A strain of Bacillus acidiceler CNBG-PGPR-17 and its application

Abstract

The present invention discloses a strain of Bacillus acidiceler CNBG-PGPR-17 and its application; the strain was deposited in the General Microbiology Center of the China Microorganism Culture Collection Administration on October 28, 2021, with a deposit number of CGMCC No. 23673, and is classified as Bacillus acidiceler. The strain has a fast growth rate, good stability, and is easy to scale up and culture; it has a strong ability to dissolve inorganic phosphorus, and the amount of inorganic phosphorus dissolved can reach 277.8 mg / kg after 7 days of inoculation; it can secrete small molecule organic acids such as lactic acid and succinic acid, and reduce the pH value of the solution; it can increase the content of soil organic matter, total nitrogen, available phosphorus and available potassium, and reduce the pH value of the soil; it promotes the growth of blueberry seedlings and improves the quality of blueberries.

Description

Technical Field

[0001] The invention relates to a Bacillus acidiceler CNBG-PGPR-17 and an application thereof, belonging to the field of biotechnology. Background Art

[0002] Blueberries are an economic berry species belonging to the genus Vaccinium in the Ericaceae family. Their fruit boasts beautiful color, unique flavor, and is rich in vitamins, minerals, and antioxidants, offering both high nutritional value and health benefits. Blueberries contain numerous polyphenols, including flavonoids, phenolic acids, and anthocyanins, which are known to significantly improve inflammation, diabetes, and cardiovascular disease. They are listed by the Food and Agriculture Organization of the United Nations as one of the five health foods for humans. In recent years, with people's growing demand for a better and healthier lifestyle, the market demand for blueberries has also grown.

[0003] Blueberries have thin, shallow roots without root hairs, and are poorly able to absorb nutrients. Blueberries are oligotrophic plants, so fertilization must meet their growth needs while avoiding excessive amounts, which can negatively impact the root system and reduce fruit yield (Bryla and Machado, 2011). Furthermore, blueberries thrive in sandy loam soils with a strong acidity (pH 4.5-4.8), high organic matter content (>5%), and good aeration. These specific soil requirements have become a major constraint on the development of my country's blueberry industry.

[0004] Soil growth-promoting rhizobacteria (PGPR) play an important regulatory role in blueberry root growth and nutrient absorption. PGPR can synthesize substances that have direct effects on plant growth and development, such as growth hormones and siderophores; modify the forms of elements in the soil, enabling their efficient absorption by plants, such as through phosphorus solubilization and nitrogen fixation; inhibit or mitigate the occurrence of soil-borne diseases, induce systemic resistance in plants, and enhance their own disease defense mechanisms, thereby improving plant growth and quality. Bacillus bacteria are currently the most commonly reported plant growth-promoting rhizobacteria. However, research on blueberry rhizobacteria has focused more on mycorrhizal fungi than on bacteria. Bacillus acidiceler is a member of the genus Bacillus. Compared to other Bacillus species, such as Bacillus subtilis, Bacillus megaterium, and Bacillus amyloliquefaciens, research on Bacillus acidiceler is currently limited. Xing Mingzhen et al. (CN111154675B) found that acid-fast Bacillus SYY15 can solubilize phosphate, produce IAA and protease, improve the germination ability of corn seeds under salt stress, and promote corn growth. Wang Qiang et al. (CN104630095B) isolated a strain of acid-fast Bacillus HS3 from the peanut rhizosphere, which can solubilize potassium and phosphate and promote peanut growth. However, there are currently no reports on acid-fast Bacillus in the blueberry rhizosphere. Therefore, exploring new, high-quality acid-fast Bacillus germplasm resources and expanding their application areas are of great significance to promoting the development of green and efficient agriculture. Summary of the Invention

[0005] Purpose of the invention: The technical problem to be solved by the present invention is to provide a strain of Bacillus acidiceler CNBG-PGPR-17 and its application.

