Bacillus acidophilus CNBG-PGPR-17 and application thereof

Abstract

The invention discloses a strain of bacillus acidophilus CNBG-PGPR-17 and an application of the bacillus acidophilus CNBG-PGPR-17. The invention further discloses a preparation method of the bacillus acidophilus CNBG-PGPR-17. The strain is preserved in the China General Microbiological Culture Collection Center (CGMCC) on October 28, 2021, the preservation number is CGMCC No.23673, and the strain is classified and named as bacillus acidogenes (Bacillus acidogenes). The strain is high in growth speed, good in stability and easy to amplify and culture; the inorganic phosphorus dissolving capacity is high, and the inorganic phosphorus dissolving amount can reach 277.8 mg / kg 7 days after inoculation; small molecular organic acids such as lactic acid and succinic acid can be secreted, and the pH of the solution is reduced; the contents of soil organic matters, soil total nitrogen, available phosphorus and quick-acting potassium can be increased, and the pH value of the soil is reduced; the growth of blueberry seedlings is promoted, and the blueberry quality is improved.

Description

Technical Field

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

[0002] Blueberry is an economic berry tree of the genus Vaccinium in the family Ericaceae. Its fruit has beautiful color and unique flavor. It is rich in vitamins, minerals and antioxidants, and has high nutritional value and medical and health care functions. Blueberry fruit contains a large number of polyphenols such as flavonoids, phenolic acids, anthocyanins, etc., which have a good improvement effect on inflammation, diabetes, cardiovascular diseases, etc., and is listed as one of the five major health foods for humans by the Food and Agriculture Organization of the United Nations. In recent years, with the growing demand for a better and healthier life, the market demand for blueberries has been increasing day by day.

[0003] Blueberries have thin roots, no root hairs, and shallow roots, and their ability to absorb nutrients is poor. Blueberries are oligotrophic plants, and fertilization must meet their growth needs, but not be excessive, otherwise it will have an adverse effect on the root system and affect fruit yield (Bryla and Machado, 2011). In addition, blueberries are suitable for growing in highly acidic (pH 4.5-4.8), high in organic matter (>5%), and well-ventilated sandy loam. The special requirements of blueberries for soil conditions have become a major problem restricting 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 certain substances that have a direct effect on plant growth and development, such as growth hormones and siderophores; change the form of elements in the soil so that they can be effectively absorbed by plants, such as phosphorus dissolution and nitrogen fixation; inhibit or reduce the occurrence of soil-borne diseases, induce plant systemic resistance, and improve the plant's own defense mechanism against diseases, thereby improving plant growth and quality. Bacillus bacteria are currently the most reported plant rhizosphere growth-promoting bacteria. However, in the research on blueberry rhizosphere growth-promoting bacteria, mycorrhizal fungi are more studied, and bacteria are less studied. Bacillus acidiceler is a type of Bacillus bacteria. Compared with other bacteria of the genus Bacillus, such as Bacillus subtilis, Bacillus megaterium, and Bacillus amyloliquefaciens, there is currently less research on Bacillus acidiceler. Xing Mingzhen et al. (CN111154675B) found that acid-fast Bacillus SYY15 can dissolve 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 rhizosphere of peanuts, which can dissolve potassium, dissolve phosphate, and promote peanut growth. However, there is no report on acid-fast Bacillus in the rhizosphere of blueberries. Therefore, it is of great significance to explore new and high-quality acid-fast Bacillus germplasm resources and expand its application fields to promote 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: 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 National Microbiological 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 the use of the Bacillus acidiceler CNBG-PGPR-17 or the microbial composite preparation in dissolving phosphorus.

[0011] The present invention also provides the 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 the 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 to 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 to 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; 5. The acid-fast growing Bacillus (Bacillus acidiceler) CNBG-PGPR-17 can secrete small molecular organic acids such as lactic acid and succinic acid to reduce the pH of the solution. acidiceler) CNBG-PGPR-17 can increase the content of soil organic matter, total nitrogen, available phosphorus and quick-acting potassium, reduce soil pH, and promote the growth of blueberry seedlings; 5. At the same time, Bacillus acidiceler (Bacillus acidiceler) CNBG-PGPR-17 can increase the content of soluble protein, soluble sugar and anthocyanin in blueberry fruit, which increased by 17.7%, 10.9% and 10.5% respectively compared with the control, thereby improving the quality of blueberry; 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 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 plant height and root length of blueberry;

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

[0024] The technical solution of the present invention is further described below in conjunction with the accompanying drawings.

