Bacillus D79, compound fungicide and application of bacillus D79 in promoting growth of fruit trees

Abstract

The invention relates to the technical field of microorganisms, and particularly discloses bacillus Bacillus sp.D79, a compound fungicide and application of the bacillus Bacillus sp.D79 in promoting growth of fruit trees, the bacillus Bacillus sp.D79 is preserved in China General Microbiological Culture Collection Center (CGMCC) on September 25, 2024, the preservation number is CGMCC No.32070, and the bacillus Bacillus sp.D79 is classified and named as bacillus Bacillus sp. The bacillus D79 provided by the invention is used for promoting the growth of cherry seedlings.

Description

Technical Field

[0001] The invention relates to the technical field, and in particular to a Bacillus D79, a compound bacterial agent and application thereof in promoting the growth of fruit trees. Background Art

[0002] Fruit trees refer to trees with edible fruits, and are a general term for perennial plants and their rootstocks that can provide edible fruits and seeds, such as apple trees, pear trees, citrus trees, apricot trees, peach trees, and cherry trees.

[0003] Sweet cherry (Prunus avium) is a popular staple fruit crop rich in sugar, organic acids, minerals and other nutrients. Due to its high economic and nutritional value, the planting area of ​​sweet cherry in China is gradually increasing. However, due to the poor site conditions of sweet cherry orchards and shallow soil layers. In addition, cherry trees have weak root development, a small root distribution range, weak absorption capacity, and are highly sensitive to changes in the soil environment. These unfavorable factors severely restrict the growth and development of cherry trees. Therefore, there is an urgent need to find economically feasible, efficient, and environmentally friendly ways to improve the root function of cherries, promote plant growth and development, and ensure the sustainable development of the sweet cherry industry.

[0004] At present, plant growth regulators are mostly used for plant growth and development, and most of them are synthetic chemicals. The concentration and dosage of plant growth regulators are extremely high, and irregular use may cause toxicity to plants. Microorganisms, as important participants in the ecosystem, are safe and friendly to the environment. Therefore, it is necessary to screen and isolate microorganisms that have a growth-promoting effect on fruit trees. Summary of the invention

[0005] In order to develop a biological product that has a growth-promoting effect on plants, the present invention provides a strain of Bacillus D79, a composite bacterial agent and the use thereof in promoting the growth of fruit trees. The Bacillus D79 provided by the present invention is used to promote the growth of cherry seedlings.

[0006] The present invention provides a Bacillus sp. D79, which is deposited in the General Microbiology Center of China National Microbiological Culture Collection Committee on September 25, 2024, with a deposit number of CGMCC No. 32070 and a classification name of Bacillus sp.

[0007] The Bacillus sp. D79 provided by the invention can improve the biomass of cherry plants, increase the total root length, root surface area, root volume and average root diameter of the cherry root system, and promote the growth of cherry seedlings.

[0008] The present invention also provides a Bacillus D79 bacterial suspension, wherein the Bacillus D79 is cultured to a bacterial concentration of 1×10 7 CFU / mL~6ⅹ10 7 CFU / mL was obtained.

[0009] The present invention also provides a composite bacterial agent obtained by compounding a Bacillus D79 bacterial suspension and an Enterobacter D27 bacterial suspension, wherein the Enterobacter D27 bacterial suspension is prepared from Enterobacter D27;

[0010] The Enterobacter D27 is deposited in the General Microbiological Center of China Microorganism Culture Collection Administration, with the deposit number: CGMCC No.29568.

[0011] Furthermore, the concentration of the Enterobacter D27 bacterial suspension is 1×10 7 CFU / mL~6ⅹ10 7 CFU / mL.

[0012] The present invention also provides a method for preparing the composite bacterial agent, wherein a suspension of Bacillus D79 and a suspension of Enterobacter D27 are prepared respectively so that the bacterial concentrations are both 1×10 7 CFU / mL~6ⅹ10 7 CFU / mL, then the Bacillus D79 bacterial suspension and the Enterobacter D27 bacterial suspension were mixed at a volume ratio of 1-2:1;

[0013] The preparation steps of the Bacillus D79 bacterial suspension are as follows: activating the Bacillus D79, inoculating it in a liquid culture medium and culturing it to a logarithmic growth phase, centrifuging it, discarding the supernatant, and resuspending the bacterial cell with sterile water to obtain the Bacillus D79 bacterial suspension.