[0006] Technical solution: In order to solve the above technical problems, the present invention provides a strain of Bacillus acidiceler CNBG-PGPR-17 isolated from the rhizosphere soil of blueberry. The Bacillus acidiceler CNBG-PGPR-17 was deposited in the General Microbiology Center of China Culture Collection Administration on October 28, 2021, with the deposit number: CGMCC No. 23673, and the classification name is Bacillus acidiceler.

[0007] The present invention also provides a microbial composite preparation, comprising the acid-fast-growing Bacillus (Bacillus acidiceler) CNBG-PGPR-17.

[0008] The present invention also provides the use of the Bacillus acidiceler CNBG-PGPR-17 or the microbial composite preparation in producing acidic substances.

[0009] The present invention also provides the use of the Bacillus acidiceler CNBG-PGPR-17 or the microbial composite preparation in nitrogen fixation.

[0010] The present invention also provides use of the Bacillus acidiceler CNBG-PGPR-17 or the microbial composite preparation in dissolving phosphorus.

[0011] The present invention also provides use of the Bacillus acidiceler CNBG-PGPR-17 or the microbial composite preparation in promoting plant growth.

[0012] Wherein, the plant comprises tomato or blueberry.

[0013] The present invention also provides a growth-promoting agent, which contains the acid-fast-growing Bacillus (Bacillus acidiceler) CNBG-PGPR-17 or the microbial composite preparation.

[0014] The present invention also provides use of the Bacillus acidiceler CNBG-PGPR-17 or the microbial composite preparation in regulating pH in an environment.

[0015] Wherein, the environment includes a culture medium environment or a soil environment.

[0016] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages: 1. The acid-fast-growing Bacillus (Bacillus acidiceler) CNBG-PGPR-17 of the present invention has a fast growth rate, good stability, and is easy to scale up and culture; 2. The acid-fast-growing Bacillus (Bacillus acidiceler) CNBG-PGPR-17 has a strong ability to dissolve inorganic phosphorus, and the amount of inorganic phosphorus dissolved can reach 277.8 mg / kg after 7 days of inoculation; 3. The acid-fast-growing Bacillus (Bacillus acidiceler) CNBG-PGPR-17 can secrete small molecular organic acids such as lactic acid and succinic acid, and reduce the pH of the solution. The lactic acid content in the fermentation broth can reach 0.27 kg / L after 24 hours of inoculation, and the pH of the culture medium is reduced by nearly 1 unit; 4. The acid-fast-growing Bacillus (Bacillus acidiceler) CNBG-PGPR-17 can secrete small molecular organic acids such as lactic acid and succinic acid, and reduce the pH of the solution. acidiceler) CNBG-PGPR-17 can increase the content of soil organic matter, total nitrogen, available phosphorus and fast-acting potassium, reduce soil pH, and promote the growth of blueberry seedlings; 5. At the same time, Bacillus acidiceler) CNBG-PGPR-17 can increase the content of soluble protein, soluble sugar and anthocyanin in blueberry fruits, which increased by 17.7%, 10.9% and 10.5% respectively compared with the control, thereby improving the quality of blueberries; 6. In addition, inoculation with Bacillus acidiceler (Bacillus acidiceler) CNBG-PGPR-17 can promote the growth of tomatoes, increasing their plant height, aboveground fresh weight and root weight by 13.9%, 40.3% and 44.1% respectively. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 The colony morphology is that of Bacillus acidiceler CNBG-PGPR-17;

[0018] Figure 2 The cell morphology of Bacillus acidiceler CNBG-PGPR-17 under a 100x microscope;

[0019] Figure 3 This is the phylogenetic tree of Bacillus acidiceler CNBG-PGPR-17;

[0020] Figure 4 This is the growth curve of Bacillus acidiceler CNBG-PGPR-17;

[0021] Figure 5The effect of pH on the growth and reproduction of Bacillus acidiceler CNBG-PGPR-17;

[0022] Figure 6 The effect of Bacillus acidiceler CNBG-PGPR-17 on the growth and development of blueberry plants.

