CN116987615B - Lecaniella albopictus YZ-151 and application thereof - Google Patents
Lecaniella albopictus YZ-151 and application thereof Download PDFInfo
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
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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
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01P—BIOCIDAL, PEST REPELLANT, PEST ATTRACTANT OR PLANT GROWTH REGULATORY ACTIVITY OF CHEMICAL COMPOUNDS OR PREPARATIONS
- A01P3/00—Fungicides
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- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
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- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
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Abstract
A strain of Listeria albopictus (Priestiaaryabhattai) YZ-151 and application thereof relate to the technical field of microbial antagonistic bacteria, and the strain YZ-151 is preserved in the microorganism strain preservation center of Guangdong province at the month of 02 and 20 of 2023, with the preservation number of GDMCCNo:62894. The bacterial strain YZ-151 has the bacteriostasis rate of more than 78% on acanthopanax black spot bacteria and gray mold bacteria; the bacteria-free filtrate of the strain YZ-151 has the bacteria inhibition rate of 48.06 percent and 57.62 percent on acanthopanax black spot bacteria and gray mold bacteria respectively; the strain YZ-151 has an inhibition effect on common fungal pathogens and shows broad-spectrum antibacterial properties; the strain YZ-151 has good acanthopanax leaf surface colonization capability and good control effect on acanthopanax black spot and gray mold.
Description
Technical Field
The invention relates to the technical field of microbial antagonistic bacteria, in particular to an Aleplerenone (Priestia aryabhattai) YZ-151 strain and application thereof.
Background
Acanthopanax senticosus Acanthopanax senticosus (rupr. Et maxim.) Harms, named cortex Acanthopancis, cantonese, radix Zanthoxyli, and radix Et caulis Acanthopanacis Senticosi, is a deciduous shrub of Acanthopanax of Araliaceae. The dry root, rhizome or stem of acanthopanax root, root bark, leaf and fruit can be used as medicine, and has the effects of replenishing qi to invigorate spleen, improving intelligence and tranquillizing. The acanthopanax root contains various active ingredients and minerals, and the modern pharmacological research results show that the acanthopanax root has pharmacological activities of resisting cancer, protecting liver, resisting inflammation, reducing blood pressure and the like.
With the continuous deep research of the pharmacological research and the chemical research of the acanthopanax in recent years by students at home and abroad, the medicinal demand of the acanthopanax increases year by year, the wild resources are severely reduced, the resources are short and the development of the acanthopanax industry is limited. In order to protect and develop the acanthopanax root resource, the sustainable utilization of the acanthopanax root is realized, and the artificial cultivation of the acanthopanax root is necessarily selected. However, with the expansion of the cultivation area and cultivation area of the acanthopanax, the disease problem of the acanthopanax limits the yield and quality of the acanthopanax. Acanthopanax black spot and gray mold are two destructive foliar spread diseases causing the loss of the yield and the quality of acanthopanax. However, conventional control methods such as pesticides and chemical control do not achieve the desired effect. Thus, the use of beneficial microbial antagonists is considered a promising strategy for managing these diseases.
Biological control is a green, safe and pollution-free control means, beneficial microorganisms inhibit the growth and reproduction of germs through antagonism, competition, production of antibacterial substances and other action mechanisms, and pathogenic germs are not easy to generate drug resistance, so that the biological control is of great interest in green planting of traditional Chinese medicinal materials. Therefore, finding a disease prevention and control way with environment-friendly characteristics is an important problem to be solved in the production of acanthopanax.
At present, research on prevention and treatment of acanthopanax black spot and gray mold diseases by utilizing the listeria albopictus Priestia aryabhattai has not been reported.
Disclosure of Invention
The invention aims to provide an Agrimonia albopictus (Priestia aryabhattai) YZ-151 and application thereof.
The technical scheme adopted by the invention for solving the technical problems is as follows:
The invention discloses an Agrimonia azedarach (Priestia aryabhattai) YZ-151, which is preserved in the microorganism strain preservation center of Guangdong province at the year 2023 and the month 20, wherein the preservation number is as follows: GDMCC No:62894.
The invention relates to an application of an Agrimonia aze-151 strain (Priestia aryabhattai) in preparing plant pathogenic bacteria antagonists.
As a preferred embodiment, the plant pathogenic bacteria include: alternaria tenuissima, botrytis cinerea, fusarium solani, sclerotinia sclerotiorum, alternaria tenuissima, leuconostoc and Fusarium oxysporum.
