CN111979149B

CN111979149B - Salt-tolerant bacillus SY1836 and application thereof

Info

Publication number: CN111979149B
Application number: CN202010825298.5A
Authority: CN
Inventors: 徐生军; 荆卓琼; 郭致杰; 张新瑞
Original assignee: INSTITUTE OF PLANT PROTECTION GANSU ACADEMY OF AGRICULTURAL SCIENCES
Current assignee: Chongqing Jiweiyuan Biotechnology Co ltd
Priority date: 2020-08-17
Filing date: 2020-08-17
Publication date: 2022-04-26
Anticipated expiration: 2040-08-17
Also published as: CN111979149A

Abstract

The invention relates to salt-tolerant bacillus SY1836 and application thereof. The preservation number of the salt-tolerant Bacillus (Bacillus halotolerans) SY1836 is CGMCC No.20080, and the Bacillus halotolerans is a biocontrol bacterium separated from saline-alkali soil. Experiments prove that the salt-tolerant Bacillus (Bacillus halotolerans) SY1836 has a good effect of antagonizing various plant pathogenic bacteria, can effectively reduce the disease index of the pepper when applied to the growth process of the pepper, and has a remarkable pepper phytophthora blight prevention and treatment effect. The salt-tolerant Bacillus (Bacillus halotolerans) SY1836 can also obviously improve the plant height and fresh weight in the growth process of the hot pepper, and has important significance in the prevention and control of the hot pepper phytophthora blight and the planting.

Description

Salt-tolerant bacillus SY1836 and application thereof

Technical Field

The invention relates to the technical field of microorganisms, in particular to salt-tolerant bacillus SY1836 and application thereof.

Background

The capsicum belongs to a perennial or limited perennial herbaceous solanaceae plant, is an important vegetable and seasoning, is edible in seed oil, and has a certain medicinal value. With the continuous increase of the cultivation area of the pepper, the production is usually accompanied with high-degree intensive planting, the multiple cropping index is high, a host and a breeding place depending on survival are provided for the occurrence of soil-borne diseases, a large amount of pathogenic bacteria in the soil are accumulated, the occurrence of the soil-borne diseases is more and more serious, and the development of pepper production is seriously restricted by epidemic diseases caused by phytophthora capsici.

At present, the control is mainly carried out by breeding disease-resistant varieties and applying chemical pesticides, but the real effective disease-resistant varieties are lacked in production, or the disease resistance of the disease-resistant varieties is easy to lose, and in order to obtain high yield and income, the application amount and application times of the chemical pesticides are increased by a producer, so that the dependence on the pesticides is generally formed in production. The application of a large amount of chemical pesticides plays an important role in improving the production of the peppers, but simultaneously brings a plurality of problems, such as a series of food safety and ecological environment problems of drug resistance generation of pathogenic bacteria, reduction of pesticide utilization rate, reduction of pepper quality, increase of pesticide residue, soil salinization, underground water pollution and the like. Meanwhile, with the continuous improvement of living standard, people have higher and higher requirements on green consumption and pollution-free vegetables, so that the production of the pollution-free vegetables is imperative, the development of the production of the pollution-free vegetables is required, besides increasing various management measures, the disease control is well done, and the reduction of the use amount of chemical pesticides is vital. The biological control can overcome a series of defects brought by chemical control, can indirectly generate certain economic benefit, ecological benefit and social benefit, and accords with the development trend of building a sustainable and eco-friendly green agricultural system. In general, biological control based on the development of plant and microorganism interactions between microorganisms would be a good complement or even an alternative to current methods of plant disease control.