[0014] Furthermore, the centrifugation condition is 4°C, 8000-12000 r / min, 10-15 min.

[0015] Furthermore, the concentration of the Bacillus D79 bacterial solution is 10 7 CFU / mL.

[0016] The present invention also provides an application of the Bacillus sp. D79, the Bacillus D79 bacterial suspension or the compound bacterial agent in promoting the growth of fruit trees.

[0017] Furthermore, the Bacillus sp. D79 or the compound bacterial agent is used to increase the biomass of cherry plants.

[0018] Furthermore, the Bacillus sp. D79 or the compound bacterial agent is used to increase the total root length, root surface area, root volume and average root diameter of the cherry root system.

[0019] Compared with the prior art, the present invention has the following beneficial effects:

[0020] The Bacillus D79 isolated and obtained by the present invention acts on cherries, increases the total root length, root surface area, root volume and average root diameter of the cherry root system, and improves the chlorophyll content, thereby promoting the growth and development of the cherry plants. The composite bacterial agent obtained by compounding Bacillus D79 and Enterobacter D27 can also increase the total root length, root surface area, root volume and average root diameter of the cherry root system, and promote the growth and development of the cherry plants. The composite bacterial agent has a better growth-promoting effect on cherry plants than using Bacillus D79 alone.

[0021] Information on the deposit of biological materials

[0022] D79, referred to as Bacillus sp. D79 in this application, was deposited in the General Microbiology Center of China Microorganism Culture Collection Administration on September 25, 2024, with the deposit number: CGMCC No. 32070. The address of the depository is the Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Postal Code: 100101, and the classification name is Bacillus sp. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0024] Figure 1 The strain characteristics of Bacillus D79 of the present invention;

[0025] In the figure, a is the color reaction of Bacillus D79;

[0026] b shows the IAA production of Bacillus D79 after different culture times;

[0027] c is the Gram staining result of Bacillus D79;

[0028] d is the growth curve of Bacillus D79.

[0029] Figure 2 This is the growth status of cherry seedlings 60 days after treatment with Bacillus D79.

[0030] Figure 3 This is the growth and development of the root system of cherry plants after being treated with Bacillus D79 and the compound bacterial agent for 60 days in the present invention.

[0031] Figure 4 This is the effect of Bacillus D79 and the compound bacterial agent on the maximum photochemical efficiency (Fv / Fm) of cherry plants in the present invention.

[0032] Figure 5 The effect of Bacillus D79 and the compound bacterial agent on the α diversity index of the rhizosphere microbial community of cherry after 60 days of treatment;

[0033] In the figure, a is the richness among bacterial community groups;

[0034] b is the diversity among bacterial community groups;

[0035] c is the diversity among fungal community groups;

[0036] d is the richness of fungal community groups.

[0037] Figure 6 The effect of Bacillus D79 and the compound bacterial agent on the β diversity of the rhizosphere microbial community of cherry after 60 days of treatment;

[0038] In the figure, a is the PCoA analysis of bacterial communities based on the Bray-Curtis algorithm;

[0039] b: PCoA analysis of fungal communities based on the unweighted-unifrac algorithm.

[0040] Figure 7 The relative abundance changes of cherry rhizosphere microorganisms at the phylum and genus levels after 60 days of treatment with Bacillus D79 and the compound bacterial agent in the present invention;

[0041] In the figure, a is the relative abundance of bacterial communities at the phylum level;

[0042] b is the relative abundance of fungal communities at the phylum level;

[0043] c is the heat map of the relative abundance of the top 50 bacterial genera at the genus level;

[0044] d is the heat map of the relative abundance of the top 50 genera in the fungal community at the genus level. DETAILED DESCRIPTION

[0045] The specific embodiments of the present invention are described in detail below, but it should be understood that the protection scope of the present invention is not limited by the specific embodiments. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in this field without creative work belong to the scope of protection of the present invention. The experimental methods described in the embodiments of the present invention are conventional methods unless otherwise specified, and the materials, reagents, etc. used in the following embodiments can be obtained from commercial channels unless otherwise specified.