[0023] Figure 7 This is the promoting effect of Bacillus acidiceler CNBG-PGPR-17 on tomato biomass. DETAILED DESCRIPTION

[0024] The technical solution of the present invention will be further described below with reference to the accompanying drawings.

[0025] Example 1 Isolation and Identification of Bacillus acidiceler CNBG-PGPR-17

[0026] (1) Isolation and purification of strains

[0027] 0.50 g of blueberry rhizosphere soil sample (collected from the blueberry planting base in Jiangning District, Nanjing, Jiangsu Province, N118°40'19", E31°50'10") was suspended and shaken in 4.50 mL of sterile saline to obtain 10 -1 Gradient); draw 0.50mL from the dilution into a sterile centrifuge tube, add to 4.50mL sterile saline, suspend and shake, and obtain 10 -2 Gradient; and so on, gradient dilution until the dilution reaches 10 -6 Gradient. Take 100 μL of each dilution from the six dilution gradients and spread it on a nutrient agar (NA) plate. Prepare two replicates in parallel, invert them, and incubate them in a 30°C constant temperature incubator for 24-36 hours. Remove all plates with colonies and select dilution plates with obvious single colonies and a moderate number. Pick the colonies with different morphologies on the plates and transfer them to new NA plates for partitioning and purification. Repeat the purification until single colonies appear, and repeat the streak culture for more than 5 times. Perform morphological determination of the strain according to the Bergey's Manual of Bacterial Identification.

[0028] (2) Strain identification

[0029] Single colonies were picked and cultured in NB medium at 30°C, shaking at 200 rpm overnight. Genomic DNA was extracted using a bacterial genomic DNA extraction kit (Hunan Aikerui Biotechnology Co., Ltd.). PCR amplification and 16S rDNA sequencing and alignment were performed using the DNA as a template. The PCR amplification system (50 μL) consisted of Taq enzyme mix (25 μL), upstream primer 27F (1 μL), downstream primer 1492R (1 μL), DNA template (1 μL), and ddH2O (22 μL). The sequence of 27F was: SEQ ID NO. 1: 5'-AGA GTT TGATCM TGG CTC AG-3', and the sequence of 1492R was: SEQ ID NO. 2: 5'-GGY TAC CTT GTT ACG ACT T-3'. PCR amplification conditions: initial denaturation temperature: 105°C; first denaturation step: 95°C for 7 min, 95°C for 30 s; second annealing step: 55°C for 30 s; third extension step: 72°C for 90 s; number of cycles: 95°C for 30 s, 33 cycles; fourth final extension step: 72°C for 5 min; fifth hold step: 12°C for 10 min. After the amplified product was confirmed by 1% agarose gel electrophoresis, it was used for AxyPrep TM

[0030] The results are as follows Figure 1 and Figure 2 The colony morphology of Bacillus acidiceler is shown in the figure. On NA agar plates, the colonies are small, round, and generally milky white to pearly gray in color. They are opaque, with a smooth, shiny surface. After several days of culture, the surface of the colonies develops a finely wrinkled texture, forming a wrinkle. The cells are short rods, approximately 4-8 μm in length and 0.8-1 μm in width. They are occasionally arranged in chains and are immotile. Staining indicates Gram-positive bacteria.

[0031] The returned sequencing results were compared with the National Center for Biotechnology Information (NCBI) GeneBank database, and the results showed that the strain provided by the present invention was Bacillus acidiceler. According to the latest reports of Peak et al. (Bacillus acidiceler sp.nov.,isolated from a forensic specimen,containing Bacillus anthracis pX02 genes.Int J Syst Evol Microbiol 2007;57:2031-2036.) and Gupta et al. (Robust demarcation of 17distinct Bacillus species clades,proposed as novel Bacillaceae genera,by phylogenomicsand comparative genomic analyses:description of Robertmurraya kyonggiensissp.nov.and proposal for an emended genus Bacillus limiting it only to the members of the Subtilis and Cereus clades of species.Int J Syst Evol Microbiol 2020;70:5753-5798.), Bacillus acidiceler was classified as Gottfriedia acidiceler. The phylogenetic tree was constructed using the maximum likelihood method using MEGA 7 software, and it was found that the strain was closely related to Gottfriedia acidiceleris CBD 119. T Gather into a cluster ( Figure 3 ), it can be determined that GNBG-PGPR-17 is an acid-fast-growing Bacillus.