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

[0026] (1) Isolation and purification of strains

[0027] Take 0.50 g of blueberry rhizosphere soil sample (collected from the blueberry planting base in Jiangning District, Nanjing, Jiangsu, N118°40'19", E31°50'10") and suspend and shake 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, perform gradient dilution until the dilution reaches 10 -6 Gradient. Take 100 μL of each of the 6 dilution gradients and spread it on a nutrient agar solid medium (NA) plate. Prepare 2 copies in parallel, invert, and culture in a 30°C constant temperature incubator for 24-36 hours. Take out all the plates with colonies, select the dilution plates with obvious single colonies and a moderate number, pick the colonies with different morphologies on the plates to new NA plates, and perform partition purification. Repeat the purification until a single colony appears, and repeat the streaking culture for more than 5 times. Perform morphological determination of the strain according to the Bergey's Bacterial Identification Manual.

[0028] (2) Strain identification

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

[0030] The results are as follows Figure 1 and Figure 2 As shown in the figure, the colony morphology of Bacillus acidiceler is small and round on the surface of NA plate culture medium. The colonies are milky white to pearl gray in color, opaque, smooth and shiny. After several days of culture, the surface of the colony will become wrinkled and wrinkled. The cells are short rod-shaped, about 4-8μm in length and about 0.8-1μm in width. The cells are occasionally arranged in chains and cannot move. Staining shows 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 by 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 17 distinct Bacillus species clades,proposed as novel Bacillaceae genera,by phylogenomicsand comparative genomic analyses:description of Robertmurraya kyonggiensis sp.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 is classified as the genus 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 China General Microbiology Center (CGMCC) of the China Microbiological Culture Collection Administration 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 culture medium. After 2 generations of activation, it was inoculated into the 4th generation NB culture medium at a 5% inoculation amount and cultured at 30°C for 72h. The OD value (absorbance value) of the bacterial solution at a wavelength of 600nm was measured every 0.5h from 0h, and the measurements were carried out in parallel three times. The growth curve was drawn with time as the horizontal axis and the absorbance value 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 7h, and OD 600 The value was 1.06, and then the growth rate slowed down, reaching the early stable stage around the 24th hour, and the stable stage lasted for a long time. 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 HCl and NaOH with a concentration of 0.1 mol / L, 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, and add them to a 50 mL sterile centrifuge tube after sterilization. Take out the CNBG-PGPR-17 seed liquid of Bacillus acidiceler (Bacillus acidiceler) from the shaker (bacterial concentration 1.56×10 8 cfu / mL), the strain was added to the NB liquid medium with different pH values ​​in a clean bench at a volume ratio of 5%, and cultured in a shaking incubator at 30°C and 160 r / min 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 and then added to the plate).

[0041] (2) Quantitative test of inorganic phosphorus solubility

[0042] The bacterium preservation tube containing Bacillus acidiceler CNBG-PGPR-17 stored in a -80°C refrigerator was taken out, thawed on ice until fluid, and mixed by shaking. The bacterial solution was picked up with an inoculation loop and three-zoned on a NA solid plate. The plate was placed at 30°C for constant temperature culture for 24 hours, and a single colony was picked for purification twice and identification and confirmation. After confirmation, a single colony was picked up and activated in NB culture medium, and the culture medium was removed by centrifugation. The bacterial mud was resuspended in an equal volume of sterile physiological saline, and the resuspended solution of Bacillus acidiceler CNBG-PGPR-17 was added to the inorganic phosphorus liquid culture medium at an inoculum volume ratio of 5%, that is, 5 mL of the resuspended bacterial solution was added to each 250 mL conical flask, and three replicates were set for each treatment group. At the same time, a blank treatment group without adding bacterial solution was set, and dynamic sampling was performed to determine the phosphorus content in the liquid.

[0043] In this embodiment, the inorganic phosphorus culture medium uses tricalcium phosphate as the phosphorus source, and the addition amount is 999.42 mg / L. The determination method refers to the "phosphomolybdenum blue spectrophotometry" 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 the culture medium (mg / L)

[0045]

