Bacillus cereus-15 and its application in phosphorus dissolution and planting of traditional chinese medicinal materials

Abstract

The present application relates to the field of microbial technology, and particularly relates to a bacillus cereus-15 and application thereof in dissolving phosphorus and planting of traditional Chinese medicinal materials. The preservation number of the bacillus cereus-15 is GDMCC No: 67524. The bacillus cereus-15 provided in the present application can increase the content of effective phosphorus and organic carbon in rhizosphere soil of magnolia officinalis, increase the activities of ALP, LAP, NAG and BG in the rhizosphere soil, and further improve the multifunctionality of the rhizosphere soil. The bacillus cereus-15 can also increase the content of total phosphorus, inorganic phosphorus, total nitrogen, nitrate nitrogen, ammonium nitrogen and soluble protein in the roots of the magnolia officinalis seedlings, and the activities of GOGAT and GS, increase the content of chlorophyll a, chlorophyll b, carotenoid and total chlorophyll in the leaves, and increase the plant height, main root length, total fresh weight, root fresh weight, leaf fresh weight and stem fresh weight, and the multifunctionality of the rhizosphere soil is significantly positively correlated with the biomass and physiological indexes.

Description

Technical Field

[0001] This invention relates to the field of microbial technology, specifically to a Bacillus cereus-15 and its application in phosphate solubilization and the cultivation of Chinese medicinal herbs. Background Technology

[0002] *Phellodendron chinense* Schneid, a deciduous tree belonging to the genus *Phellodendron* in the family Rutaceae, is a cold- and drought-resistant Class II protected plant in China, widely distributed in Hunan, Hubei, Sichuan, and Yunnan provinces in southern China. It is a traditional and valuable woody medicinal herb in China. Its stem and root bark contain alkaloids, flavonoids, and other active compounds, with berberine, a major component, possessing anti-tumor, anti-inflammatory, antibacterial, and blood sugar- and lipid-lowering effects. It is widely used in the treatment of diabetes, enteritis, cancer, gout, and other diseases. Because *Phellodendron chinense* is generally not a dominant or leading species in a community, and its extremely high medicinal value has led to over-logging, resulting in a sharp decline in wild populations and a fragmented distribution. The growth and medicinal component content of *Phellodendron chinense* are influenced by factors such as rhizosphere growth-promoting bacteria, nutrients, and site conditions, thus affecting its yield and economic value.

[0003] Soil multifunctionality refers to the ability of a soil ecosystem to simultaneously provide and maintain multiple ecological functions, including microbial activity, nutrient cycling, and soil nutrient storage, which is fundamental to the realization of plant ecosystem functions. The soils in Hunan Province are mainly red soils, with high total phosphorus content and low available phosphorus content (≤0.01%), limiting plant growth and the synthesis of important secondary metabolites. Inoculating rhizosphere with phosphate-solubilizing bacteria is one effective way to improve soil multifunctionality and alleviate phosphorus limitation. For example, inoculating rhizosphere with phosphate-solubilizing bacteria (Pseudomonas moraviensis, Bacillus safensis, Burkholderia multivorans WS-FJ9, etc.) can significantly increase the content of available phosphorus in the soil, thereby promoting plant growth. However, research on the resource types of rhizosphere soil phosphate-solubilizing bacteria for the medicinal herb *Phellodendron amurense*, and their role in enhancing soil multifunctionality and promoting growth is relatively limited.

[0004] Therefore, the isolation and identification of phosphorus-solubilizing bacteria in the rhizosphere soil of Phellodendron amurense and their functional research on enhancing soil multifunctionality and promoting growth can provide a theoretical basis for the development of special microbial fertilizers for Phellodendron amurense and scientific planting. Summary of the Invention

[0005] To address the aforementioned technical problems, this invention provides Bacillus cereus-15 and its application in phosphorus solubilization and the cultivation of traditional Chinese medicinal herbs. Bacillus cereus-15 can increase the content of available phosphorus and organic carbon in the rhizosphere soil of *Phellodendron amurense*, and increase the activities of ALP, LAP, NAG, and BG in the rhizosphere soil, thereby enhancing the multifunctionality of the rhizosphere soil. It can also increase the content of total phosphorus, inorganic phosphorus, total nitrogen, nitrate nitrogen, ammonium nitrogen, and soluble protein in the roots of *Phellodendron amurense* seedlings, as well as the activities of GOGAT and GS. Furthermore, it increases the content of chlorophyll a, chlorophyll b, carotenoids, and total chlorophyll in the leaves, and increases plant height, taproot length, total fresh weight, root fresh weight, leaf fresh weight, and stem fresh weight. Moreover, the multifunctionality of the rhizosphere soil is significantly positively correlated with biomass and physiological indicators.