[0032] The obtained Bacillus acidiceler CNBG-PGPR-17 strain was deposited in the General Microbiology Center of China Culture Collection Administration (CGMCC) on October 28, 2021, with the deposit number: CGMCC No. 23673, and the classification name was Bacillus acidiceler.

[0033] Example 2 Growth Curve of Bacillus acidiceler CNBG-PGPR-17

[0034] A single colony of Bacillus acidiceler CNBG-PGPR-17 was picked and activated in NA liquid medium. After activation for 2 generations, it was inoculated into the 4th generation NB medium at a 5% inoculum size and cultured at 30°C for 72 h. The OD value (absorbance value) of the bacterial solution at a wavelength of 600 nm was measured every 0.5 h starting from 0 h. The measurements were repeated three times in parallel, and a growth curve was plotted with time as the horizontal axis and absorbance as the vertical axis.

[0035] Depend on Figure 4 It can be seen that Bacillus acidiceler CNBG-PGPR-17 can enter the mid-logarithmic phase in 7 hours, and OD 600 The value was 1.06, and then the growth rate slowed down and reached the early stage of stability around the 24th hour. The stable period lasted for a long time. The OD 600 The value is between 1.54 and 1.59, indicating that the acid-fast growing Bacillus (Bacill usacidiceler) CNBG-PGPR-17 of the present invention has a fast growth rate and good stability.

[0036] Example 3 Suitable pH range for growth of Bacillus acidiceler CNBG-PGPR-17

[0037] Prepare 0.1 mol / L HCl and NaOH to adjust the pH of nutrient agar liquid medium (NB) to 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10.0, 11.0, and 12.0, respectively. Sterilize and add to a 50 mL sterile centrifuge tube. Remove the Bacillus acidiceler CNBG-PGPR-17 seed solution (bacterial concentration 1.56 × 10 8 cfu / mL), the strain was added to the above-mentioned NB liquid culture medium with different pH values ​​at a volume ratio of 5% in a clean bench, and cultured in a shaking incubator at 30°C and 160 rpm for 3 days. The OD 600Value, see the result Figure 5 It can be seen that the pH range suitable for the growth of Bacillus acidiceler CNBG-PGPR-17 is 5.0-9.0, and the growth is best in the weakly acidic to neutral range.

[0038] Example 4 Phosphate Solubilization Ability of Bacillus acidiceler CNBG-PGPR-17

[0039] (1) Culture medium configuration

[0040] Prepare inorganic phosphorus liquid culture medium, whose ingredients are 10g glucose, 5g tricalcium phosphate, 0.1g calcium carbonate, 0.5g magnesium sulfate, 0.5g ammonium sulfate, 0.1g calcium sulfate, 0.005g ferric chloride, and 1L water (Note: calcium carbonate needs to be sterilized separately before adding to the plate).

[0041] (2) Quantitative experiment on inorganic phosphorus solubility

[0042] The tube containing Bacillus acidiceler CNBG-PGPR-17 stored in a -80°C refrigerator was removed, thawed on ice until fluid, and mixed by vortexing. The bacterial solution was picked with an inoculating loop and streaked onto a NA solid plate. The plate was incubated at 30°C for 24 hours. A single colony was picked and purified twice and identified and confirmed. After confirmation, a single colony was picked and activated in NB medium. The medium was removed by centrifugation, and the bacterial sludge was resuspended in an equal volume of sterile saline. The resuspended solution of Bacillus acidiceler CNBG-PGPR-17 was added to the inorganic phosphorus liquid medium at an inoculum size of 5% by volume, that is, 5 mL of the resuspended bacterial solution was added to each 250 mL conical flask. Three replicates were set for each treatment group, and a blank treatment group without bacterial solution was set up. Dynamic sampling was performed to determine the phosphorus content in the liquid.

