Acinetobacter of peony rhizosphere and application thereof

Abstract

The present application discloses acaibacterium of peony rhizosphere and application thereof, and belongs to the technical field of microorganisms.The acalbacterium strain provided by the present application has good phosphorus solubilization effect, and can effectively convert insoluble inorganic phosphorus into soluble inorganic phosphorus.The strain can also be applied to the soil environment of plant growth, improve the utilization of phosphorus in the soil and promote the growth of plants, and has important agricultural and environmental values.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology. It relates to Acinetobacter rhizosphereus of peony and its applications, specifically to a strain of Acinetobacter rhizosphereus of peony, and the application of this strain in soil phosphorus solubilization and plant growth promotion. Background Technology

[0002] Phosphorus is one of the essential macroelements for plant growth and development. Natural soils are rich in phosphorus, but due to the strong fixation and adsorption of phosphorus by soil minerals, most phosphorus exists in the form of insoluble phosphates such as calcium phosphate, aluminum phosphate, and iron phosphate. Therefore, over 90% of phosphorus in the soil is in an unavailable state, which limits the absorption and utilization of phosphorus nutrients by plants. Phosphate-solubilizing bacteria can convert insoluble phosphorus in the soil into soluble phosphates that plants can absorb, improving the utilization rate of phosphorus in the soil and promoting plant growth. Current research has identified phosphate-solubilizing bacteria mainly from genera such as *Leptochloa*, *Bacillus*, *Pseudomonas*, and *Klebsiella*. However, research on the rhizosphere strain of *Fengdan* as a phosphate-solubilizing bacterium has not yet been reported. Summary of the Invention

[0003] (1) An Acinetobacter sp. strain was deposited at the China General Microbiological Culture Collection Center on December 12, 2024, with accession number CGMCC No.33053.

[0004] (2) A microbial agent containing Acinetobacter sp. as described in (1).

[0005] (3) The microbial agent according to (2), wherein the content of *Lactobacillus plantarum* as described in (1) in the microbial agent is not less than 10%. 6 cfu / mL or 10 6 cfu / g.

[0006] (4) A method for reducing the content of insoluble inorganic phosphorus, wherein Acinetobacter sp. of (1) or the microbial agent of (2) or (3) is added to the system containing insoluble inorganic phosphorus.

[0007] (5) According to the method of (4), wherein the insoluble inorganic phosphorus is converted into soluble inorganic phosphorus.

[0008] (6) The method according to ((4) or (5), wherein the insoluble inorganic phosphorus includes calcium phosphate, iron phosphate, and aluminum phosphate.

[0009] (7) A method for promoting plant growth, wherein Acinetobacter sp. as described in (1) or a microbial agent as described in (2) or (3) is added to the system for culturing the plant.

[0010] (8) The method according to (7), wherein promoting plant growth includes one or more of increasing plant height, increasing plant stem girth, and increasing plant fresh weight;

[0011] Optionally, the plants include tomatoes, wheat, rice, corn, cotton, rapeseed, soybeans, and peas.

[0012] (9) The application of Acinetobacter sp. as described in (1) or microbial agents as described in (2) or (3) in reducing the content of insoluble inorganic phosphorus.

[0013] (10) The application according to (9), wherein the insoluble inorganic phosphorus is converted into soluble inorganic phosphorus.

[0014] Beneficial effects: Acinetobacter rhizosphereus PoPSB-1 ​​has significant soil phosphorus solubilization capacity and plant growth promotion effect. It can be used to improve soil phosphorus utilization and promote plant growth, and has important agricultural and environmental value.

[0015] Preservation of biological materials

[0016] The Acinetobacter PoPSB-1 ​​provided by this invention, classified and named Acinetobacter sp., was deposited on December 12, 2024, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 33053, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences. Attached Figure Description

[0017] Figure 1 This is a colony morphology diagram of strain PoPSB-1.

[0018] Figure 2 Phylogenetic tree of the 16S rRNA gene of strain PoPSB-1. Detailed Implementation

[0019] In this specification, "alkaline-available nitrogen" refers to the form of nitrogen produced by the decomposition of organic matter under alkaline conditions. It is mainly found in soil organic matter and is one of the main forms of nitrogen in the soil. The release rate of alkaline-available nitrogen is relatively slow, requiring microbial decomposition to be converted into a form that can be absorbed by plants.

[0020] In this specification, "available potassium" refers to potassium in the soil that is easily absorbed and utilized by crops, including soil solution potassium and soil exchangeable potassium. Available potassium accounts for 0.1% to 2% of the total potassium in the soil, of which soil solution potassium accounts for 1% to 2%. Due to its very low proportion, it is often included in exchangeable potassium. The content of available potassium is one of the important indicators characterizing the potassium supply status of the soil.

