inoculum size of 2%, and fermenting at 28 deg.C and 180rpm for 48h to form fermentation liquid, i.e. microbial inoculum.
The concentration of live bacillus beilesiensis BvBv1-4 in the fermentation liquor is 107～108CFU/mL。
The LB culture medium is as follows: 10g of tryptone, 5g of yeast extract and 10g of sodium chloride, adding distilled water to a constant volume of 1000mL, uniformly mixing, subpackaging in triangular flasks, and sterilizing at 121 ℃ for 30 min.
The using method of the microbial inoculum comprises the following steps: the microbial inoculum is diluted by 10 times by sterile water to ensure that the viable bacteria concentration of the diluted solution is 106～ 107CFU/ml, irrigating roots around roots of Salvia miltiorrhiza; the root irrigation is carried out 1 time every 7 days, and the root irrigation is carried out 2 times.
The invention has the beneficial effects that: the invention aims to prevent and treat root rot of salvia miltiorrhiza by using microorganisms, so that Bv1-4 capable of preventing and treating root rot of salvia miltiorrhiza is separated from rhizosphere soil of salvia miltiorrhiza through separation identification, plate antagonism and disease-preventing potting experiments and is identified as Bacillus belief (Bacillus velezensis). Tests show that the Bacillus belgii has a good inhibiting effect on root rot pathogenic bacteria of salvia miltiorrhiza, has a control effect on root rot diseases of salvia miltiorrhiza of more than 25%, can effectively reduce the incidence of root rot diseases of salvia miltiorrhiza, and has an effect superior to that of the existing common chemical pesticide. The Bacillus belgii screened by the invention can not generate the common problems of chemical pesticides such as drug resistance, pesticide residue, environmental pollution and the like after being used, is healthy and environment-friendly to people and livestock, and meets the requirements of people on ecological agriculture.
Drawings
FIG. 1 is a graph comparing the results of plate antagonism tests of strain Bv1-4 against Fusarium heterosporum, Fusarium solani, Fusarium oxysporum and Alternaria tenuis.
FIG. 2 is a colony morphology map of the strain Bv 1-4.
FIG. 3 is a graph comparing the results of the allergy test of strain Bv1-4 to tobacco leaves, wherein the left side of the graph is strain Bv1-4, and the right side of the graph is a positive control.
Detailed Description
The present invention will be further described with reference to the following examples and the accompanying drawings.
Isolation of biocontrol bacteria
Weighing 1g of soil, adding 10ml of sterile water, fully oscillating to obtain a soil suspension, then shaking the soil suspension uniformly, diluting, coating 200 mu L of diluent on an LB solid plate, culturing until bacterial colonies grow out, selecting bacterial colonies with different sizes, colors or forms, streaking, separating and purifying on the surface of an LB solid culture medium to obtain 1 strain, which is named as Bv 1-4.
(II) screening of biocontrol bacteria (Flat plate antagonism test)
Respectively inoculating fusarium, fusarium solani, fusarium oxysporum and alternaria tenuis on the root rot pathogenic bacteria layer of salvia miltiorrhiza on a PDA culture medium, culturing for 5 days at 28 ℃, preparing cultured root rot pathogenic bacteria of salvia miltiorrhiza into bacterial cakes by using a perforator with a sterilized diameter of 8mm, connecting the bacterial cakes to one side of a new PDA flat plate (the diameter is 90 mm), sterilizing a paper disc ring with the diameter of 8mm, placing the paper disc ring on the other side of the PDA culture medium, sucking 10 mu l of biocontrol bacteria suspension, and slowly dripping the biocontrol bacteria and the paper disc ring into the paper disc ring, wherein the biocontrol bacteria and the paper disc ring are positioned on the same straight line during inoculation. Sealing with sealing film, culturing at 28 deg.C, recording the size of the antibacterial band when the control pathogenic bacteria grow to the paper disc, and measuring the colony radius of the corresponding pathogenic bacteria on the plate. The contrast is the culture of Fusarium, Fusarium solani, Fusarium oxysporum and Alternaria gracilis on the root rot pathogenic bacteria layer of the salvia miltiorrhiza and the culture of inoculated sterile water paper discs on PDA, the colony treatment is repeated for at least 3 times, and the result of the culture of confrontation is checked after 5 days, and the result is shown in table 1 and figure 1, wherein Bv1-4 has effects on 4 pathogenic bacteria.