[0043] In this example, the inorganic phosphorus culture medium used tricalcium phosphate as the phosphorus source, and the added amount was 999.42 mg / L. The determination method was based on the "phosphomolybdenum blue spectrophotometry method" in the national standard "Marine Monitoring Specification Part 4: Seawater Analysis" GB 17378.4-2007 to determine the inorganic phosphate in the solution. The results are shown in Table 1.

[0044] Table 1 Inorganic phosphorus content released in culture medium (mg / L)

[0045]

[0046] It can be seen that after inoculating acid-fast bacillus (Bacillus acidiceler) CNBG-PGPR-17, phosphate content in solution gradually increases with incubation time. When inoculating 1d, the inorganic phosphorus content dissolved by acid-fast bacillus (Bacillus acidiceler) CNBG-PGPR-17 provided by the present invention has reached 168.7mg / L, and the amount of dissolved phosphorus increases rapidly over time. Inoculate the 7th day, the inorganic phosphorus content dissolved reaches 277.8mg / L, which is much higher than the reported bacterial strain SYY15D incubated for 7 days. The inorganic phosphorus solubility capacity 152.36mg / L (CN111154675B). The above results show that the acid-fast bacillus CNBG-PGPR-17 provided by the present invention can quickly dissolve inorganic phosphorus and release it into the surrounding environment, and can maintain a long-term release capacity.

[0047] Example 5 Acid production capacity of Bacillus acidiceler CNBG-PGPR-17

[0048] Remove the tube containing Bacillus acidiceler CNBG-PGPR-17 from a -80°C freezer, thaw on ice until fluid, and vortex to mix. Use an inoculating loop to streak the culture solution onto a NA solid plate. Incubate the plate at 30°C for 24 hours. Single colonies are picked and purified twice and confirmed. Once confirmed, single colonies are activated in NA liquid medium. Subsequently, a 5% by volume inoculum of Bacillus acidiceler CNBG-PGPR-17 is inoculated into a sterile centrifuge tube containing 20 mL of NB medium. NB medium without the strain is used as a control. Three replicates are used for each treatment. Incubate the tube in a shaker at 30°C, 160 rpm, for 24 hours. Centrifuge the tube, and measure the pH of the filtrate. The results showed that the pH of the NB medium without inoculation was 7.18±0.02, and the pH of the medium inoculated with Bacillus acidiceler CNBG-PGPR-17 was 6.21±0.03, indicating that Bacillus acidiceler CNBG-PGPR-17 has the ability to secrete acidic substances.

[0049] Furthermore, 1.00 mL of the fermentation broth from the strain after 24 h of culture was accurately pipetted into a centrifuge tube, centrifuged, and filtered through a 0.22 μm aqueous filter. The types and quantities of small-molecule organic acids in the culture broth were analyzed using ESI-HPLC-MS / MS. Instrumental conditions included an LC-30A liquid chromatograph (Shimadzu, Japan) and a SCIEX-4500Qtrap (MSMS) triple quadrupole liquid chromatography-mass spectrometer (AB, USA). Chromatographic conditions included a Waters BEH C18 column, column temperature of 30°C, mobile phase A consisting of methanol and mobile phase B consisting of 1‰ formic acid, flow rate of 0.3 mL / min, and elution gradient: 0 min, 20% A; 1 min, 20% A; 3 min, 50% A; 9 min, 80% A; 10.5 min, 80% A; 10.6 min, 20% A; 13.5 min, stop. The mass spectrometry parameters were: ESI negative ion mode, MRM scan type, curtain gas 15 psi, spray voltage -4000 V, nebulizer gas pressure 65 psi, auxiliary gas pressure 70 psi, and nebulizer temperature 400° C. The results are shown in Table 2.