[0006] Therefore, the present invention provides the following technical solution:

[0007] In a first aspect, the present invention provides a Bacillus cereus-15 in an optional embodiment, the Bacillus cereus-15 having the accession number GDMCC No:67524.

[0008] In this invention, the accession number of Bacillus cereus-15 is GDMCC No:67524, the accession date is December 24, 2025, the accession center is Guangdong Provincial Microbial Culture Collection Center, the accession address is 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, Guangdong Academy of Sciences Institute of Microbiology, the accession classification name is Bacillus cereus BC-15, and the accession status is live.

[0009] Preferably, the nucleotide sequence of the Bacillus cereus-15 is shown in SEQ ID NO.1.

[0010] In a second aspect, the present invention provides a Bacillus cereus-15 bacterial agent in an optional embodiment, which is prepared using the aforementioned Bacillus cereus-15.

[0011] Thirdly, in an optional embodiment, the present invention provides the application of the above-mentioned Bacillus cereus-15 in increasing the available phosphorus content, ammonium nitrogen content and / or organic carbon content in the rhizosphere soil of Phellodendron amurense seedlings.

[0012] Fourthly, in an optional embodiment, the present invention provides the application of the above-mentioned Bacillus cereus-15 in improving the ALP activity, LAP activity, NAG activity and / or SMF index in the rhizosphere soil of Phellodendron amurense seedlings.

[0013] Fifthly, in optional embodiments, the present invention provides the application of the above-mentioned Bacillus cereus-15 in increasing the total phosphorus content, inorganic phosphorus content, total nitrogen content, nitrate nitrogen content, ammonium nitrogen content, GOGAT activity and / or GS activity in the roots of Phellodendron amurense seedlings.

[0014] In a sixth aspect, the present invention provides, in an optional embodiment, the application of the above-mentioned Bacillus cereus-15 in regulating the content of soluble protein in the root of Phellodendron amurense.

[0015] In a seventh aspect, the present invention provides, in optional embodiments, the application of the above-mentioned Bacillus cereus-15 in increasing the pigment content and / or photosynthetic rate in the leaves of Phellodendron amurense seedlings.

[0016] Eighthly, in an optional embodiment, the present invention provides the application of the above-mentioned Bacillus cereus-15 in increasing the plant height, root length, total fresh weight, root fresh weight, leaf fresh weight and / or stem fresh weight of Phellodendron amurense seedlings.

[0017] In a ninth aspect, the present invention provides, in an optional embodiment, the application of the above-mentioned Bacillus cereus-15 in improving the multifunctionality of rhizosphere soil of Phellodendron amurense.

[0018] The nucleotide sequence shown in SEQ ID NO.1 is as follows:

[0019]

[0020] Compared with the prior art, the present invention has one of the following beneficial effects:

[0021] 1. The Bacillus cereus-15 provided by this invention can increase the content of available phosphorus and organic carbon in the rhizosphere soil of Phellodendron amurense, increase the activity of ALP, LAP, NAG and BG in the rhizosphere soil, and thus improve the multifunctionality of the rhizosphere soil. It can also increase the content of total phosphorus, inorganic phosphorus, total nitrogen, nitrate nitrogen, ammonium nitrogen and soluble protein in the roots of Phellodendron amurense seedlings, as well as the activity of GOGAT and GS, increase the content of chlorophyll a, chlorophyll b, carotenoids and total chlorophyll in leaves, and increase plant height, taproot length, total fresh weight, root fresh weight, leaf fresh weight and stem fresh weight. Moreover, the multifunctionality of the rhizosphere soil is significantly positively correlated with biomass and physiological indicators. Attached Figure Description

[0022] Figure 1 This is a schematic diagram showing the isolation, identification, and homology analysis results of Bacillus cereus-15 in Examples 1 and 2 of the present invention;

[0023] Figure 2 This is a schematic diagram showing the effects of Bacillus cereus-15 on the physicochemical properties and multifunctionality of rhizosphere soil in Phellodendron amurense seedlings in Example 3 of the present invention.