Table 1: bacteriostatic effect of Bacillus belgii (Bv1-4) on root rot pathogenic bacteria of salvia miltiorrhiza
(III) identification of biocontrol bacteria
And performing morphological identification on the screened strains with good antagonistic effect, and determining the strains to be the Bellis bacillus subtilis through morphological, physiological and biochemical analysis (see table 2) and molecular biological identification. The Bacillus belgii forms milky white colonies on the culture medium, is opaque, has folds on the surface, slightly bulges in the middle, is uneven on the edge and has strong lifting property (see figure 2).
Table 2: bv1-4 physiological and biochemical characteristics
Note: "-" is negative and "+" is positive.
(IV) sequencing of biocontrol bacteria
Sequencing: the strain Bv1-4 is sequenced, and the identification result is Bacillus belgii (Bacillus velezensis).
The 16SrDNA gene sequencing result is as follows:
SEQUENCE LISTING
(V) preparation of microbial inoculum
Inoculating Bacillus belgii Bv1-4 into LB culture medium, performing shake culture at 28 deg.C and 180rpm for 12h to prepare seed bacteria, inoculating seed bacteria into LB culture medium at an inoculum size of 2%, and fermenting at 28 deg.C and 180rpm for 48h to form fermentation liquid, i.e. microbial inoculum. The concentration of live bacteria of the Bacillus beiLeisi Bv1-4 in the fermentation liquor is 107～108CFU/mL。
(VI) test of tobacco leaf allergy caused by microbial inoculum
Inoculating the lower epidermis of tobacco subcutaneously with prepared microbial inoculum, and inoculating with 10mM MgSO4As a negative control, the wild tobacco fire was used as a positive control (YH). Under the conditions of 24 ℃ and 60% -80% high humidity, allergic necrotic spots appear after 24-48h as positive reaction, and yellow spots appear after 3d as negative reaction. As a result, as shown in FIG. 3, the Bacillus bleekii Bv1-4 anaphylaxis test is a negative reaction, so that the Bacillus bleekii Bv1-4 is nonpathogenic and can be used as a biocontrol bacterium for preventing and treating the root rot of salvia miltiorrhiza.
(VII) disease prevention experiment of biocontrol bacteria
Germinating Saviae Miltiorrhizae radix seed on a tray filled with sterilized soil, transferring into nutrition pot, culturing for 30 days, inoculating biocontrol bacteria by root irrigation method, wetting soil with small amount of clear water before root irrigation, inoculating Bacillus beili Bv1-4 fermentation broth (viable bacteria concentration of 10) at rhizosphere7～108CFU/mL), diluting the fermentation broth 10 times with sterile water, 10mL for each plant, and inoculating 10mL again at intervals of 7 d; the control group is inoculated with LB culture medium (CK) with the same quantity by the same method, in addition, a medicament positive control group is arranged, the control medicament is 1000-time diluent of 50 percent carbendazim wettable powder, 10ml of each salvia miltiorrhiza seedling is irrigated with roots for 1 time after 7 days, and the roots are irrigated for 2 times totally.After the root of each group is irrigated for 3 days for the last time, fusarium solani which is the pathogen of root rot of salvia miltiorrhiza is inoculated. The inoculation method comprises the following steps: irrigating root, inoculating, collecting 10ml root rot pathogenic bacteria spore suspension (concentration is 1 × 10)6Root/ml) of Salvia miltiorrhiza seedlings, watering for 1 time in 3d, culturing in a constant temperature incubator at 28 deg.C, and periodically observing the morbidity of Salvia miltiorrhiza. And after culturing for 30 days, investigating and calculating the morbidity, disease index and prevention and treatment effect. The grading standard of the root rot disease of salvia miltiorrhiza is carried out according to related documents, and the result is shown in table 3.