Relevant researches show that the biological control fungicide developed by using the microorganisms has good application effects in the aspects of preventing and controlling plant diseases, promoting plant growth, improving plant stress resistance and the like. The bacillus subtilis TG26 strain separated from the loofah rhizosphere soil has strong inhibiting effect on wheat scab and watermelon fusarium wilt, the pot control effect on the wheat scab reaches 89.8%, and the seedling control effect on the watermelon fusarium wilt reaches 100%. The field test shows that the paenibacillus polymyxa WY110 separated from the rice rhizosphere has high in-vitro inhibitory activity on 3 main pathogenic bacteria of rice, namely rice blast, rhizoctonia solani and bacterial blight, and has the nitrogen fixation function. The spore strain Bn-130 separated and screened from the pepper rhizosphere soil shows that the control effect on pepper phytophthora blight reaches 64.17 percent through a greenhouse disease control test. The microbial pesticide 'Baikang' (bacillus subtilis wettable powder) jointly developed by Yunnan agricultural university and Chinese agricultural university mainly prevents and treats rice sheath blight, root rot of panax notoginseng and tobacco black cavity disease. "Maifengning" developed by Nanjing agriculture university is a living biological bactericide prepared from bacillus subtilis strain B3, has a field prevention effect on wheat sharp eyespot of 50% -80%, and has a disease prevention mechanism mainly expressed in the production of antibacterial substances for inhibiting the growth of hypha, the formation of sclerotium and the germination of sclerotium of wheat sharp eyespot.

However, a biocontrol agent that promotes the growth of one crop does not necessarily have a similar effect on other crops. Some bacteria produce a general growth promoting effect on several crops, while others exhibit strong host-crop selectivity and colonize individual or limited crops. In addition, the interaction between the biocontrol microorganism and the microbial community in the soil ecological environment often exists, and the introduction of the exogenous biocontrol microorganism can cause the composition of the microbial community of the rhizosphere soil of crops to change, so that the use of the biocontrol microbial inoculum has great regional adaptability.

Disclosure of Invention

In order to at least solve the problems in the prior art, the invention provides salt-tolerant bacillus SY1836 and application thereof.

In a first aspect, the invention provides a Bacillus halodurans SY1836 with preservation information as follows: the preservation number is: CGMCC No. 20080; the classification is named as: bacillus halotolerans (Bacillus halotolerans); the preservation unit is as follows: china general microbiological culture Collection center (CGMCC); the preservation address is as follows: the institute of microbiology, national academy of sciences No. 3, Xilu No. 1, Beijing, Chaoyang, Beijing; the preservation date is as follows: year 2020, 6, 15.

The invention collects soil samples from water source town salinization plots in Jinchang city, Gansu province, separates a strain with inhibition capability to various pathogenic bacteria, identifies the strain as salt-tolerant Bacillus (Bacillus halotolrans) after identifying the physicochemical property and comparing 16S rRNA, gyrA, cheA and amyA gene sequences, and is named as salt-tolerant Bacillus (Bacillus halotolrans) SY1836, and the physiological characteristics are as follows:

the spores are rod-shaped, have endospores, are gram-positive, grow anaerobically, grow on glucose, sucrose, fructose, mannitol, glycerol and cellulose, and grow on 15% NaCl.

The invention further provides a microbial inoculum containing the Bacillus halodurans SY 1836.

In a second aspect, the present invention provides a method of promoting the growth of a pepper plant, comprising:

applying the Bacillus halodurans SY1836 to the pepper plant, or applying the inoculant.

The microbial inoculum can be a fermentation product of Bacillus halodurans SY 1836.

Further, the total number of viable bacteria of the Bacillus halodurans SY1836 is not less than 10⁸CFU/mL。

The invention further provides the function of the Bacillus halodurans SY1836 or the microbial inoculum in promoting the growth of the pepper.

The invention further provides the effect of said Bacillus halodurans SY1836 or said bacterial agent in inhibiting pathogenic bacteria which are one or more of Botrytis cinerea (Botrytis cinerea), colletotrichum oxysporum (colletotrichum acutum), Fusarium graminearum (Fusarium graminearum), Fusarium oxysporum f.sp.globosum, Fusarium solani (f.solani), Fusarium verticillioides (f.verticillioides), Phytophthora capsici (Phytophthora capsici), Rhizoctonia solani (Rhizoctonia solani) and Sclerotinia sclerotiorum (sclerotiorum) in.

The invention further provides application of the Bacillus halodurans SY1836 or the microbial inoculum in improving the plant height and/or fresh weight of the capsicum.