[0046] It can be seen that after inoculating acid fast-growing bacillus (Bacillus acidiceler) CNBG-PGPR-17, the phosphate content in the solution gradually increases with the culture time. When inoculating 1d, the inorganic phosphorus content dissolved by acid fast-growing bacillus (Bacillus acidiceler) CNBG-PGPR-17 provided by the present invention has reached 168.7mg / L, and the amount of dissolved phosphorus increases rapidly with the extension of time. Inoculate the 7th day, the inorganic phosphorus content dissolved reaches 277.8mg / L, which is much higher than the reported bacterial strain SYY15D culturing 7 days when the inorganic phosphorus solubility capacity 152.36mg / L (CN111154675B). The above results show that the acid fast-growing 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] Take out the preservation tube containing Bacillus acidiceler CNBG-PGPR-17 stored in a -80°C refrigerator, thaw on ice until fluid, shake and mix, pick the bacterial solution with an inoculation loop and make three-zone lines on the NA solid plate, place the plate at 30°C for constant temperature culture for 24 hours, pick a single colony for purification twice and identify and confirm, pick a single colony to activate in NA liquid culture medium after confirmation, then inoculate the strain Bacillus acidiceler CNBG-PGPR-17 in a sterile centrifuge tube containing 20mL NB culture medium at a volume ratio of 5%, and use NB culture medium without the strain as a control, and each treatment is repeated three times. Cultivate in a constant temperature shaker at 30°C and 160r / min for 24 hours, centrifuge, and measure the pH value of the filtrate. The results showed that the pH of the NB medium without inoculation of the strain 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] Further, 1.00 mL of the fermentation broth of the strain after 24 h of culture was accurately taken into a centrifuge tube with a pipette, centrifuged, passed through a 0.22 μm aqueous filter membrane, and analyzed the types and quantities of small molecular organic acids in the culture broth using ESI-HPLC-MS / MS. The instrument conditions were: LC-30A liquid chromatograph (Shimadzu, Japan), SCIEX-4500Qtrap (MSMS) triple quadrupole liquid chromatography-mass spectrometer (AB, USA). The chromatographic conditions were: Waters BEH C18 column, column temperature 30 ° C, mobile phase A was methanol, B was 1‰ formic acid, flow rate 0.3 mL / min, 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, scan type MRM, curtain gas 15 psi, spray voltage -4000 V, nebulizer gas pressure 65 psi, auxiliary gas pressure 70 psi, 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 0.27 g / L and 12.1 mg / L respectively. In addition, Bacillus acidiceler CNBG-PGPR-17 can also secrete a certain amount 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 strains, 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] Single colonies of Bacillus acidiceler CNBG-PGPR-17 were picked from the NA solid medium plate and inoculated on the Ashby nitrogen-free solid medium. Plate streaking method: The Ashby nitrogen-free medium was divided into three areas: I, II, and III; a single colony was picked from the NA solid medium and streaked in a zigzag pattern in area I of the Ashby nitrogen-free medium; the bacterial solution was dipped from area I and streaked in a zigzag pattern in area II of the Ashby nitrogen-free medium; the bacterial solution was dipped from area II and streaked in a zigzag pattern in area III of the Ashby nitrogen-free medium. The inoculated Ashby nitrogen-free solid medium plate was placed in a constant temperature incubator at 30°C for 48 hours, and the growth of the strain on the plate was observed. The results showed that Bacillus acidiceler CNBG-PGPR-17 could grow in all three streaked areas of the Ashby nitrogen-free solid medium, proving that it has a certain nitrogen fixation ability.

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

[0059] The growth-promoting ability of Bacillus acidiceler CNBG-PGPR-17 on blueberry seedlings was evaluated at the blueberry base of the Institute of Botany, Chinese Academy of Sciences, Jiangsu Province. The cultivation medium used in the experiment was garden soil: peat: perlite = 6:2:2. Two treatments were set up: untreated control (CK) and root irrigation with bacterial agent (Ba), and each treatment was set up with 6 replicates. The matrix was mixed according to the above ratio. 1.0 kg of mixed matrix was weighed and potted (15 cm in diameter and 15 cm in height). After watering, the six-month-old blueberry tissue culture seedlings (Vaccinium corymbosum'Lanmei 1') with consistent growth were transplanted into plastic pots. A single colony of Bacillus acidiceler CNBG-PGPR-17 was picked from the NA medium with an inoculation loop and activated in the NB medium. The strain was then inoculated into the NB medium at a volume ratio of 5%, cultured in a constant temperature shaker at 30°C and 160 r / min for 48 h, centrifuged, and the bacterial sludge was resuspended in sterile water and diluted to 1×10 7cfu / mL. One week after the blueberry seedlings were transplanted, 200mL of bacterial solution was taken to water the roots of the blueberry. Thereafter, the bacteria were watered once every two weeks. At the same time, 200mL of water was used to irrigate the roots of the control group. The potted plants were cultured in the open field. During the experiment, 0.1 or 0.2L of water was poured into each pot each time depending on the soil moisture, and the soil was kept moist without water accumulation. All treatments were irrigated once every two weeks with 5‰ mass fraction fertilizer solution, (NH4)2SO4:KH2PO4=3:2 in the fertilizer solution, and 100mL was poured into each plant each time. Pest and disease control was carried out in time to ensure the normal growth of the plants. The plant height, total branch length, leaf area and dry weight of the blueberry seedlings were measured after 5 months (Table 3). Soil samples were collected, air-dried and sieved, and their basic physical and chemical properties were 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 can be seen from Table 3, Bacillus acidiceler CNBG-PGPR-17 significantly promoted the growth of blueberry seedlings. Compared with the control, the plant height, total branch length and leaf area of ​​blueberries in the Bacillus acidiceler CNBG-PGPR-17 treatment group increased by 7.2%, 10.5% and 24.2% respectively; the 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 can be seen from Table 4, Bacillus acidiceler CNBG-PGPR-17 can significantly reduce soil pH, increase soil organic matter and effective nutrient content, and improve soil water and fertilizer retention capacity. Compared with the control, under the treatment of Bacillus acidiceler CNBG-PGPR-17, soil pH decreased by 0.3 units; organic carbon increased by 22.6%, and soil total nitrogen, effective phosphorus and available potassium increased 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 Bacillus acidophilus CNBG-PGPR-17 on blueberry fruits