In a preferred embodiment, the plant pathogenic bacteria antagonist is an acanthopanax pathogenic bacteria antagonist.
As a preferred embodiment, the active ingredient of the plant pathogenic bacteria antagonist is a bacterial suspension or sterile filtrate of Lecaniella albopictus (Priestia aryabhattai) YZ-151.
The invention discloses an application of an Alternaria alternata (Priestia aryabhattai) YZ-151 in preventing and treating acanthopanax black spot.
The invention discloses application of a strain of Agrimonia azedarach (Priestia aryabhattai) YZ-151 in preventing and treating gray mold of acanthopanax.
The beneficial effects of the invention are as follows:
In the invention, a strain YZ-151 is screened from healthy acanthopanax root soil, identified as Agrimonia arundinacea (Priestia aryabhattai) and deposited in the microorganism strain collection of Guangdong province at the year of 2023, 02 and 20, which is abbreviated as GDMCC, and the address is: building 5, 30 # of university, road 100, guangzhou City martyr, guangdong province, accession number: GDMCC No:62894.
The strain YZ-151 screened by the invention has good antagonism effect on acanthopanax black spot pathogenic bacteria (Alternaria tenuissima) and acanthopanax gray mold pathogenic bacteria (Botrytis cinerea B. Cinerea), and the bacteriostasis rate is more than 78%; the bacteria-free filtrate of the strain YZ-151 has the bacteriostasis rate of 48.06% and 57.62% on the pathogenic bacteria of acanthopanax black spot (Alternaria tenuissima) and the pathogenic bacteria of acanthopanax gray mold (Botrytis cinerea B.cinerea); the strain YZ-151 has different degrees of inhibition on common fungal pathogens, and shows broad-spectrum antibacterial properties. Meanwhile, the strain YZ-151 screened by the invention has good acanthopanax leaf surface colonization capability and good control effect on acanthopanax black spot and gray mold.
The invention provides a theoretical basis for green prevention and treatment of acanthopanax black spot and gray mold and a scientific basis for development and application of acanthopanax biocontrol bacteria resources.
Drawings
FIG. 1 shows the antagonistic effect of the strain YZ-151 on the pathogenic bacteria of acanthopanax black spot and gray mold. In the figure, A is acanthopanax black spot pathogen (Alternaria minutissima A. Tenuissima), and B is acanthopanax gray mold pathogen (Botrytis cinerea B. Cinerea).
FIG. 2 shows colony morphology of strain YZ-151.
FIG. 3 is a phylogenetic tree constructed based on the 16S rDNA partial sequence of strain YZ-151.
FIG. 4 shows the antagonistic effect of strain YZ-151 on 5 common fungal pathogens. In the figure, 1: strain YZ-151, fusarium solani, 2: strain YZ-151 against sclerotinia, 3: bacterial strain YZ-151, alternaria alternata, 4: the strain YZ-151 destroyed the cyclosporin in the opposite direction, 5: fusarium oxysporum strain YZ-151.
FIG. 5 shows the control effect of strain YZ-151 on acanthopanax black spot. In the figure, a: strain YZ-151; CK: and (5) treating with clear water.
FIG. 6 shows the control effect of the strain YZ-151 on gray mold of acanthopanax. In the figure, a: strain YZ-151; CK: and (5) treating with clear water.
Detailed Description
The technical solutions of the embodiments of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention, and it is apparent that the described embodiments are only some embodiments of the present invention, not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Test materials
Root soil of acanthopanax root: is obtained from a medicinal plant garden planting base (43 DEG 48 '23' N,125 DEG 24 '57' W) of Jilin agricultural university.
Test pathogenic bacteria: alternaria tenuissima, botrytis cinerea B.cinerea, fusarium solani Fusarium solani, sclerotinia sclerotiorum Sclerotinia ginseng, alternaria tenuissima Alternariapanax, cyclosporum catastrophe Cylindrocarpon destructans (Zinss.) Scholtan, and Fusarium oxysporum Fusarium oxysporum are all provided by the Jilin agricultural university plant disease comprehensive management laboratory.
The formulation of each medium is shown in the table.
Bacterial extraction kit TaKaRa MiniBEST Universal Genomic DNAExtraction Kit ver.5.0: purchased from vinca, sciences.