[0046] Example 1: Isolation and screening of Bacillus sp. D79, determination of growth-promoting properties and identification of the genus.

[0047] 1. Experimental Methods

[0048] 1. Sampling

[0049] The rhizosphere soil samples used in the present invention were collected from the robust Northeastern Mountain Cherry (Cerasus sachalinensis Kom.) in the Northeastern Mountain Cherry Ex Situ Conservation Nursery of Shenyang Agricultural University, Shenyang, Liaoning Province, China (41°49′31″N, 123°34′0″E, which has been in a natural growth state since its construction in 2006). In June 2022, three trees were selected, and a fan-shaped pit with a radius of 50 cm, a depth of 30 cm, and an angle of 45° was dug around them to observe the root distribution of the Northeastern Mountain Cherry. Capillary roots with a depth of less than 15 cm were selected, and the rhizosphere soil was obtained by the root shaking method. Weigh 10 g of soil sample and dissolve it in 90 mL of sterile water to make a mother solution, which was then diluted to 10 -2 ~10 -8 The rhizospheric bacteria were isolated and cultured on beef extract peptone (NB) solid medium.

[0050] 2. Strain screening and characteristic test

[0051] (1) Qualitative test for IAA-producing strains: The isolated strains were cultured in NB liquid culture medium (added with 100 mg / L L-tryptophan) for 24 h, and then 50 μl of the bacterial solution was subjected to a color reaction with 50 μl of Salkowski reagent (50 mL 35% HClO4 + 1 mL 0.5 mol / LFeCl3). The darker the red color, the stronger the ability of the strain to produce IAA. The IAA-producing strain D79 was screened and obtained.

[0052] (2) Quantitative determination of IAA production: After culturing the selected strain D79 for 12 h, 24 h, 36 h, and 48 h, 5 mL of the bacterial suspension was centrifuged at 10,000 r / min for 10 min, 3 mL of the supernatant was taken, an equal volume of Salkowski colorimetric reagent was added, and the supernatant was kept in the dark for 30 min to measure the OD 530The IAA yield was calculated by the IAA standard curve; the sterilized NB medium was used as the blank control, the IAA standard sample concentration was used as the horizontal axis, and the OD 530 The values ​​were used as the ordinate to draw the IAA standard curve (Table 1).

[0053] Table 1 IAA standard curve

[0054]

[0055] (3) Determination of plant growth-promoting properties

[0056] The screened IAA-producing strain D79 was subjected to Gram staining and growth curve determination; the nitrogen fixation ability of the strain was detected using Ashby medium, and the nitrogenase activity of the strain was determined using a nitrogenase ELISA research kit; the ability of the strain to dissolve inorganic and organic phosphorus was determined using PKO medium (containing Ca3(PO4)2 and egg yolk lecithin, respectively).

[0057] 3. Molecular identification of strain D79

[0058] The strain was cultured in NB liquid medium until the logarithmic growth phase, and the bacteria were collected by centrifugation. The 16S rDNA sequence was amplified by PCR using the bacterial 16SrDNA universal primers 27F and 1492R; the sequence was sequenced by Shanghai Sangon Biotechnology Co., Ltd., and then the sequence was compared with the bacterial sequence in the NCBI database to identify the species of strain D79.

[0059] 2. Experimental Results

[0060] The results are as follows Figure 1 In a, the color of strain D79 is red, which shows that strain D79 has a stronger ability to produce IAA compared with CK. Figure 1 According to b, the highest IAA production capacity of Bacillus D79 is 12.44 mg / L; Figure 1 From the c, we can see that Bacillus D79 is a Gram-positive bacterium; from the growth curve analysis of Bacillus D79, it can be concluded that Bacillus D79 reaches the logarithmic growth phase when it is cultured for 40 hours ( Figure 1 d).

[0061] As shown in Table 2, Bacillus D79 has the ability to fix nitrogen and dissolve phosphorus. The nitrogenase activity of D79 strain is 28.25 IU / L; the soluble phosphorus content in the organic phosphorus fermentation broth inoculated with Bacillus D79 is about 1.42 mg / L; the soluble phosphorus content in the inorganic phosphorus fermentation broth is about 47.49 mg / L.