[0050] Table 2 Types and contents of organic acids released from strain culture filtrates (ng / mL)

[0051]

[0052] It can be seen that the organic acids secreted by Bacillus acidiceler CNBG-PGPR-17 are mainly lactic acid and succinic acid, reaching 0.27g / L and 12.1mg / L respectively. In addition, Bacillus acidiceler CNBG-PGPR-17 can also secrete certain amounts of malic acid, citric acid, oxalic acid, malonic acid, and shikimic acid.

[0053] Example 6 Nitrogen Fixation Ability of Bacillus acidiceler CNBG-PGPR-17

[0054] (1) Ashby nitrogen-free solid culture medium (Catalogue of Chinese Agricultural Fungi, 2001)

[0055] Components: glucose 10g, potassium dihydrogen phosphate 0.2g, magnesium sulfate heptahydrate 0.2g, sodium chloride 0.2g, calcium sulfate dihydrate 0.1g, calcium carbonate 5g, agar 20g, distilled water 1000mL.

[0056] (2) Slice culture

[0057] A single colony of Bacillus acidiceler CNBG-PGPR-17 was picked from a NA solid medium plate and inoculated onto Ashby nitrogen-free solid medium. The streak method involved dividing the Ashby nitrogen-free medium into three zones: I, II, and III. A single colony was picked from the NA solid medium and streaked in a zigzag pattern across Zone I of the Ashby nitrogen-free medium. A sample of the bacterial suspension from Zone I was then streaked in a zigzag pattern across Zone II of the Ashby nitrogen-free medium. A sample of the bacterial suspension from Zone II was then streaked in a zigzag pattern across Zone III of the Ashby nitrogen-free medium. The inoculated Ashby nitrogen-free solid medium plates were incubated at 30°C for 48 hours, and the growth of the strain on the plates was observed. The results showed that Bacillus acidiceler CNBG-PGPR-17 grew in all three streaked zones of the Ashby nitrogen-free medium, demonstrating its ability to fix nitrogen.

[0058] Example 7 Growth-promoting ability of Bacillus acidiceler CNBG-PGPR-17 on blueberry seedlings

[0059] A study evaluating the growth-promoting ability of Bacillus acidiceler CNBG-PGPR-17 on blueberry seedlings was conducted at the Blueberry Base of the Institute of Botany, Chinese Academy of Sciences, Jiangsu Province. The cultivation medium used consisted of garden soil: peat: perlite in a ratio of 6:2:2. Two treatments were set up: an untreated control (CK) and a root irrigated with the inoculum (Ba), with six replicates per treatment. The medium was mixed according to the above proportions. 1.0 kg of the mixed medium was weighed and placed in a pot (15 cm diameter, 15 cm height). After watering to settle, six-month-old blueberry tissue culture seedlings (Vaccinium corymbosum 'Lanmei 1') of uniform growth were transplanted into the plastic pot. A single colony of Bacillus acidiceler CNBG-PGPR-17 was picked from the NA medium with an inoculation loop and activated in NB medium. The strain was then inoculated into NB medium at a volume ratio of 5%, cultured in a constant temperature shaker at 30°C and 160 rpm for 48 h, centrifuged, and the bacterial sludge was resuspended in sterile water and diluted to 1×10 7cfu / mL. One week after transplanting the blueberry seedlings, 200 mL of bacterial solution was applied around the root system. Thereafter, this was repeated every two weeks. A control group received 200 mL of water for root irrigation. Potted plants were cultured in the open field. During the experiment, 0.1 or 0.2 L of water was applied to each pot, depending on soil moisture conditions, to keep the soil moist but not waterlogged. All treatments were irrigated every two weeks with a 5‰ fertilizer solution containing (NH₄)₂SO₄:KH₂PO₄ in a ratio of 3:2, with 100 mL applied to each plant. Pest and disease control measures were implemented promptly to ensure normal plant growth. Plant height, total branch length, leaf area, and dry weight of the blueberry seedlings were measured five months later (Table 3). Soil samples were collected, air-dried, sieved, and their basic physical and chemical properties determined (Table 4).