Example 1 screening, identification and preservation of Strain YZ-151
1. Screening
1) Collecting a plurality of healthy acanthopanax root rhizosphere soil samples, and sieving the soil samples with a 20-mesh sieve after air-drying the soil samples. 10g of the sample is weighed and put into a triangular flask filled with glass beads and 90mL of sterile water, fully oscillated for 10-30min, uniformly mixed and stood for 5min. The solution was diluted to 10 -4 in a sterile condition and applied to NA medium in 200. Mu.L of each dilution. Each treatment was repeated 3 times, the plates were placed in a constant temperature incubator and subjected to purification culture at 30℃for 24-48 hours, a plurality of pure colonies were isolated, and were numbered and stored at 4℃for further use.
2) Under the aseptic condition, each single bacterial colony is picked up and respectively streaked and inoculated on a flat plate containing an NA culture medium, after the flat plate is inversely cultured for 24 hours at the temperature of 30 ℃ in a constant temperature incubator, 3 bacterial cakes with the diameter of about 1cm are picked up and inoculated into a triangular flask (the liquid loading amount is 50mL/100 mL) containing an NB culture medium, and shake culture is carried out for 24 hours at the temperature of 32 ℃ and 180r/min, so that the NB bacterial culture solution is obtained.
3) The filter paper sheet method is adopted for primary screening: preparing activated radix Acanthopanacis Senticosi black spot pathogenic bacteria (Alternaria tenuissima) and radix Acanthopanacis Senticosi gray mold pathogenic bacteria (Botrytis cinerea B.cinerea) into bacterial cakes with diameter of 8mm respectively, inoculating to the center of a plate (plate diameter of 90 mm) containing PDA culture medium under aseptic condition, soaking sterilized filter paper sheet in NB bacterial culture solution for 1min, drying in shade, sticking the filter paper sheet to 4 symmetrical corner points at position about 25mm from the center of the plate, inoculating pathogenic bacteria alone as control, repeating each treatment for 3 times, and culturing in a dark box at 25deg.C. And (5) after the control group grows up on the flat plate, measuring the growth condition of pathogenic bacteria, and observing whether a bacteria inhibition zone is generated.
4) The oxford cup method is adopted for re-screening: the NB bacterial culture was centrifuged at 12000r/min for 20min at 4℃to collect the supernatant, which was subjected to degerming with a 0.22 μm filter, and the sterile filtrate was collected. Inoculating bacterial cake with diameter of 8mm at center of plate containing PDA culture medium, placing 4 sterile oxford cups at 4 symmetrical corners at position about 25mm from center of the plate, adding 200 μl sterile filtrate into each sterile oxford cup, inoculating pathogenic bacteria alone as control, repeating each treatment for 3 times, culturing in a dark box in a constant temperature incubator at 25deg.C, and calculating antibacterial rate after the control group is full of the plate. The calculation formula of the bacteriostasis rate is as follows: antibacterial ratio (%) = (radius of control colony-radius of counter cultured colony)/(radius of control colony-4) ×100%.
5) Results
By analysis, the strain with the highest bacteriostasis rate is named YZ-151, wherein the bacteriostasis rate of the strain YZ-151 on acanthopanax black spot pathogenic bacteria (Alternaria tenuissima) and acanthopanax gray mold pathogenic bacteria (Botrytis cinerea) is more than 78%, and the results are shown in table 1 and figure 1; the average antibacterial rate of the sterile filtrate of the strain YZ-151 on the acanthopanax black spot pathogenic bacteria (Alternaria tenuissima) and the acanthopanax gray mold pathogenic bacteria (Botrytis cinerea) is 52.84%, and the results are shown in Table 2.
Table 1 inhibition of Acanthopanax senticosus and Gray mold by YZ-151 strain
| Pathogenic bacteria | Alternaria tenuissima (A. Tenuissima) | Botrytis cinerea (B.cinerea) |
| Antibacterial rate% | 78.86±1.41 | 79.57±5.61 |
Table 2 inhibition of the sterile filtrate of Strain YZ-151 against Acanthopanax senticosus and Botrytis cinerea
| Pathogenic bacteria | Alternaria tenuissima (A. Tenuissima) | Botrytis cinerea (B.cinerea) |
| Antibacterial rate% | 48.06％±1.00 | 57.62％±5.61 |
2. Authentication
1) Morphological characteristics and physiological and biochemical characteristics identification: reference materials of microbiological test handbook, common bacterial System identification handbook and Berger's bacterial identification handbook, strains YZ-151 are streaked and inoculated in NA culture medium, placed in a constant temperature incubator at 30 ℃ for 2-3 days, single colony morphological characteristics are observed, gram staining and spore staining observation are carried out, and physiological and biochemical indexes such as carbohydrate metabolism test, enzyme test, citrate utilization test and salt tolerance are measured by adopting a bacterial trace biochemical reaction tube.