[0062] Table 2 Quantitative results of nitrogen fixation and phosphorus solubilization of strain D79

[0063] strain Nitrogenase activity IU / L Dissolving organic phosphorus capacity mg / L Dissolving inorganic phosphorus capacity mg / L CK － 0.45±0.09b 36.83±1.64b D79 28.25±2.27a 1.42±0.07a 47.49±2.12a

[0064] Note: Data are expressed as mean ± standard deviation (n = 3). Different lowercase letters in the same column represent significant differences between different treatments, p < 0.05.

[0065] 2. Molecular identification of strain D79

[0066] According to 16S rDNA sequencing, strain D79 belongs to the genus Bacillus.

[0067] Example 2: Growth-promoting effect of Bacillus sp. D79 on cherry seedlings.

[0068] 1. Test methods

[0069] 1. Preparation of inoculated bacterial solution: Inoculate the activated Bacillus D79 into 50 mL NB liquid medium and culture it to the logarithmic growth phase, centrifuge at 4°C 10000r / min for 10 min, discard the supernatant, and resuspend the cells with sterile water to a concentration of about 10 7 CFU / mL(OD 600 =0.8), to obtain Bacillus D79 bacterial suspension for later use.

[0070] The activated Enterobacter D27 was inoculated into 50 mL NB liquid medium and cultured to the logarithmic growth phase. The culture was centrifuged at 10000 r / min at 4°C for 10 min, the supernatant was discarded, and the cells were resuspended with sterile water to a concentration of about 10 7 CFU / mL(OD 600 =0.8), and obtain D27 bacterial suspension for standby use. The Enterobacter D27 is deposited in the General Microbiological Center of China Microbiological Culture Collection Administration, with the deposit number: CGMCC No.29568.

[0071] 2. Planting and treatment of experimental seedlings: The planting soil was a mixture of experimental garden soil and river sand from the fruit tree research base of Shenyang Agricultural University in a ratio of 2:1 (the contents of organic matter, alkaline nitrogen, available phosphorus and available potassium were 17.42 g / kg, 127.33 mg / kg, 173.42 mg / kg and 123.75 mg / kg, respectively); the experimental seedlings were 'Gisela No. 6' cherry seedlings, which were produced by plant tissue culture technology in advance and transferred to nutrient pots for cultivation after hardening; the formal experimental treatment began on June 25, 2023. The seedlings were about 10 cm tall. There were three treatments in the experiment: (1) control (CK), plants were inoculated with 50 mL of sterile water; (2) D79 strain, inoculated with 50 mL of Bacillus D79 bacterial suspension; (3) mixed bacterial agent (MIX), inoculated with 25 mL of D27 bacterial suspension and 25 mL of D79 bacterial suspension. Each treatment was repeated 5 times. Sterile water, D79 strain and MIX were inoculated by root irrigation, once every 5 days, for a total of 6 times; the inoculation was stopped 30 days after inoculation, and then entered a 30-day normal management period, for a total of 60 days; the experiment began sampling on August 25, 2023 to determine the plant biomass and root growth indicators.

[0072] 3. Determination of plant biomass and root system indicators

[0073] The height and stem diameter of the seedlings were measured with a tape measure and a vernier caliper. The fresh weight and dry weight of the aboveground and underground parts of the plants were weighed after the roots were cleaned (dried at 80°C for 72 h); the surface area, volume, and total length of the roots were determined using a digital scanner (Epson, Long Beach, USA).

[0074] To determine the chlorophyll content in leaves, fresh leaves were chopped and extracted in 5 ml of 80% acetone in the dark for 24 h, and then the absorbance of the mixture at 663 nm and 645 nm was recorded spectrophotometrically. Plant leaves were treated in the dark for 30 min in advance, and leaf chlorophyll fluorescence parameters were measured using a multifunctional plant efficiency meter.