TABLE 3 Bacillus beilesiensis Pp1-4 potted plant disease-preventing effect on root rot of salvia miltiorrhiza
The result shows that the control effect of the Bacillus beilesensis Bv1-4 on the root rot of the salvia miltiorrhiza is 63.3%, and the effect is obvious.
The foregoing description is only a preferred embodiment of the present invention, and various modifications and changes will occur to those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Sequence listing
<110> institute of plant protection of academy of agricultural sciences of Henan province
<120> Bacillus belgii capable of preventing and treating root rot of salvia miltiorrhiza and application thereof
<141> 2021-01-07
<160> 1
<170> SIPOSequenceListing 1.0
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accagagagt cgccttcgcc actggtgttc ctccacatct ctacgcattt caccgctaca 840
cgtggaattc cactctcctc ttctgcactc aagttcccca gtttccaatg accctccccg 900
gttgagccgg gggctttcac atcagactta agaaaccgcc tgcgagccct ttacgcccaa 960
taattccgga caacgcttgc cacctacgta ttaccgcggc tgctggcacg tagttagccg 1020
tggctttctg gttaggtacc gtcaaggtgc cgccctattt gaacggcact tgttcttccc 1080
taacaacaga gctttacgat ccgaaaacct tcatcactca cgcggcgttg ctccgtcaga 1140
ctttcgtcca ttgcggaaga ttccctactg ctgcctcccg taggagtctg ggccgtgtct 1200
cagtcccagt gtggccgatc accctctcag gtcggctacg catcgttgcc ttggtgagcc 1260
gttacctcac caactagcta atgcgccgcg ggtccatctg taagtggtag ccaaagccac 1320
cttttatgtc tgaaccatgc ggttcaaaca accatccggt attagccccg gtttcccgga 1380
gttatcccag tcttacaggc aggttaccca cgtgttactc acccgtccgc cgctaacatc 1440
agggagcaag ctcccatctg tccgctcgac ttgcatgtat taggcacgcc gccagcgttc 1500
gtcctgagcc atgatcaaac tct 1523
Claims (7)
1. The Bacillus beiLeisi capable of producing root rot of red sage root is Bacillus velezensis Bv1-4, which has been preserved in China Center for Type Culture Collection (CCTCC) with the preservation address: the preservation number of the registration book is CCTCC M2020926, and the preservation date is 2020, 12 months and 21 days.
2. The bacillus beilesensis capable of producing the compound capable of preventing and treating root rot of salvia miltiorrhiza according to claim 1, wherein the screening method comprises the following steps: collecting salvia miltiorrhiza rhizosphere soil in a salvia miltiorrhiza field, weighing 1g of soil, adding 10ml of sterile water, fully oscillating to obtain a soil suspension, shaking the soil suspension uniformly, diluting, coating 0.2ml of diluent on an LB solid plate, culturing, separating and purifying after bacterial colonies grow out, screening out a strain with an inhibiting effect on salvia miltiorrhiza root rot by using a plate antagonism test, and identifying to obtain bacillus beilesiensis Bv 1-4.
3. The Bacillus belgii capable of producing the root rot disease prevention and treatment effects on Salvia miltiorrhiza Bunge according to claim 1 or 2, wherein the Bacillus belgii is applied to the prevention and treatment effects on the root rot disease of Salvia miltiorrhiza Bunge.
4. The Bacillus belgii capable of producing bacterial agents for preventing and treating root rot of Salvia miltiorrhiza according to claim 1 or 2, wherein the Bacillus belgii is used for preparing bacterial agents for preventing and treating root rot of Salvia miltiorrhiza.
5. The use of claim 4, wherein the preparation method of the microbial inoculum comprises the following steps: inoculating Bacillus belgii Bv1-4 into LB culture medium, performing shake culture at 28 deg.C and 180rpm for 12h to prepare seed bacteria, inoculating seed bacteria into LB culture medium at an inoculum size of 2%, and fermenting at 28 deg.C and 180rpm for 48h to form fermentation liquid, i.e. microbial inoculum.