The invention has the following beneficial effects:

according to the invention, the salt-tolerant Bacillus SY1836 is obtained by separating from soil, has an inhibiting effect on various plant pathogenic bacteria, can effectively reduce disease index of pepper when the salt-tolerant Bacillus SY1836 is applied to the pepper growth process, and has a good biological control effect on pepper phytophthora blight. In addition, the salt-tolerant Bacillus (Bacillus halotolerans) SY1836 can also effectively promote the growth of the pepper and improve the plant height and fresh weight in the growth process of the pepper. The salt-tolerant Bacillus (Bacillus halotolerans) SY1836 provided by the invention has important significance in controlling and planting pepper phytophthora blight.

Drawings

FIG. 1 is a detection result of substances with characteristics related to disease prevention and growth promotion generated by Bacillus halodurans SY1836 provided in example 2 of the present invention; wherein A is phosphate dissolution; b is a siderophore; c is protease; d is alpha-amylase; e is cellulase; f is pectinase; g is NH 3; h is IAA;

FIG. 2 is a result of antagonistic ability test of various plant pathogenic bacteria of Bacillus halodurans SY1836 provided in example 2 of the present invention; wherein: a is Botrytis cinerea (Botrytis cinerea); b is Colletotrichum anthracis (Colletoichum acutatum); c is Fusarium graminearum (Fusarium graminearum); d is a specialized form (F.oxysporum f.sp.conglutinans) of the family of Fusarium oxysporum Cruciferae; e is fusarium solani (f.solani); f is Fusarium verticillium (F.verticillioides); g is Phytophthora capsici (Phytophthora capsicii); h is Rhizoctonia solani (Rhizoctonia solani); i is Sclerotinia sclerotiorum (sclerotirotiorum);

FIG. 3 shows the effect of Bacillus halotolerans SY1836 on controlling phytophthora blight of capsicum annuum in greenhouse seedlings and potted plants in the embodiment 3 of the invention; wherein, the sequence of the diagram from left to right and from top to bottom is A-H; wherein A-D is the seedling tray control effect, and E-H is the potting control effect; wherein A and E are Bacillus halodurans SY1836, B and F are Bacillus subtilis, C and G are fluazinam, and D and H are clear water contrast;

FIG. 4 shows the effect of Bacillus halotolerans SY1836 on controlling phytophthora blight in field according to embodiment 4 of the present invention; wherein A is salt-tolerant Bacillus (Bacillus halotolerans) SY1836, B is Bacillus subtilis, C is fluazinam, and D is clear water contrast.

Detailed Description

The following examples are intended to illustrate the invention but are not intended to limit the scope of the invention.

Example 1

The embodiment provides a process for separating and purifying Bacillus halodurans (SY 1836) from soil, which specifically comprises the following steps:

the soil sample is collected from a water source saline land in Jinchang City of Gansu province. Randomly selecting 5 points in a sampling plot, taking 0-5cm of salinized soil, mixing, bringing back by using a sealing bag, removing soil blocks and plant residues by using a sieve with a mesh of 0.2cm, and air-drying at room temperature. Weighing 10g of soil sample, placing the soil sample into a triangular flask containing 90mL of sterilized water, and shaking for 10min to prepare 10g of soil sample^-1Standing the soil suspension with the concentration until the soil particles precipitate, sucking 1mL of supernatant, transferring into a test tube containing 9mL of sterilized water, and making into 10^-2The soil suspension with the concentration is sequentially subjected to gradient dilution to obtain the final concentration of 10^-5The soil suspension of (1). Selection 10^-3，10^-4And 10^-5Three concentrations of the suspension were pipetted 200. mu.L each, added to LB medium plates with 10% NaCl content, spread evenly with a spreading bar and repeated 3 times. Culturing the culture dish at a constant temperature of 28 ℃ for 5 days, selecting colonies with different forms, transferring the colonies to a new LB culture medium plate with 10 percent of NaCl, carrying out purification culture, and storing at 4 ℃ to be tested. The strain is frozen in a freezing storage tube (the content of glycerol is 20 percent) at the temperature of minus 80 ℃ in an ultralow temperature refrigerator for long-term storage.