[0024] Figure 3 This is a schematic diagram showing the effects of Bacillus cereus-15 on the physicochemical properties and enzyme activities of Phellodendron amurense seedling roots in Example 3 of the present invention;

[0025] Figure 4 This is a schematic diagram showing the effect of Bacillus cereus-15 on the pigment content and photosynthetic rate in the leaves of Phellodendron amurense seedlings in Example 3 of the present invention.

[0026] Figure 5 This is a schematic diagram showing the effect of Bacillus cereus-15 on the growth of Phellodendron amurense seedlings in Example 3 of the present invention;

[0027] Figure 6 This is a schematic diagram showing the correlation analysis results between soil multifunctionality of Bacillus cereus-15 and the biomass of Phellodendron amurense seedlings in Example 3 of the present invention. Detailed Implementation

[0028] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

[0029] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0030] Unless otherwise specified, all materials and reagents used in the following examples are commercially available.

[0031] The preparation method of R2A solid culture medium is as follows: Weigh 18.12g of R2A solid culture medium powder and 3.2g of R2A liquid culture medium powder (purchased from Haibo Biotechnology Co., Ltd.), dissolve them in distilled water, and bring the volume to 1L. Autoclave the prepared culture medium (121℃, 15min). After sterilization with 75% ethanol solution, place it in a prepared laminar flow hood to cool. When the culture medium temperature drops to 40-50℃, add 0.8mL of prepared sterile amphotericin B (final concentration 0.8mg·L⁻¹). -1 Then dispense the contents into sterile petri dishes and 50mL Erlenmeyer flasks, seal them, and set aside for later use.

[0032] The PVK medium was prepared as follows: 17.0g of PVK liquid medium powder (purchased from Haibo Biotechnology Co., Ltd.) was weighed, dissolved in distilled water, and the volume was adjusted to 1L. The prepared medium was then autoclaved (121℃, 15min) and dispensed in a clean bench for subsequent use.

[0033] Example 1

[0034] This embodiment provides a method for preparing Bacillus cereus-15, including the following steps:

[0035] S1: Collect rhizosphere soil from one-year-old Phellodendron amurense plants in the field (≤2mm from the roots), place it in a sealed bag, and bring it back to the laboratory promptly. Weigh 0.2g of fresh Phellodendron amurense rhizosphere soil sample using an analytical balance, then transfer it to an Erlenmeyer flask containing 50mL of sterile water. After sealing, place the flask in an ultrasonic cleaner and treat it at 100rpm for 3min at room temperature. Collect the bacterial suspension for later use.

[0036] S2: Take 1 mL of the isolated rhizosphere soil bacteria suspension and centrifuge at 6000 rpm for 3 min, then collect the supernatant. In a clean bench, take 50 µL of the supernatant and spread it evenly on an R2A solid plate (containing amphotericin B), seal it, and incubate it upside down in a 28℃ incubator for 18–24 h. In a clean bench, pick single colonies from the plate and transfer them to R2A liquid medium, then incubate at 28℃ and 200 rpm for 18–24 h in a shaker. After five plate-liquid culture cycles, 20 single rhizosphere soil bacteria strains of *Phellodendron amurense* were finally obtained through observation and identification under a regular microscope.

[0037] S3: The 20 obtained rhizosphere soil bacteria were subjected to streak plating and liquid culture according to the above method. Then, 500 µL of bacterial solution was taken and mixed with 500 µL of 80% glycerol solution. After labeling, the mixture was immediately placed in liquid nitrogen for quick freezing and then transferred to an ultra-low temperature freezer at -80℃ for long-term storage.