[0069] An experiment was conducted to evaluate the effect of acid-fast-growing Bacillus CNBG-PGPR-17 on the size and quality of blueberry fruit at Lanhui Manor, a blueberry planting base in Jiangning District, Nanjing. Four-year-old O'Neill blueberry trees with consistent growth were selected from the same field, and two treatments were set up: an untreated control (CK) and a root irrigation with a bacterial 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 the NA medium with an inoculation loop and activated in the NB medium. Subsequently, the strain was inoculated into the NB medium at a volume ratio of 5%, and cultured in a constant temperature shaker at 30°C and 160r / min for 48h. The culture was centrifuged, and the bacterial sludge was resuspended in sterile water and diluted to 1×10 7 cfu / mL. Take 200mL of bacterial solution and pour the bacteria around the blueberry root system. Then pour the bacteria once every two weeks. At the same time, the control group uses 200mL of water for root irrigation. Other field management methods are consistent with routine orchard management. The experiment started on March 15, 2022. When the blueberry fruits were ripe, blueberry fruits were randomly collected from the southeast, northwest, and northeast directions, 40 per plant, and the single fruit weight, soluble protein, anthocyanin and other quality indicators were measured (Table 5).

[0070] The results are shown in Table 5. It can be seen that the acid-fast Bacillus CNBG-PGPR-17 has a certain promoting effect on the growth of blueberry fruit. Compared with the control, the average single fruit weight of blueberry fruit treated with acid-fast Bacillus CNBG-PGPR-17 increased by nearly 5%, but the weight gain effect was not significant. In addition, acid-fast Bacillus CNBG-PGPR-17 can increase the content of soluble protein, soluble sugar and anthocyanin in blueberry fruit, which increased by 17.7%, 10.9% and 10.5% respectively compared with the control, improving the quality of blueberry fruit.

[0071] Table 5 Effects of acid-fast growing 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] The growth-promoting ability of Bacillus acidiceler CNBG-PGPR-17 on tomato seedlings was evaluated 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. The soil was air-dried and put into plug trays. Two treatments were set up: untreated control (CK) and root irrigation with bacterial agent (Ba). Each treatment group was set up with 4 replicates, and each replicate contained 4 tomato seedlings. A single colony of Bacillus acidiceler CNBG-PGPR-17 was picked from the NA medium with an inoculation loop and activated in the liquid culture medium. Then, 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. Centrifugation was performed, and the bacterial sludge was resuspended in sterile water and diluted to 1×10 7 cfu / mL. Tomato seeds were sown in advance for seedling raising. After the seedlings grew two leaves and one heart, seedlings of the same size were selected and transplanted into plug trays. Each seedling had one plug tray, and the bacteria were poured one week after transplanting, with 10 mL per plug tray. The control treatment was not poured with bacteria, but with 10 mL of clean water instead. After 30 days of the bacteria pouring treatment, the plant height and fresh weight of 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 the plant height and biomass of tomatoes and promoted the growth of tomato seedlings. Compared with the control without bacteria, the plant height, aboveground fresh weight and root weight of Bacillus acidiceler CNBG-PGPR-17 treatment increased by 13.9%, 40.3% and 44.1%, respectively.

Claims

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

2. A microbial composite preparation, characterized in that: The microbial composite preparation comprises the Bacillus acidiceler CNBG-PGPR-17 described in claim 1.

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

4. Use of the Bacillus acidiceler CNBG-PGPR-17 according to claim 1 or the microbial composite preparation according to claim 2 in nitrogen fixation.

5. Use of the Bacillus acidiceler CNBG-PGPR-17 according to claim 1 or the microbial composite preparation according to claim 2 in dissolving phosphorus.

6. Use of the Bacillus acidiceler CNBG-PGPR-17 according to claim 1 or the microbial composite preparation according to claim 2 in promoting plant growth.

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

8. A growth-promoting agent, characterized in that It contains the acid-fast-growing Bacillus (Bacillus acidiceler) CNBG-PGPR-17 described in claim 1 or the microbial composite preparation described in claim 2.

9. Use of the Bacillus acidiceler CNBG-PGPR-17 according to claim 1 or the microbial composite preparation according to claim 2 in regulating pH in an environment.

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

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