In the embodiment, the colony morphology and color of bacteria are directly observed and distinguished, 66 bacterial isolates are obtained through co-separation, and antagonistic capacity is screened and detected by a plate confronting culture method, so that 5 bacterial strains can antagonize 9 pathogenic fungi, wherein the strain SY1836 shows strong antagonistic capacity. This example further identifies the strain SY1836 obtained by separation and purification, and its physiological characteristics are as follows: the spores are rod-shaped, have endospores, are gram-positive, grow anaerobically, grow on glucose, sucrose, fructose, mannitol, glycerol and cellulose, and grow on 15% NaCl.

In this example, the gene sequence analysis and identification is further performed by the following specific method:

this example uses Fast TIANAmp Bacteria DNAkit (Tiangen, Beijing) to extract the whole genome DNA of biocontrol bacterium SY 1836. The 16S rRNA gene of the strain was PCR-amplified with universal primers 27F (5'-AGAGTTTGATCCTGGCTCAG-3') and 1492R (5'-GGTTACCTTGTTACGACTT-3') for the bacterial 16S rRNA gene. The gyrA gene of the strain was PCR amplified with primers GyrA-F (5'-CAGTCAGGAAATGCGTACGTCCTT-3') and GyrA-R (5'-CAAGGTAATGCTCCAGGCATTGCT-3'). The cheA gene of the strain was PCR amplified with primers CheA-F (5'-AGGGCTGCCCATACATTAAAAGGAATGAGC-3') and CheA-R (5'-AGTTTCAACCGGGACCATTCTCATATTCAG-3'). The amyA gene of the strain was PCR amplified with primers ABa3(5'-GCACGCTGATGCAGTATTTTGAATGG-3') and ABa4 (5'-GGAGCTGTCACCTTCCCTCGTCC-3'). 25 μ L of the reaction mixture was amplified on a PCR thermal cycler. The PCR product was purified and sent to Biotechnology (Shanghai) Inc. (Shanghai, China) for sequencing. The sequencing results were compared in the NCBI database, the 16S rRNA, gyrA, cheA and amyA gene sequences of related species were downloaded from the NCBI database, and phylogenetic studies were performed using MEGA 6.0. The 16S rRNA, gyrA, cheA and amyA gene sequences of strain SY1836 were submitted to the NCBI database.

Finally, the conclusion is drawn: the lengths of the 16S rRNA, gyrA, cheA and amyA gene sequences of Bacillus halodurans SY1836 are 1458bp, 990bp, 826bp and 1184bp respectively, homology comparison analysis is carried out on the obtained sequences and the 16S rRNA, gyrA, cheA and amyA gene sequences of some strains in NCBI databases, and the results show that the similarity of the 16S rRNA, gyrA, cheA and amyA gene sequences of the strain SY1836 and Bacillus halodurans (Bacillus halodurans) is 99.79%, 99.69%, 99.64% and 99.74% respectively. Based on the results of homology comparison analysis, the strain SY1836 was identified as Bacillus halolerans.

In this embodiment, Bacillus halotolerans SY1836 is further preserved in the common microorganism center of china committee for culture collection of microorganisms (CGMCC), under the classification name: bacillus halotolerans (Bacillus halotolerans); and (4) storage address: the institute of microbiology, national academy of sciences No. 3, Xilu No. 1, Beijing, Chaoyang, Beijing; the preservation number is CGMCC No. 20080; the preservation date is as follows: year 2020, 6, 15.

Example 2

In this embodiment, the specific method for detecting disease prevention and growth promotion related substances of Bacillus halodurans SY1836 is as follows:

1.1 production of siderophiles

And (3) inoculating the strain to a CAS culture medium, culturing in an incubator at 30 ℃ for 14d, and observing whether a yellow orange color circle appears, wherein if the yellow orange color circle appears, the strain has the ability of producing siderophin.