[0021] In this specification, "available phosphorus" refers to the phosphorus components in the soil that can be absorbed by plants. It includes all water-soluble phosphorus, some adsorbed phosphorus, and organic phosphorus; some soils also contain certain precipitated phosphorus.

[0022] In this specification, "organic matter" refers to substances in the soil that are derived from life, including soil microorganisms and soil animals and their secretions, as well as plant residues and plant secretions in the soil.

[0023] This invention first screened a phosphorus-solubilizing strain, PoPSB-1, from the soil in which the peony 'Fengdan' grows. This strain can convert insoluble phosphorus into soluble phosphorus and has a good conversion effect.

[0024] Tomatoes, as plants that require soluble phosphorus for growth and development, are highly dependent on it. Soluble phosphorus is a key factor in the biological processes of tomato plants, including photosynthesis, energy metabolism, and signal transduction. Soluble phosphorus in the soil can be absorbed by tomato roots and converted into phosphorus compounds needed by the plant, promoting fruit formation and ripening. Therefore, in some exemplary embodiments, the screened phosphorus-solubilizing strain PoPSB-1 ​​was added to the soil in which tomatoes were cultivated. The results showed that after adding the phosphorus-solubilizing strain PoPSB-1, the tomatoes exhibited better growth, and the organic matter content in the soil was significantly increased compared to the control due to the effect of the phosphorus-solubilizing strain PoPSB-1.

[0025] Example

[0026] In the examples, the determination of alkaline nitrogen, available potassium, available phosphorus and organic matter content were carried out in accordance with LY / T 1228-2015, NY / T889-2004, NY / T 1121.7-2014 and NY / T 1121.6-2006, respectively.

[0027] Example 1

[0028] 1. Prepare liquid and solid culture media containing insoluble inorganic phosphorus for the isolation and screening of phosphate-solubilizing bacteria.

[0029] PVF Inorganic Phosphorus Liquid Culture Medium: 10.0g glucose, 0.5g ammonium sulfate, 0.3g sodium chloride, 0.3g magnesium sulfate heptahydrate, 0.03g manganese sulfate, 0.3g potassium chloride, 0.03g ferrous sulfate heptahydrate, 5.0g calcium phosphate, 1000mL distilled water, pH 6.8-7.2, sterilized at 121℃ for 20min.

[0030] PVF Inorganic Phosphorus Solid Medium: Add 15.0g of agar to PVF Inorganic Phosphorus Liquid Medium.

[0031] 2. Using the plate coating method, the soil bacterial suspension was spread on PVF inorganic phosphorus solid medium and cultured at 30°C in an incubator until colonies grew. Colonies with phosphate-solubilizing zones were selected for purification to obtain strain PoPSB-1.

[0032] 3. Determination of the phosphate solubilization ability of strain PoPSB-1

[0033] After inoculating strain PoPSB-1 ​​onto PVF inorganic phosphorus solid medium, distinct phosphate-solubilizing zones appeared around the colonies after 1 to 2 days of cultivation. These zones gradually expanded with increasing cultivation time, stabilizing by the seventh day. The phosphate-solubilizing ability of the phosphate-solubilizing bacteria was qualitatively analyzed using the ratio of the diameter of the phosphate-solubilizing zone (D) to the diameter of a single colony (d) on the PVF inorganic phosphorus solid medium (D / d) (Table 1).

[0034] Table 1

[0035] strain name Diameter ratio (D / d) PoPSB-1 2.58±0.12

[0036] Take OD 600 Fresh POPSB-1 ​​bacterial suspension with a concentration of 1.0 was inoculated into PVF inorganic phosphorus liquid medium at an inoculum volume of 2% (v / v), with an equal volume of sterilized LB liquid medium as a control. After incubation at 30℃ for 7 days, the supernatant was collected after centrifugation, and the soluble inorganic phosphorus content in the supernatant was determined by molybdenum antimony spectrophotometry.

[0037] The results showed that the soluble inorganic phosphorus content in the fermentation supernatant of strain PoPSB-1 ​​reached 628.29 mg / L, while the soluble inorganic phosphorus content in the control group was less than 10 mg / L.

[0038] 4. 16S rDNA sequencing and physiological and biochemical identification of strain PoPSB-1.

[0039] The highly efficient phosphate-solubilizing bacterium PoPSB-1 ​​is rod-shaped and Gram-negative. On inorganic phosphorus solid medium, its colonies are characterized by being round, milky white, flat in the middle, moist on the surface, and with neat and transparent edges. Figure 1 ).