[0075] 4. Assessment of the rhizosphere microbial community

[0076] At the same time, the root shaking method was used to collect the rhizosphere soil of the plants in each group after 60 days of treatment, and the large soil particles and the remaining roots were removed through a 20-mesh sieve. Finally, a total of 9 rhizosphere soil samples (3 treatments × 3 replicates) were obtained and stored at -80 °C for future use. 5 g of each sample was weighed and sent to Shanghai Meiji Biomedical Technology Co., Ltd. for sequencing, and the Meiji interactive analysis cloud platform was used to analyze the microbial diversity of rhizosphere soil. Uparse software was used to cluster tags with an identification rate of more than 97% into OTUs, and then an OTU table was generated. Mothur software was used for Alpha diversity analysis, including sobs index, shannon index, and simpson index. The principal coordinates analysis of bacterial and fungal communities was performed using R language, using the bray-curtis algorithm and the unweighted-unifrac algorithm. The bar graphs and heat maps of bacterial and fungal community composition were drawn using R language.

[0077] 5. Statistical analysis

[0078] The data were analyzed using SPSS26.0 software, the graphics were drawn using Origin2021 software, the calculated data were presented in the form of mean ± standard deviation, and one-way analysis of variance (ANOVA) was used to evaluate the differences in the data obtained from the experiment, and the significance level was set at P < 0.05.

[0079] 2. Experimental Results

[0080] 1. Cherry plant biomass

[0081] The growth status of Gisela 6 seedlings after 60 days of treatment is as follows Figure 2 As shown in Table 3, Bacillus D79 treatment had a growth-promoting effect on the biomass of cherry plants. Compared with CK, plant height, stem diameter, aboveground and underground biomass increased significantly after Bacillus D79 treatment (P<0.05). The mixed bacterial agent (MIX) treatment had a more significant growth-promoting effect on cherry plants than Bacillus D79 alone.

[0082] Table 3 Effects of Bacillus D79 on the biomass of cherry (Gisela 6) seedlings

[0083]

[0084] Note: Data are expressed as mean ± standard deviation (n = 5). Different lowercase letters in the same column represent significant differences between different treatments, p < 0.05.

[0085] 2. Root structure of cherry plants

[0086] The results are as follows Figure 3As shown in Table 4, compared with CK, the root volume and average root tip diameter of the D79 strain inoculant treatment increased significantly by 35.92% and 9.62%, respectively.

[0087] Table 4 Effects of Bacillus D79 on root architecture of cherry (Gisela 6) seedlings

[0088] deal with Total root length(m) <![CDATA[Root surface area (cm 2 )]]> <![CDATA[Root volume (cm 3 )]]> Average root diameter (mm) CK 13.46±2.60a 221.10±46.10a 2.84±0.59b 0.52±0.01b D79 15.22±0.47a 271.48±12.70a 3.86±0.36a 0.57±0.03a MIX 14.73±1.39a 274.86±31.39a 4.00±0.64a 0.59±0.03a

[0089] Note: Data are expressed as mean ± standard deviation (n = 3). Different lowercase letters in the same column represent significant differences between different treatments, p < 0.05.

[0090] 3. Determination of maximum photochemical efficiency (Fv / Fm) and chlorophyll content

[0091] The results are as follows Figure 4 As shown, compared with CK, the maximum photochemical efficiency (Fv / Fm) of D79 strain and mixed bacterial agent (MIX) treatment was significantly higher than that of CK treatment.

[0092] As shown in Table 5, compared with CK, the D79 strain inoculation treatment significantly increased the leaf chlorophyll a, chlorophyll b and total chlorophyll (a+b) contents, while the MIX strain inoculation treatment had no significant effect on the chlorophyll content.

[0093] Table 5 Effect of Bacillus D79 on chlorophyll content in cherry (Gisela 6) seedlings

[0094] deal with Chlorophyll a(mg / g) Chlorophyll b(mg / g) Total chlorophyll a+b (mg / g) CK 1.84±0.10b 0.72±0.04b 2.56±0.13b D79 2.95±0.12a 1.29±0.04a 4.24±0.15a MIX 2.00±0.09b 0.78±0.04b 2.77±0.14b

[0095] Note: Data are expressed as mean ± standard deviation (n = 4). Different lowercase letters in the same column represent significant differences between different treatments, P < 0.05.