1.2 production of Amylase

And (3) inoculating the strain to an NA culture medium containing 1% soluble starch, culturing in an incubator at 30 ℃ for 5d, and observing whether a transparent ring appears, wherein if the transparent ring appears, the strain has the amylase producing capability.

1.3 production of proteases

The strain is picked up and spotted on a casein yeast culture medium (5 g of casein, 2.5g of yeast extract, 1g of glucose and 1000mL of water) containing 7% skimmed milk powder, the strain is placed in an incubator at 30 ℃ for culture for 5d, and whether a transparent ring appears is observed, if so, the strain has the capability of producing protease.

1.4 production of pectinase

The strain is picked and inoculated on a basic culture medium (1 g of sodium nitrate, 1g of dipotassium phosphate, 1g of potassium chloride, 0.5g of magnesium sulfate, 0.5g of yeast powder, 1g of glucose, 18g of agar and 1000mL of water) containing 1% pectin, the strain is placed in a 30 ℃ incubator for culture for 7d, Gram iodine solution (1.0 g of iodine, 2.0g of potassium iodide and 300mL of water) is poured into the pectin culture medium, whether a transparent ring appears or not is observed under a dark blue background, and if yes, the strain has the capability of producing pectinase.

1.5 production of cellulase

The strain is picked and spotted on a basic culture medium containing 1% of cellulose, the strain is placed in an incubator at 30 ℃ for 5d, 0.01% Congo red solution submerges the cellulose culture medium for 15min, the culture medium is washed by water for 3 times, whether a transparent ring appears or not is observed, and if yes, the strain has the capability of producing the cellulase.

1.6 production of IAA

The strain was subjected to shake cultivation at 30 ℃ for 72 hours in TSB medium containing 0.1% L-tryptophan, the bacterial suspension was centrifuged to remove the cells, 2.0mL of Salkawski color developer (50mL, 35% perchloric acid, 1mL of 0.5M ferric chloride) was added to 1.0mL of the sterile filtrate, and the mixture was allowed to develop at room temperature for 30min, indicating the generation of IAA if pink color was generated, and the medium was used as a control.

1.7 dissolution of phosphorus

The strain is picked up and spotted on PVK culture medium (0.5 g of yeast powder, 10.0g of glucose, 5.0g of calcium phosphate, 0.5g of ammonium sulfate, 0.2g of potassium chloride, 0.1g of magnesium chloride, 0.0001g of manganese sulfate, 0.0001g of ferrous sulfate, 18.0g of agar and 1000mL of water), the strain is placed in a 30 ℃ incubator for culture for 14d, whether a transparent ring appears or not is observed, and if yes, the strain has phosphorus dissolving capacity.

1.8 Potassium dissolution

The strain is selected and spotted on an Aleksandrov culture medium (5.0 g of glucose, 0.5g of magnesium sulfate heptahydrate, 0.005g of ferric chloride, 2.0g of calcium carbonate, 2.5g of yeast powder, 18.0g of agar and 1000mL of water) containing 0.2% potassium feldspar, the strain is placed in an incubator at 30 ℃ for culture for 14d, whether a transparent circle appears or not is observed, and if yes, the strain has the potassium-dissolving capacity.

1.9 Nitrogen fixation

The strain is selected and inoculated on an Azadirachta culture medium (10.0 g of glucose, 0.2g of dipotassium phosphate, 0.5g of calcium carbonate, 0.2g of magnesium sulfate heptahydrate, 0.2g of sodium chloride, 0.1g of calcium sulfate, 18.0g of agar and 1000mL of water), the strain is placed in an incubator at 30 ℃ for culture for 14d, whether the strain normally grows or not is observed, and if the strain normally grows, the strain has the nitrogen fixing capacity.

1.10NH₃Generation of

Inoculating 50. mu.L of the bacterial suspension into 30mL of peptone ammoniation medium (5.0 g of peptone, 0.5g of dipotassium phosphate, 0.25g of sodium chloride, 0.5g of magnesium sulfate heptahydrate, 0.01g of ferrous sulfate heptahydrate, 1000mL of water), culturing in a 30 ℃ incubator for 3d, adding 1.0mL of Nessler color developing agent (6.0 g of mercuric chloride, 12.4g of potassium iodide, 30mL of 20% sodium hydroxide, 70mL of water), and if the yellow color becomes brown, indicating that there is NH₃Production, medium as control.