[0038] S4: Bacterial DNA from the rhizosphere soil of *Berberis thunbergii* was extracted and isolated using a bacterial genomic DNA extraction kit (purchased from Omega Bio-Tek). The 16S rRNA sequence was then amplified using universal bacterial primers 27F (5'-AGAGTTTGATCCTGGCTCAG-3') and 1492R (5'-CTACGGTACCTTGTTACGA-3'). Finally, the sequence was detected and analyzed on the Illumina NovaSeq 6000 platform. (See [link to relevant documentation]). Figure 1 A. 15 # The sequencing sequences of the strains were BLAST-aligned and homology-analyzed in the NCBI nucleic acid database. A phylogenetic tree was then constructed using MEGA 5.1 software. (See [link to documentation]). Figure 1 B. Based on the results of nucleic acid sequence analysis, 15 samples of rhizosphere soil from *Berberis thunbergii* were analyzed. # The strain was named Bacillus cereus-15 (BC-15).

[0039] Example 2

[0040] This embodiment provides a phosphorus solubility effect test of Bacillus cereus-15 prepared in Example 1. The specific steps are as follows:

[0041] (1) Preparation of potassium antimony tartrate solution: Weigh 0.5g of potassium antimony tartrate [K(SbO)C4H4O6], dissolve it in 100mL of pure water to prepare a 0.5% potassium antimony tartrate solution.

[0042] (2) Preparation of molybdenum-antimony mixed solution: Take 10g of ammonium molybdate [(NH4)6Mo7O 24 Dissolve [·4H2O] in 450mL of pure water, then slowly add 153mL of concentrated sulfuric acid while stirring. Next, add 100mL of 0.5% potassium antimony tartrate solution, and finally add water to make up to 1L. Shake well and store in a brown bottle for later use.

[0043] (3) Preparation of molybdenum-antimony anti-coloring agent: Before use (i.e., prepare on the same day), weigh 1.5g of L-ascorbic acid, dissolve it in 100mL of molybdenum-antimony mixture, mix well, and the effective period is 24h.

[0044] (4) Prepare PVK liquid culture medium containing ferric phosphate, adjusting the pH to 5, 6, 7, 8, and 9. Measure 50 mL of each pH gradient PVK liquid culture medium and place it into Erlenmeyer flasks. Finally, autoclave the flasks. Next, in a clean bench, add 100 µL of the cultured BC-15 bacterial suspension to each sterilized PVK liquid culture medium and incubate at 37°C and 160 rpm for 3 days with shaking. Then, centrifuge the mixed culture at 5000 rpm for 10 min at room temperature and collect the supernatant. Take 10 mL of the supernatant and dilute it to 50 mL with sterile distilled water. Slowly add 3 drops of 2,6-dinitrophenol indicator to the prepared solution, mix well, and then add 10% sodium hydroxide solution or 10% dilute hydrochloric acid solution until the indicator turns slightly yellow. Finally, add 10 mL of molybdenum antimony anti-chromic agent, mix well, and let stand at room temperature for 30 min. Measure the absorbance at 680 nm and calculate the available phosphorus content. See [link to relevant documentation]. Figure 1 C. Experimental results showed that the available phosphorus content in PVK liquid medium for BC-15 bacteria was 0.2910 mg·mL⁻¹ at pH 5, 6, and 9, respectively. -1 0.2364 mg·mL -1 and 0.3649 mg·mL -1 In the CK group, the available phosphorus content in the PVK liquid medium was 0.2205 mg·mL at pH 5, 6, and 9, respectively. -1 0.2339 mg·mL -1 and 0.2711 mg·mL -1 Compared with the control group, the available phosphorus content in the BC-15 group increased by 31.97%, 1.07%, and 34.60%, respectively. At pH 7 and 8, the available phosphorus content in the PVK liquid medium containing BC-15 was 0.2416 mg / mL, respectively. -1 and 0.2247 mg·mL -1 In the CK group, the available phosphorus content in the PVK liquid medium was 0.2443 mg·mL at pH 7 and 8, respectively. -1 and 0.2592 mg·mL -1 At pH 7 and 8, the content of BC-15 bacteria in the BC-15 group decreased by 1.11% and 13.31% respectively compared with the CK group at the same time, indicating that BC-15 bacteria have good phosphorus solubilization efficiency at pH 5 and 9.