[0096] 4. Effects of inoculation with Bacillus D79 on rhizosphere microbial communities

[0097] Table 6 Effect of strain D79 and MIX on the relative abundance of bacterial genera with PGP traits (%)

[0098] Serial number Genus name CK D79 MIX 1 Arthrobacter 5.62 6.88 4.11 2 Sphingomonas 3.61 2.41 2.91 3 Pseudolabrys 1.69 1.46 1.59 4 Streptomyces 1.65 1.40 1.28 5 Bacillus 1.11 1.33 1.80 6 Bradyrhizobium 1.32 1.26 1.40 7 Novosphingobium 1.39 1.24 0.89 8 Nocardioides 1.18 1.22 1.10 9 Mesorhizobium 0.82 1.13 1.16 10 Mycobacterium 0.72 0.82 0.70 11 Aeromicrobium 0.76 0.85 0.59 12 Pseudarthrobacter 0.60 0.96 0.57 13 Solirubrobacter 0.56 0.67 0.76 14 Kribbella 0.57 0.69 0.62 15 Devosia 0.76 0.53 0.44 16 Phycicoccus 0.58 0.69 0.47 17 Microlunatus 0.51 0.63 0.51 18 Blastococcus 0.52 0.51 0.52 total 23.97 24.68 21.42

[0099] Note: The bacterial genera with PGP traits were obtained from the PlaBase database by comparing the top 50% relative abundance of bacteria at the genus level in this experimental data.

[0100] The results are as follows Figure 5 As shown in Table 6, the Shannon index of D79 and MIX compound bacterial agent treatments was significantly higher than that of CK, and the Simpson index was significantly lower than that of CK.

[0101] like Figure 6As shown in the figure, the PCoA-based β diversity analysis of rhizosphere soil microorganisms showed that the rhizosphere soil microorganisms treated with D79 and MIX mixed bacterial agents had obvious clustering. The results showed that there was a clear separation between the rhizosphere soil bacterial community and fungal community in the strain treatment and the control treatment. Inoculation with different strains had a significant effect on the β diversity of rhizosphere soil bacterial and fungal communities in Gisela 6.

[0102] 5. Rhizosphere microbial community structure

[0103] Different strain treatments had different effects on the species composition of rhizosphere microorganisms of cherry plants. At the bacterial phylum level, the top 10 dominant bacterial phyla in abundance were Proteobacteria, Actinobacteriota, Acidobacteriota, Chloroflexi, Gemmatimonadota, unclassified Bacteria, Firmicutes, Bacteroidota, Patescibacteria, and Myxococcota, accounting for more than 95% of the entire bacterial community. Compared with CK, the abundance of unclassified Bacteria decreased significantly in each inoculation group, while the abundance of Bacteroidota, Patescibacteria, and Myxococcota increased to varying degrees in each inoculation group. In addition, the abundance of Firmicutes and Verrucomicrobiota was most significant in the MIX compound bacterial agent.

[0104] At the bacterial genus level, the top 10 dominant bacterial genera in abundance were Arthrobacter, Gaiellales, Vicinamibacterales, Sphingomonas, Gemmatimonadaceae, Gaiella, Vicinamibacteraceae, RB41, Pseudolabrys, and Streptomyces. There were some differences in the changes of each genera between different inoculation groups. Compared with CK, the abundance of Arthrobacter increased significantly in the D79 inoculation group, but decreased significantly in the MIX inoculation group; the abundance of unclassified Bacteria and Sphingomonas decreased to varying degrees. However, the abundance of Bacillus increased to varying degrees in each inoculation group, with the most obvious change in MIX.