The detection result is shown in figure 1, Bacillus halodurans SY1836 can produce siderophin, alpha-amylase, protease, pectinase, cellulase, IAA and NH3, and has nitrogen fixation and phosphorus dissolution capability and no potassium dissolution capability.

In this example, the antagonistic activity of Bacillus halodurans SY1836 against various plant pathogens was further examined, after cutting Botrytis cinerea (Botrytis cinerea), colletotrichum oxysporum (colletotrichum acutum), Fusarium graminearum (Fusarium graminearum), Fusarium oxysporum (f.sp.globosum), Fusarium solani (f.solani), Fusarium verticillioides (f.verticillioides), Phytophthora capsici (Phytophthora capsici), Rhizoctonia solani (Rhizoctonia solani) and Sclerotinia sclerotiorum (sclerotiorum) at a constant temperature of 25 ℃ for 7 days with a punch (6.5mm), inoculating a cake 2cm from the culture medium, inoculating the cake at the other side of the culture medium, and culturing the cake in a parallel zone at a distance of 2cm from the culture medium, and measuring the colony size of 10 ℃ in a single spot at the same time. Each treatment was repeated 3 times. The results are shown in table 1 and fig. 2:

TABLE 1 antagonistic ability of strain SY1836 against various phytopathogens

Example 3

In this embodiment, the greenhouse disease prevention and growth promotion effects of Bacillus halodurans SY1836 are further detected, and the specific method is as follows:

2.1 preparation of salt tolerant Bacillus (Bacillus halotolerans) SY1836 biocontrol microbial inoculum

Bacillus halodurans SY1836 was inoculated on LB solid medium for activation, transferred to a 100mL Erlenmeyer flask containing 50mL LB liquid medium, and shake-cultured at 28 ℃ and 150rpm for 24h to obtain a seed solution. The seed solution was inoculated into a fermentation medium (1/2PDB +1/2LB) at a ratio of 1:100, and cultured at 28 ℃ for 24 hours with shaking at 150 rpm.

2.2 determination of seedling tray control effect of Bacillus halotolerans SY1836 on pepper phytophthora blight

Seeds of a pepper variety named Changfeng are surface-sterilized in 0.1% sodium hypochlorite for 20min and then washed with sterile water four times. The seeds were sown in 50-well seedling trays filled with horticultural soil, one seed per well. The seedling grows in a greenhouse with no disease and insect at 20-30 deg.C, after 8 weeks, before inoculating pathogenic bacteria 3d and 4h, 20mL strain SY1836 fermentation broth (viable bacteria concentration 10)⁸CFU/mL) were poured into the horticultural soil of the plant roots, respectively, to prevent excess liquid from flowing out from the bottom of the seedling tray. Clear water, Bacillus subtilis (a.i.1X 10)¹¹Spores/gram, WP, debang bio ltd, harbin, china) and fluazinam (a.i.50%, SC, jiangyin suli chemical ltd, jiangyin, china) were treated as blank control and positive control, respectively, using the same treatment method described above. Each treatment was repeated 3 times for a total of 30 strains. Pipette 10mL of P.capsicii inoculum (1X 10)⁵sporangia/mL) was poured into horticultural soil at the root of each plant. The inoculated plants were placed in a growth chamber to maintain humidity above 90% and temperature control at 20-30 ℃. And (4) investigating disease progression at 7d after pathogen inoculation, and calculating prevention and control effects. Grading the disease condition standard: level 0: no symptoms of health; level 1: the seedling has slightly black root and stem, and leaves do not wither or can recover; and 2, stage: the stem of the seedling becomes black to 1-2cm, the leaves are irreversibly wilted, and the lower leaves occasionally fall off; and 3, level: the stem of the seedling becomes black by more than 2cm, and the leaves are obviously wilted or shed; 4, level: the root and stem of the seedling become black and contract, and all the fallen leaves or plants except the growing point are wilted; and 5, stage: the whole plant died.