[0040] The selected strains were cultured and sent to Beijing Qingke Biotechnology Co., Ltd. for 16S rDNA sequencing. BLAST comparison showed a sequence similarity of <90% with existing strains, classifying them as novel strains. A phylogenetic tree of the 16S rRNA gene was constructed using MEGA 7.0 software, showing high homology with some bacteria in the genus *Acinetobacter*. Figure 2 ).

[0041] Physiological and biochemical identification of bacteria was performed in accordance with the "Handbook for Systematic Identification of Common Bacteria", including catalase test, oxidase test, methyl red test, VP test, indole test, starch hydrolysis, glucose oxidation, sucrose fermentation, and gelatin liquefaction.

[0042] The results are shown in Table 2. The strain PoPSB-1 ​​was positive for catalase, while all other indicators were negative.

[0043] Therefore, PoPSB-1 ​​was identified as Acinetobacter.

[0044] Table 2

[0045] index PoPSB-1 index PoPSB-1 catalase + glucose - Oxidase - sucrose - Methyl red - Starch hydrolysis - VP - Gelatin liquefaction - Indole -

[0046] Example 2

[0047] The PoPSB-1 ​​strain was inoculated into LB liquid medium to prepare a bacterial suspension, which was used for plant growth promotion experiments.

[0048] The selected phosphate-solubilizing bacterium PoPSB-1 ​​was inoculated into 40 mL of LB liquid medium and cultured overnight at 30 °C with a speed of 180 rpm. After 18 h, the bacterial culture was collected by centrifugation, and the bacterial cells were washed twice with 40 mL of sterile water. Finally, sterile water was added to adjust the bacterial concentration to 10. 8 CFU / mL available for use.

[0049] Another control strain of Bacillus subtilis WB800 (a commonly used commercial phosphorus-dissolving bacterium) was purchased, and a WB800 bacterial suspension was prepared using the same steps.

[0050] Each pot was filled with 1.0 kg of test soil, and 30 tomato seedlings of similar growth were planted in it. The treatment group received 40 mL of bacterial suspension (bacterial concentration 10). 8 cfu / mL), with 40 mL of sterile water or 40 mL of WB800 bacterial suspension (bacterial concentration 10). 8 The control group (cfu / mL) was used as the reference. Both the treatment and control groups had six replicates. Inoculation was performed using the root drenching method, and the plants were then placed in an artificial climate incubator. After 25 days of culture, samples were taken to measure tomato seedling growth indicators and soil nutrient content. The results are shown in Tables 3 and 4: Tomatoes treated with strain PoPSB-1 ​​grew better, and the soil in which the tomatoes were cultured had a higher organic matter content.

[0051] Table 3 Tomato growth indicators:

[0052] deal with Plant height (cm) Stem diameter (mm) Fresh weight of underground portion (g) Fresh weight of above-ground parts (g) water 20.18±1.82 4.96±0.45 3.84±0.23 5.34±0.20 PoPSB-1 23.67±1.81 5.24±0.52 4.22±0.17 6.82±0.17 WB800 22.72±2.12 4.91±0.44 3.91±0.20 6.30±0.28

[0053] Table 4 Soil nutrient content:

[0054]

[0055]

[0056] Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Anyone skilled in the art can make various modifications and alterations without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be determined by the claims.

Claims

1. An Acinetobacter sp. strain was deposited on December 12, 2024, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 33053.

2. A microbial inoculant, characterized in that, Contains Acinetobacter sp. as described in claim 1.

3. The microbial agent according to claim 2, characterized in that, The content of Acinetobacter bacillus as described in claim 1 in the microbial inoculant is not less than 10%. 6 cfu / mL or 10 6 cfu / g.

4. A method for reducing the content of insoluble inorganic phosphorus, characterized in that, Add the Acinetobacter sp. of claim 1 or the microbial agent of claim 2 or 3 to a system containing poorly soluble inorganic phosphorus.

5. The method according to claim 4, characterized in that, The insoluble inorganic phosphorus is converted into soluble inorganic phosphorus.

6. The method according to claim 4 or 5, characterized in that, The insoluble inorganic phosphorus includes calcium phosphate, iron phosphate, and aluminum phosphate.

7. A method for promoting plant growth, characterized in that, Add Acinetobacter sp. as described in claim 1 or the microbial agent as described in claim 2 or 3 to the plant culture system.

8. The method according to claim 7, characterized in that, The promotion of plant growth includes one or more of the following: increasing plant height, increasing plant stem girth, and increasing plant fresh weight; the plants include tomatoes, wheat, rice, corn, cotton, rapeseed, soybeans, and peas.

9. The use of Acinetobacter sp. as described in claim 1 or the microbial agent as described in claim 2 or 3 in reducing the content of insoluble inorganic phosphorus.

10. The application according to claim 9, characterized in that, The insoluble inorganic phosphorus is converted into soluble inorganic phosphorus.

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