[0105] At the same time, the present invention also conducted the same analysis on the fungal community. At the phylum level, Ascomycota (68.44% to 74.71%), Mortierellomycota (10.17% to 13.40%) and Basidiomycota (11.66% to 13.09%) were the dominant phyla in the four inoculation groups, accounting for more than 90% of the entire fungal community. Among them, the abundance of Ascomycota showed a significant downward trend compared with the CK group, while the abundance of Mortierellomycota showed a significant upward trend compared with the CK group. At the genus level, Cyberlindnera (15.95%-32.52%), Mortierella (10.06%-13.10%), Pseudeurotium (3.29%-12.88%), Tausonia (4.61%-6.95%), and Fusarium (2.99%-3.97%) were the dominant genera in the four inoculation groups. The abundance of Cyberlindnera showed a significant downward trend compared with the CK group, while the abundance of Mortierella showed a significant upward trend compared with the CK group. Compared with the CK group, the abundance of Pseudeurotium showed a significant upward trend in both the D79 and MIX groups.

[0106] In summary, the present invention isolated a Bacillus sp. D79 from the rhizosphere of northeastern mountain cherry, and resuspended Bacillus sp. D79 to a concentration of about 10 7 CFU / mL was used to obtain the Bacillus D79 suspension, and the root irrigation of cherry seedlings was carried out with the Bacillus D79 suspension, which promoted the growth and development of cherry seedlings. The compound bacterial agent MIX was obtained by compounding the Bacillus D79 suspension with the D27 suspension, and the cherry seedlings were treated with the compound bacterial agent for root irrigation. It was found that the promotion effect on cherry growth was significantly higher than that of the single application of the Bacillus D79 suspension.

[0107] Although preferred embodiments of the present invention have been described, additional changes and modifications may occur to these embodiments once those skilled in the art are aware of the basic inventive concepts.

[0108] Obviously, those skilled in the art may make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include these modifications and variations.

Claims

1. A strain of Bacillus sp. D79, characterized in that: The Bacillus sp.D79 was deposited in the General Microbiology Center of the China Microbiological Culture Collection Administration on September 25, 2024, with the deposit number: CGMCC No.32070, and the classification name is Bacillus sp.

2. A Bacillus D79 bacterial suspension, characterized in that: The Bacillus D79 of claim 1 is cultured to a bacterial concentration of 1 x 10 7 CFU / mL~6ⅹ10 7 CFU / mL was obtained.

3. A composite bacterial agent, characterized in that: The method is obtained by compounding the Bacillus D79 bacterial suspension and the Enterobacter D27 bacterial suspension according to claim 2, wherein the Enterobacter D27 bacterial suspension is prepared from Enterobacter D27; The Enterobacter D27 is deposited in the General Microbiology Center of China Microorganism Culture Collection Administration, with the deposit number: CGMCC No.29568.

4. The composite bacterial agent according to claim 3, characterized in that: The bacterial concentration of the Enterobacter D27 bacterial suspension is 1×10 7 CFU / mL~6ⅹ10 7 CFU / mL.

5. A method for preparing the composite bacterial agent according to claim 3, characterized in that: Prepare Bacillus D79 bacterial suspension and Enterobacter D27 bacterial suspension respectively so that the bacterial concentration is 1×10 7 CFU / mL~6ⅹ10 7 CFU / mL, then the Bacillus D79 bacterial suspension and the Enterobacter D27 bacterial suspension were mixed in a volume ratio of 1 to 2:1; The preparation steps of the Bacillus D79 bacterial suspension are as follows: activating the Bacillus D79, inoculating it in a liquid culture medium and culturing it to a logarithmic growth phase, centrifuging it, discarding the supernatant, and resuspending the bacterial cell with sterile water to obtain the Bacillus D79 bacterial suspension.

6. The method for preparing the composite bacterial agent according to claim 5, characterized in that: The centrifugation condition is 4°C, 8000-12000 r / min, for 10-15 min.

7. The method for preparing the composite bacterial agent according to claim 5, characterized in that: The concentration of the Bacillus D79 bacterial suspension is 10 7 CFU / mL.

8. Use of the Bacillus sp. D79 according to claim 1, the Bacillus D79 bacterial suspension according to claim 2, or the composite bacterial agent according to claim 3 in promoting the growth of fruit trees.

9. The use according to claim 8, characterized in that: The Bacillus sp. D79 or the compound bacterial agent is used for increasing the biomass of cherry plants.

10. The use according to claim 8, characterized in that: The Bacillus sp. D79 or the compound bacterial agent is used for increasing the total root length, root surface area, root volume and average root diameter of the cherry root system.

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