The method for calculating the disease index and the prevention and control effect comprises the following steps:

in the formula: CK is the disease index of blank control treatment, and PT is the disease index of medicament (biocontrol bacteria) treatment.

2.3 determination of potted plant control effect of Bacillus halotolerans SY1836 on pepper phytophthora blight

Pepper seeds were treated as described above. Seeds were sown in 105-well trays filled with horticultural soil, one seed per well. The seedlings were grown in a greenhouse at a temperature of 20-30 ℃ without disease and after 4 weeks, the seedlings were transplanted into plastic pots (10 cm in diameter) filled with horticultural soil and continued to grow in a greenhouse at a temperature of 20-30 ℃ without disease and for 4 weeks. 50mL of strain SY1836 fermentation liquor (viable bacteria concentration 10) 3d and 4h before inoculation of pathogenic bacteria⁸CFU/mL) were separately poured into the horticultural soil at the roots of the plants to prevent excess liquid from flowing out of the bottom of the plastic pots. And (3) treating the plants by using the same treatment method by using clear water, the bacillus subtilis and the fluazinam as a blank control and a positive control respectively. Each treatment was repeated 3 times for a total of 30 strains. Pipette 10mL of P.capsicii inoculum (1X 10)⁵sporangia/mL) was poured into horticultural soil at the root of each plant. The inoculated plants were placed in a growth chamber to maintain humidity above 90% and temperature control at 20-30 ℃. And (4) investigating disease progression at 14d after pathogen inoculation, and calculating prevention and control effects. The disease grading criteria are as above.

The control effect on pepper phytophthora blight of the embodiment is shown in fig. 3, wherein a-H are sequentially shown in the sequence diagram from left to right and from top to bottom; wherein A-D is the seedling tray prevention and control effect, and E-H is the potted plant prevention and control effect; wherein A and E are Bacillus halodurans SY1836, B and F are Bacillus subtilis, C and G are fluazinam, and D and H are clear water contrast; as can be seen from fig. 3, the effect of Bacillus halodurans SY1836 used in this example is similar to that of fluazinam in controlling phytophthora blight of pepper, and is significantly better than that of Bacillus subtilis. Through calculation, the prevention and control effects of Bacillus halodurans SY1836 on pepper phytophthora blight are 58.45% (seedling tray test) and 69.33% (pot culture test) respectively.

2.4 Bacillus halodurans SY1836 Pot culture test for promoting growth of Capsicum plants

Biocontrol bacterial SY1836 bacterial suspension was prepared and pepper seeds were treated as described above. Seeds were sown in 105-well trays filled with horticultural soil, one seed per well. Seedlings were grown in a greenhouse at a temperature of 20-30 ℃ without pests, and after 4 weeks, the seedlings were transplanted into plastic pots (10 cm in diameter) filled with horticultural soil. 50mL of strain SY1836 fermentation liquor (viable bacteria concentration 10)⁸CFU/mL) was separately poured into the wet horticultural soil at the roots of the plants to prevent excess liquid from flowing out of the bottom of the plastic pots, for a total of 3 applications every 7 d. Plants were treated with clear water as a blank control using the same treatment method described above. Each treatment was repeated 3 times for a total of 30 strains. Plant height, root length, fresh weight of aerial parts and fresh weight of roots of pepper plants were investigated at 14d after the last treatment.