2) 16S rDNA identification: single colony of strain YZ-151 is inoculated in NB culture solution, cultured at 32 ℃ and 180r/min for 24 hours, and bacterial DNA is extracted by adopting a bacterial extraction kit after centrifugally collecting thalli. 16S rDNA selection Universal primers 27F (5-AGAGTTTGATCCTGGGCTCAG-3) and 1492R (5-TACGGCTACCTTGTTACGACTT-3), PCR reaction System: genomic DNA 2. Mu. L, dNTP (10. 10 mmoL/L) 2. Mu.L, 10 XBuffer 5. Mu.L, universal primer 27F 2. Mu.L, universal primer 142r2mu. L, taq enzyme (2.5U/. Mu.L) 2. Mu. L, ddH 2 O35. Mu.L. 16S rDNA gene PCR amplification conditions: 94 ℃ for 5min;94 ℃ for 1min;58 ℃ for 30s; 90s at 70 ℃ for 35 cycles; and at 72℃for 10min. The amplified products were detected by 1% agarose gel electrophoresis and then sent to the Shanghai Co., ltd for sequencing.
3) Results
3-1) Morphological Properties: on NA medium, the colony of strain YZ-151 became milky, the surface of the colony was wet, opaque (FIG. 2), and the gram stain was in the form of a rod with spores.
3-2) Physiological and biochemical characteristics: the strain YZ-151 can utilize glucose, sucrose and mannitol, xylose and L-arabinose cannot be utilized, the V-P test is negative, the OF test is positive, the hydrogen sulfide test result is negative, the gelatin hydrolysis and the starch hydrolysis are both positive, the arginine double hydrolase is negative, the citrate, the tartrate is negative, the casein hydrolysis, the contact enzyme, the oxidase and the nitrate reduction are both positive, and the highest salt tolerance concentration is 7%.
3-3) Molecular identification: the 16S rDNA gene PCR amplified product of the strain YZ-151 is detected by 1% agarose gel electrophoresis and then is sent to the Wuhan division company of the Shanghai Co., ltd. For sequencing, and the effective sequence length is 1434bp (SEQ ID NO: 1), and the GenBank accession number is OK474770. A phylogenetic tree is constructed at NCBI by BLAST sequence alignment and by adopting MEGAX64 software through an N-J method, and as a result, FIG. 3 shows that the 16S rDNA effective sequence of the strain YZ-151 is clustered with the Acerist (Priestia aryabhattai) and has the closest relationship and higher confidence. The strain YZ-151 was identified as Lecanier ardsteria by combination of morphological, physiological and biochemical features, and 16S rDNA identification comprehensive analysis (Priestia aryabhattai).
3. Preserving
The screened strain of the Agrimonia azedarach (Priestia aryabhattai) YZ-151 is preserved in the microorganism strain collection of Guangdong province, namely GDMCC in 2023 and 20 days, and the address is: building 5, 30 # of university, road 100, guangzhou City martyr, guangdong province, accession number: GDMCC No:62894.
EXAMPLE 2 antibacterial Spectrometry test of Strain YZ-151
The bacterial strain YZ-151 is subjected to antibacterial spectrum measurement by using 5 common fungal pathogens (Fusarium solani, sclerotinia sclerotiorum Sclerotinia ginseng, alternaria alternifolia ALTERNARIA PANAX, synechococcus destroyed Cylindrocarpon destructans (Zinss.) Scholtan and Fusarium oxysporum Fusarium oxysporum) as targets, each treatment is repeated for 3 times, and the bacterial strain YZ-151 is placed in a incubator at 25 ℃ for culturing until a control group grows up to a flat plate, and the antibacterial rate is calculated. The calculation formula of the bacteriostasis rate is as follows: antibacterial ratio (%) = (radius of control colony-radius of counter cultured colony)/(radius of control colony-4) ×100%.