[0060] Table 3 Effect of acid-fast Bacillus CNBG-PGPR-17 on the growth of blueberry seedlings

[0061]

[0062] Note: Different letters in each column indicate significant differences among treatments, p < 0.05.

[0063] As shown in Table 3, Bacillus acidiceler CNBG-PGPR-17 significantly promoted the growth of blueberry seedlings. Compared with the control, plant height, total branch length, and leaf area of ​​the blueberry treated with Bacillus acidiceler CNBG-PGPR-17 increased by 7.2%, 10.5%, and 24.2%, respectively. Root, stem, leaf, and total plant dry weight increased by 25%, 15.9%, 36%, and 25.1%, respectively.

[0064] Table 4 Effects of acid-fast-growing Bacillus CNBG-PGPR-17 on basic physical and chemical properties of soil

[0065]

[0066] Note: Different letters in each column indicate significant differences among treatments, p < 0.05.

[0067] As shown in Table 4, Bacillus acidiceler CNBG-PGPR-17 significantly reduced soil pH, increased soil organic matter and available nutrient content, and improved soil water and fertilizer retention. Compared to the control, treatment with Bacillus acidiceler CNBG-PGPR-17 decreased soil pH by 0.3 units, increased soil organic carbon by 22.6%, and increased soil total nitrogen, available phosphorus, and available potassium by 34.9%, 50.7%, and 21.6%, respectively. These changes in soil physical and chemical properties are beneficial to the growth of blueberry seedlings.

[0068] Example 8 Effect of Acidobacillus CNBG-PGPR-17 on Blueberry Fruit

[0069] An experiment was conducted at the Blueberry Planting Base in Jiangning District, Nanjing City to evaluate the effect of acid-fast-growing Bacillus CNBG-PGPR-17 on the size and quality of blueberry fruit. Four-year-old O'Neill blueberry trees with consistent growth were selected from the same field. Two treatments were set up: an untreated control (CK) and a root irrigated with a fungal agent (Ba). Each treatment group had four replicates, and each replicate treated six fruit trees. A single colony of acid-fast-growing Bacillus (Bacillus acidiceler) CNBG-PGPR-17 was picked from NA medium with an inoculation loop and activated in NB medium. Subsequently, the strain was inoculated into NB medium at a volume ratio of 5%, cultured in a constant temperature shaker at 30°C and 160r / min for 48h, centrifuged, and the bacterial sludge was resuspended in sterile water and diluted to 1×10 7 cfu / mL. 200 mL of bacterial solution was poured around the blueberry root system, and then poured once every two weeks. At the same time, the control group was irrigated with 200 mL of water. Other field management methods were consistent with routine orchard management. The experiment began on March 15, 2022. When the blueberry fruits were ripe, blueberry fruits were randomly collected from the four directions of east, south, west and north, with 40 fruits per plant. Single fruit weight, soluble protein, anthocyanin and other quality indicators were measured (Table 5).

[0070] The results are shown in Table 5. As can be seen, the strain CNBG-PGPR-17 promoted blueberry fruit growth to a certain extent. Compared with the control, the average weight of blueberries treated with CNBG-PGPR-17 increased by nearly 5%, but the weight gain was not significant. Furthermore, CNBG-PGPR-17 increased the content of soluble protein, soluble sugar, and anthocyanin in blueberries by 17.7%, 10.9%, and 10.5%, respectively, compared with the control, improving blueberry fruit quality.