The growth promoting effect of the Bacillus halodurans SY1836 is as follows:

TABLE 2 growth promoting effect of biocontrol bacterial strain SY1836 greenhouse potted plant

Example 4

In the embodiment, the field disease prevention and growth promotion effects of Bacillus halodurans SY1836 are further detected, and the specific method comprises the following steps:

in the embodiment, the prevention and control effect of the biocontrol bacterial strain SY1836 on pepper epidemic diseases is measured by a field test of artificial inoculation in a Lanzhou test station and a Yuzhong test station of agricultural academy of sciences of Gansu province in the growing season of the pepper. The strain SY1836 biocontrol microbial inoculum and pepper seedlings were prepared in the same manner as shown in example 3. At days 5 and 25, pepper seedlings (9 weeks old) were transplanted to 300m²And 500m²Lanzhou and Ulmus. The row spacing and the plant spacing were kept at 50cm and 30cm, respectively. The biocontrol bacterial strain SY1836, bacillus subtilis (a.i.1X 10) are set for the treatment¹¹Spore/g, WP, Deqiang biology Ltd, Harbin, China), fluazinam (a.i.50%, SC, Jiangyin Soli Chemie Ltd, China Jiangyin) and water (blank control), inThe rows were randomized block trial, with 3 replicates per treatment. 100mL of strain SY1836 fermentation broth (viable bacteria concentration 10) at 8 months, 1 day, 8 days and 15 days⁸CFU/mL), Bacillus subtilis and fluazinam (at the recommended dosages by the supplier) and clear water were poured separately into the plant roots. At day 8, 15, 10mL of P.capsicii inoculum (1X 10)⁵sporangia/mL) was poured into the soil surrounding the roots of each plant. And (4) investigating disease progression at 15d after pathogen inoculation, and calculating prevention and control effects. Grading the disease condition standard: level 0: no symptoms of health; level 1: the aerial parts only have scabs on leaves and fruits; and 2, stage: brown rot on the overground stems and branches; and 3, level: brown rot spots are formed at the base of the stem; 4, level: the overground stems, branches and stem bases have brown rot spots, and part of branches die; and 5, stage: the whole plant died.

The control effect of the Bacillus halodurans SY1836 used in the embodiment on the field phytophthora blight of the pepper is similar to that of fluazinam in the control effect of the phytophthora blight of the pepper and is significantly better than that of the Bacillus subtilis, and the control effects of the Bacillus halodurans SY1836 on the phytophthora blight of the pepper are 62.81 +/-1.21% (Lanzhou test station) and 65.08 +/-2.67% (Yuzhou test station) respectively by calculation as can be seen from fig. 4.

Although the invention has been described in detail hereinabove with respect to a general description and specific embodiments thereof, it will be apparent to those skilled in the art that modifications or improvements may be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.

Claims

1. A salt-tolerant Bacillus (Bacillus halotolerans) SY1836 is characterized in that the preservation number of the salt-tolerant Bacillus (Bacillus halotolerans) SY1836 is as follows: CGMCC No. 20080.

2. An agent, characterized in that the agent contains Bacillus halodurans (Bacillus halotolerans) SY1836 of claim 1.

3. A method of promoting the growth of a pepper plant, comprising:

applying to said pepper plant Bacillus halodurans SY1836 according to claim 1 or an inoculant according to claim 2.

4. The method according to claim 3, wherein the total viable count of Bacillus halodurans SY1836 is not less than 10⁸CFU/mL。

5. The method according to claim 3 or 4, wherein said applying is to the roots of said pepper plants.

6. Use of Bacillus halodurans SY1836 as claimed in claim 1 or a bacterial agent as claimed in claim 2 for promoting the growth of pepper.

7. The use of Bacillus halodurans SY1836 or the inoculant of claim 2 for inhibiting the action of a pathogen selected from the group consisting of Botrytis cinerea (Botrytis cinerea), colletotrichum oxysporum (Colletorichum acutum), Fusarium graminearum (Fusarium graminearum), Fusarium oxysporum f.sp.conduranans, Fusarium solani (f.solani), Fusarium verticillioides (f.verticillioides), Phytophthora capsici (Phytophthora capsici), Rhizoctonia solani (Rhizoctonia solani) and Sclerotinia sclerotiorum (sclerotiorum).

8. Use of Bacillus halodurans SY1836 as claimed in claim 1 or a bacterial agent as claimed in claim 2 for increasing the height of pepper plants.

9. Use of Bacillus halodurans SY1836 as claimed in claim 1 or a bacterial agent as claimed in claim 2 for increasing fresh weight of pepper.