As shown in Table 3 and FIG. 4, the strain YZ-151 has obvious antagonism to Fusarium solani, sclerotinia sclerotiorum Sclerotinia ginseng, alternaria Alternariapanax, leuconostoc mesenteroides Cylindrocarpon destructans (Zinss.) Scholtan and Fusarium oxysporum Fusarium oxysporum, the bacterial inhibition rate of the strain YZ-151 to Alternaria Alternariapanax reaches 81.10%, the bacterial inhibition rates to Fusarium solani and Leuconostoc mesenteroides Cylindrocarpon destructans (Zinss.) Scholtan are 77.13% and 76.22%, respectively, and the bacterial inhibition rates to Sclerotinia sclerotiorum ALTERNARIA PANAX and Fusarium oxysporum Fusarium oxysporum are 67.07% and 68.90%, respectively.
TABLE 3 inhibition of 5 common fungal pathogens by strain YZ-151
| Pathogenic bacteria | Fusarium solani | Sclerotinia sclerotiorum (L.) kuntze | Alternaria alternata (L.) Kuntze | Destruction of post-sporulation bacteria | Fusarium oxysporum |
| Antibacterial rate% | 77.13±0.43b | 67.07±1.83c | 81.10±0.86a | 76.22±0.61b | 68.90±1.72c |
Example 3 screening of Strain YZ-151 for Rifampicin resistance and test for colonization on Acanthopanax leaf surface
Culturing the strain YZ-151 in a rifampicin NB culture solution with the concentration of 10 mug/mL, 20 mug/mL, 30 mug/mL and 40 mug/mL … mug/mL in sequence, and inducing the resistance mutation of rifampicin step by step to obtain a mutant strain YZ Rif -151; the original strain YZ-151 is used as a control, and whether the antibacterial activity of the mutant strain YZ Rif -151 on acanthopanax black spot pathogenic bacteria (Alternaria tenuissima) and acanthopanax gray mold pathogenic bacteria (Botrytis cinerea) is different or not is observed. Inoculating the mutant strain YZ Rif -151 into a triangular flask (with a liquid loading amount of 500mL/1000 mL) containing a BPY culture medium fermentation broth, culturing at 34 ℃ and 180r/min for 48h, and preparing a bacterial suspension with a bacterial content of 10 7 by using the sterile BPY culture medium fermentation broth; uniformly spraying 10mL of bacterial suspension on the leaf surfaces of acanthopanax, sampling and recovering the acanthopanax in 1d, 3d, 7d, 11d, 17d, 25d and 35d respectively, setting 3 basins for each treatment, randomly taking acanthopanax leaf tissues from each basin, sterilizing the surfaces of the acanthopanax leaf tissues by using 75% ethanol solution for 15s, washing the acanthopanax leaf tissues with sterile water for several times, weighing a plurality of acanthopanax leaf tissues, grinding the acanthopanax leaf tissues into homogenate in a sterile mortar, dissolving the homogenate in 10mL of sterile water, standing for 30min, diluting the homogenate to 10 -3 for standby respectively, measuring the bacterial count by adopting a flat plate gradient dilution method, coating 200 mu L of diluent on NA culture medium containing 200 mu g/mL of rifampicin, repeating each treatment for 3 times, and culturing at 30 ℃ for 3d, and counting the bacterial colony count on the surfaces of the culture medium. The number of colonies per gram of fresh leaf tissue was counted (cfu. G -1). The calculation formula of the bacteria content is as follows: bacterial load = colony count x dilution fold x water for isolation/plate water for plating x tissue mass for isolation (g).
After the mutant strain YZ Rif -151 is continuously cultured for 3 generations on NA culture medium without rifampicin, the mutant strain YZ Rif -151 can still grow normally on rifampicin NB culture solution with the concentration of 200 mug/mL, and the fermentation liquor of the mutant strain YZ-151 still has antagonism on acanthopanax black spot pathogenic bacteria (Alternaria graciliata) and acanthopanax gray mold pathogenic bacteria (Botrytis cinerea), and the antagonism effect is not obviously different from that of the original strain YZ-151, so that the strain YZ-151 has genetic stability and can still keep good antibacterial activity on acanthopanax black spot pathogenic bacteria (Alternaria graciliata) and acanthopanax gray mold pathogenic bacteria (Botrytis cinerea).