[0071] Table 5 Effects of acid-fast Bacillus CNBG-PGPR-17 on basic physical and chemical properties of soil

[0072]

[0073] Example 9 Growth-promoting ability of acid-fast Bacillus CNBG-PGPR-17 on tomato seedlings

[0074] An experiment to evaluate the growth-promoting ability of Bacillus acidiceler CNBG-PGPR-17 on tomato seedlings was conducted in the light room of the Institute of Botany, Chinese Academy of Sciences, Jiangsu Province. The soil used in the experiment was collected from the experimental field of the Institute of Botany, Chinese Academy of Sciences, Jiangsu Province. After the soil was air-dried, it was placed in a plug tray. Two treatments were set up: an untreated control (CK) and a root irrigation with a bacterial agent (Ba). Each treatment group had 4 replicates, and each replicate contained 4 tomato seedlings. A single colony of Bacillus acidiceler CNBG-PGPR-17 was picked from the NA culture medium with an inoculation loop and activated in a liquid culture medium. Subsequently, the strain was inoculated into the NA liquid culture medium at a volume ratio of 5%, and cultured in a constant temperature shaker at 30°C and 160r / min for 48h. After centrifugation, the bacterial sludge was resuspended in sterile water and diluted to 1×10 7 cfu / mL. Tomato seeds were sown in advance for seedling cultivation. After the seedlings grew two leaves and a heart, uniformly sized seedlings were transplanted into plug trays, one per seedling. One week after transplanting, 10 mL of fungus solution was applied to each tray. A control treatment was not treated with fungus, but with 10 mL of water. Thirty days after treatment, the plant height and fresh weight of the tomatoes were measured.

[0075] The results of the effect of Bacillus acidiceler CNBG-PGPR-17 on the growth of tomato seedlings are shown in Figure 6 and 7 The results showed that the application of Bacillus acidiceler CNBG-PGPR-17 significantly increased tomato plant height and biomass, and promoted the growth of tomato seedlings. Compared with the untreated control, plant height, aboveground fresh weight, and root weight of plants treated with Bacillus acidiceler CNBG-PGPR-17 increased by 13.9%, 40.3%, and 44.1%, respectively.

Claims

1. A strain of acid-fast Bacillus ( Bacillus acidiceler ) CNBG-PGPR-17, characterized in that The acid-fast Bacillus ( Bacillus acidiceler ) CNBG-PGPR-17 was deposited in the General Microbiology Center of China Culture Collection Administration on October 28, 2021, with the deposit number: CGMCC No. 23673, and the classification name was Bacillus acidophilus ( Bacillus acidiceler ).

2. A microbial composite preparation, characterized in that: The microbial composite preparation comprises the acid-fast-growing Bacillus ( Bacillus acidiceler )CNBG-PGPR-17.

3. The acid-fast-growing Bacillus according to claim 1 ( Bacillus acidiceler ) Use of CNBG-PGPR-17 or the microbial composite preparation according to claim 2 in the production of acidic substances.

4. The acid-fast-growing Bacillus according to claim 1 ( Bacillus acidiceler ) Use of CNBG-PGPR-17 or the microbial composite preparation described in claim 2 in dissolving phosphorus.

5. The acid-fast-growing Bacillus according to claim 1 ( Bacillus acidiceler ) Use of CNBG-PGPR-17 or the microbial composite preparation according to claim 2 in promoting plant growth.

6. The use according to claim 5, characterized in that The plants include tomatoes or blueberries.

7. A growth-promoting agent, characterized in that It contains the acid-fast-growing Bacillus ( Bacillus acidiceler ) CNBG-PGPR-17 or the microbial composite preparation according to claim 2.

8. The acid-fast-growing Bacillus according to claim 1 ( Bacillus acidiceler ) Use of CNBG-PGPR-17 or the microbial composite preparation according to claim 2 in regulating pH in an environment.

9. The use according to claim 8, characterized in that The environment includes a culture medium environment or a soil environment.

10. The acid-fast-growing Bacillus according to claim 1 ( Bacillus acidiceler ) Use of CNBG-PGPR-17 or the microbial composite preparation according to claim 2 in improving the quality of blueberry fruit.

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