6. The method for preparing the microbial inoculum according to claim 5, wherein the concentration of the live bacillus belgii Bv1-4 in the microbial inoculum is 107～108CFU/mL。
7. The use of claim 5, wherein the microbial inoculum is used by the method comprising the following steps: the microbial inoculum is diluted by 10 times by sterile water to ensure that the viable bacteria concentration of the diluted solution is 106～107CFU/ml, irrigating roots around roots of Salvia miltiorrhiza; the root irrigation is carried out 1 time every 7 days, and the root irrigation is carried out 2 times.
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Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105543132A (en) * | 2015-12-31 | 2016-05-04 | 河南省农业科学院植物保护研究所 | Bacillus methylotrophicus YB-F7 and application thereof in preventing plant diseases |
| CN107446847A (en) * | 2017-08-14 | 2017-12-08 | 云南农业大学 | One plant of Bei Laisi bacillus GT11 and its application |
| CN109576188A (en) * | 2018-12-28 | 2019-04-05 | 天津开发区坤禾生物技术有限公司 | A kind of prevention and treatment tuber of pinellia root rot microbial inoculum and its preparation method and application |
| US20190230938A1 (en) * | 2015-05-22 | 2019-08-01 | Institut National De La Recherche Scientifique | Bacterial And Fungal Metabolites Possessing Anti-Microbial Activity Against Xanthomonas Species, Compositions, Methods, Kits And Uses Relating To Same |
| KR102078252B1 (en) * | 2018-12-18 | 2020-02-17 | 경기도 | Bacillus velezensis ARRI17 strain having antifungal activity against Root Rot Pathogen of ginseng and Use Thereof |
| CN111471624A (en) * | 2020-04-25 | 2020-07-31 | 浙江师范大学 | Bacillus belgii CSQXDZ26 strain and application thereof |
-
2021
- 2021-01-07 CN CN202110019594.0A patent/CN112795506B/en not_active Expired - Fee Related
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20190230938A1 (en) * | 2015-05-22 | 2019-08-01 | Institut National De La Recherche Scientifique | Bacterial And Fungal Metabolites Possessing Anti-Microbial Activity Against Xanthomonas Species, Compositions, Methods, Kits And Uses Relating To Same |
| CN105543132A (en) * | 2015-12-31 | 2016-05-04 | 河南省农业科学院植物保护研究所 | Bacillus methylotrophicus YB-F7 and application thereof in preventing plant diseases |
| CN107446847A (en) * | 2017-08-14 | 2017-12-08 | 云南农业大学 | One plant of Bei Laisi bacillus GT11 and its application |
| KR102078252B1 (en) * | 2018-12-18 | 2020-02-17 | 경기도 | Bacillus velezensis ARRI17 strain having antifungal activity against Root Rot Pathogen of ginseng and Use Thereof |
| CN109576188A (en) * | 2018-12-28 | 2019-04-05 | 天津开发区坤禾生物技术有限公司 | A kind of prevention and treatment tuber of pinellia root rot microbial inoculum and its preparation method and application |
| CN111471624A (en) * | 2020-04-25 | 2020-07-31 | 浙江师范大学 | Bacillus belgii CSQXDZ26 strain and application thereof |
Non-Patent Citations (3)
| Title |
|---|
| LI MA ET AL.: "Phylogenetic diversity of bacterial endophytes of Panax notoginseng with antagonistic characteristics towards pathogens of root-rot disease complex", 《ANTONIE VAN LEEUWENHOEK》 * |
| 何碧珀等: "2 株芝麻真菌病害生防菌的抑菌特征、促生评价及鉴定", 《河南农业科学》 * |
| 姚锦爱等: "贝莱斯芽胞杆菌ZZBV-3的鉴定及其对草莓根腐病的防效", 《中国生物防治学报》 * |
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