The acanthopanax leaf surface is sprayed with bacterial liquid of anti-rifampicin marker strain YZ-151, which is in a trend of 'increasing after decreasing' within 0-35d, and the leaf surface colonization amount of mutant strain YZ Rif -151 is different in different periods, the leaf surface colonization amount of mutant strain YZ Rif -151 is shown in table 4, the colonization amount of mutant strain YZ Rif -151 reaches the lowest value at day 7, the peak value at day 11, and the colonization amount reaches 1.24×10 6 CFU/g, which is 4.89 times of day 7. The mutant strain YZ Rif -151 can still keep the amount of the colonitis to be more than 10 3 CFU/g fresh leaves within 0-35d, which shows that the strain YZ-151 has good acanthopanax leaf surface colonisation capability and can be used for potted plant disease prevention tests.
TABLE 4 leaf surface colonization amount of mutant strain YZ Rif -151
Example 4 potted biocontrol test of Strain YZ-151
1) Preparation of pathogenic bacteria suspension: inoculating preserved radix Acanthopanacis Senticosi black spot pathogenic bacteria (Alternaria tenuissima) and radix Acanthopanacis Senticosi gray mold pathogenic bacteria (Botrytis cinerea B.cinerea) into PDA culture medium respectively, inversely culturing at 25deg.C until the whole plate grows, collecting bacterial cake, inoculating into PDB culture medium to obtain mycelium suspension, filtering, drying water, weighing mycelium, diluting with sterile water, finally preparing Alternaria tenuissima (A.tenuissima) into mycelium suspension with mycelium concentration of 8.76g/L, preparing Botrytis cinerea (B.cinerea) into mycelium suspension with mycelium concentration of 4.68g/L, and storing mycelium suspension at 4deg.C for use.
2) Preparation of antagonistic bacteria suspension: inoculating the strain YZ-151 into NA culture medium, culturing at 30 ℃ for 48 hours, picking 2-3 colonies with the diameter of about 1cm, inoculating into NB culture medium (50 mL/100mL of liquid loading), culturing at 32 ℃ for 12 hours at 180r/min, inoculating into BPY culture medium fermentation broth with the inoculum size of 1%, culturing at 34 ℃ for 24 hours at 180r/min to obtain strain YZ-151 bacterial suspension, and regulating the concentration of the strain YZ-151 bacterial suspension to 10 7 cfu/mL by using the BPY culture medium fermentation broth.
3) Potted plant test of acanthopanax black spot: selecting 3-year-old acanthopanax for potting test, needling 5-6 wounds on acanthopanax leaves by a needling method, smearing 10mL of mycelium suspension (mycelium concentration is 8.76 g/L) of Alternaria tenuissima (A. Tenuissima) on each plant of needled leaves, and setting 2 treatments, wherein each treatment comprises 5 pots and 1 plant per pot. A: strain YZ-151 (strain YZ-151 bacterial suspension concentration 10 7 cfu/ml); CK: and (5) treating with clear water.
Acanthopanax black spot disease grading standard: level 0: complete leaf, no disease spot: stage 1: a small amount of lesions (1% -5% of leaf area); 3 stages: moderate plaque formation (6% -10% of leaf area); 5 stages: the number of the lesions is large (accounting for 11% -20% of the area of the leaf blade); 7 stages: the disease spots are many and large (accounting for 21% -50% of the leaf area); stage 9: the lesions are numerous and large (51% -100% of the leaf area).
Disease index = { Σ [ (number of patients at each stage×number of patients at each stage)/(total number of patients examined×9) ] } ×100.
4) Potted plant test of acanthopanax gray mold
Selecting 3-year-old acanthopanax for potting test, needling 5-6 wounds on acanthopanax leaves by a needling method, smearing 10mL of mycelium suspension of Botrytis cinerea (B.cinerea) on each plant of needled leaves, wherein the mycelium concentration is 4.68g/L, setting 2 treatments, and setting 5 pots for each treatment, wherein each pot is 1 plant. A: strain YZ-151 (strain YZ-151 bacterial suspension concentration 10 7 cfu/ml); CK: and (5) treating with clear water.
Gray mold grading standard of acanthopanax: level 0: complete leaf, no disease spot: stage 1: a small amount of lesions (1% -5% of leaf area); 3 stages: moderate plaque formation (6% -10% of leaf area); 5 stages: the number of the lesions is large (accounting for 11% -20% of the area of the leaf blade); 7 stages: the disease spots are many and large (accounting for 21% -50% of the leaf area); stage 9: the lesions are numerous and large (51% -100% of the leaf area).
Disease index = { Σ [ (number of patients at each stage×number of patients at each stage)/(total number of patients examined×9) ] } ×100.
Control effect = [ (control disease index-treatment disease index)/control disease index ] ×100%.
5) The acanthopanax black spot disease potting test and the gray mold potting test are both carried out for 35 days, and the condition of the acanthopanax is investigated after 35 days of inoculation. As shown in Table 5, FIG. 5 and FIG. 6, the strain YZ-151 has a certain control effect on acanthopanax black spot and gray mold, and compared with CK clear water control, the strain YZ-151 can effectively reduce the disease index of acanthopanax black spot and gray mold. The strain YZ-151 has good control effect on acanthopanax black spot and gray mold.
Table 5 prevention and treatment effects of strain YZ-151 on acanthopanax black spot and gray mold
According to the invention, a strain of Agrimonia azedarach (Priestia aryabhattai) YZ-151 is screened out from healthy acanthopanax root soil, and an antibacterial test proves that the strain YZ-151 has antagonistic effect on acanthopanax black spot pathogenic bacteria (Alternaria tenuissima) and acanthopanax gray mold pathogenic bacteria (Botrytis cinerea) and the antibacterial rate is over 78%; the bacteria inhibition rate of the sterile filtrate of the strain YZ-151 on the pathogenic bacteria of acanthopanax black spot (Alternaria tenuissima) and the pathogenic bacteria of acanthopanax gray mold (Botrytis cinerea B.cinerea) is 48.06% and 57.62% respectively; the strain YZ-151 has different degrees of inhibition effects on 5 pathogenic fungi such as Fusarium solani, sclerotinia sclerotiorum Sclerotinia ginseng, alternaria alternifolia Alternariapanax, synechococcus Cylindrocarpon destructans (Zinss.) Scholtan, fusarium oxysporum Fusarium oxysporum and the like, and has broad-spectrum antibacterial properties.
Screening a strain of Agrimonia arvensis (Priestia aryabhattai) YZ-151 from healthy acanthopanax root soil, and carrying out an acanthopanax leaf surface colonization test to prove that the strain YZ-151 has good acanthopanax leaf surface colonization capability; potted plant experiments prove that the strain YZ-151 has good control effect on acanthopanax black spot and gray mold.
The invention discloses an Aleplerenone (Priestia aryabhattai) YZ-151 strain and application thereof, and the skilled person can properly improve the technological parameters by referring to the content of the present disclosure. It is expressly noted that all such similar substitutions and modifications will be apparent to those skilled in the art, and are deemed to be included in the present invention. While the invention has been described with reference to preferred embodiments, it will be apparent to those skilled in the relevant art that variations and modifications can be made in the invention described herein without departing from the spirit or scope of the invention.
Claims (7)
1. An aprepitant ardsteria (Priestia aryabhattai) YZ-151, characterized in that the strain was deposited at the cantonese province microorganism strain collection at 2023, month 02 and day 20 under the deposit number: GDMCC No: 62894.
2. Use of a strain of listeria albopictus (Priestia aryabhattai) YZ-151 as defined in claim 1 for the preparation of a plant pathogenic antagonist; the method is characterized in that the plant pathogenic bacteria are as follows: alternaria tenuissima, botrytis cinerea, fusarium solani, sclerotinia panaxatriformis, alternaria panaciens, leptosporum vulgare and Fusarium oxysporum.
3. The use according to claim 2, wherein the plant pathogenic bacteria antagonist is an acanthopanax pathogenic bacteria antagonist.
4. Use according to claim 3, characterized in that the active ingredient of the plant pathogenic antagonist is a bacterial suspension of the species listeria albopictus (Priestia aryabhattai) YZ-151.
5. Use according to claim 3, characterized in that the active ingredient of the plant pathogenic antagonist is a sterile filtrate of the bacterium ambrisentan (Priestia aryabhattai) YZ-151, the plant pathogenic bacteria being: alternaria tenuissima and Botrytis cinerea.
6. The use of a strain of listeria albopictus (Priestia aryabhattai) YZ-151 according to claim 1 for the control of acanthopanax black spot.
7. Use of a strain of listeria albopictus (Priestia aryabhattai) YZ-151 according to claim 1 for controlling acanthopanax gray mold.
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