[0045] Prepare PVK liquid medium containing aluminum phosphate, and adjust the pH of the medium to 5, 6, 7, 8, and 9. Subsequent operations are the same as those described for ferric phosphate. See [link to results]. Figure 1 D. Experimental results showed that the available phosphorus content in PVK liquid medium for BC-15 bacteria was 0.1496 mg·mL⁻¹ at pH 5, 6, 7, and 9, respectively. -10.1501 mg·mL -1 0.1361 mg·mL -1 and 0.1326 mg·mL -1 In the CK group, the available phosphorus content in the PVK liquid medium was 0.1128 mg·mL at pH 5, 6, 7, and 9, respectively. -1 0.1015 mg·mL -1 0.1209 mg·mL -1 and 0.1287 mg·mL -1 Compared with the control group, the available phosphorus content in the BC-15 group increased by 32.62%, 47.88%, 12.57%, and 3.03%, respectively. At pH 8, the available phosphorus content in the PVK liquid medium containing BC-15 was 0.1150 mg / mL. -1 In the CK group, at pH 8, the available phosphorus content in the PVK liquid medium was 0.1297 mg·mL⁻¹. -1 At pH 8, the content of BC-15 bacteria decreased by 11.33% compared with the CK group at the same time, indicating that BC-15 bacteria have good phosphorus solubilizing effect at pH 5 and 6.

[0046] Example 3

[0047] This embodiment provides a method for preparing a bacterial agent based on Bacillus cereus-15 prepared in Example 1, including the following steps:

[0048] (1) In the clean bench, the BC-15 bacteria stored at -80℃ will be activated on the sterilized R2A solid plate;

[0049] (2) In a clean bench, pick a single colony of BC-15 and inoculate it into an Erlenmeyer flask containing 20 mL of R2A liquid medium. Incubate at 28°C and 200 rpm for 10 h. OD of the bacterial culture is... 600 A value of 0.7 indicates BC-15 bacterial culture.

[0050] Example 4

[0051] This embodiment provides an efficacy test of Bacillus cereus-15 prepared in Example 1, specifically as follows:

[0052] (1) Sow Sichuan Phellodendron seeds (collected from Hunan Botanical Garden) in 50-cell tree seedling trays, planting 2 seeds in each cell, and place them in a greenhouse for germination for 20-30 days.

[0053] (2) When the seedlings of *Phellodendron amurense* reach a height of 10cm, select 18 seedlings with similar growth and transplant them into plastic pots (diameter × height: 19.7cm × 20.5cm) containing 3000g of field soil. Plant one seedling in each pot, for a total of 2 groups, with 9 seedlings in each group. Transfer the potted seedlings to the *Phellodendron amurense* experimental base on the third floor of the School of Life Sciences and Technology for further cultivation for 30 days, while watering and weeding are carried out regularly. When the *Phellodendron amurense* seedlings grow to the 3-5 leaf stage (about 30cm), conduct an inoculation experiment with rhizosphere BC-15 inoculant.

[0054] (3) 5 mL of sterile R2A liquid culture medium and 5 mL of BC-15 bacterial suspension were taken from the control group (CK) and experimental group (BC-15), respectively, and then diluted with sterile distilled water to 50 mL to prepare experimental inoculum. Inoculation was carried out by ring drenching, once every 7 days, with 50 mL of inoculum each time, for a total of 5 inoculations. After 35 days of cultivation, the photosynthetic parameters of the third layer of leaves (from top to bottom) of Phellodendron amurense seedlings were measured using a Li-6400 XT photosynthesis system. Subsequently, destructive sampling was performed and rhizosphere soil was collected to determine the physicochemical properties and enzyme activity of the rhizosphere soil. Plant height, leaf fresh weight, root fresh weight and total fresh weight were measured using a tape measure and analytical balance. At the same time, the third layer of leaves and roots were collected for subsequent determination of pigment and nutrient content and enzyme activity.

[0055] 1. BC-15 phosphate-solubilizing bacteria enhance the multifunctionality of rhizosphere soil in Phellodendron amurense seedlings.

[0056] After 35 days of cultivation, the total phosphorus, total nitrogen, and nitrate nitrogen contents in the rhizosphere soil of *Phellodendron amurense* seedlings in the BC-15 group were 0.70 mg·g⁻¹. -1 0.79 mg·g -1 and 0.23 mg·g -1 The total phosphorus, total nitrogen, and nitrate nitrogen contents in the rhizosphere soil of *Phellodendron amurense* seedlings in the control group were 0.87 mg·g⁻¹. -1 0.92 mg·g -1 and 0.43 mg·g -1 The levels of bacteria in the BC-15 group were reduced by 19.54%, 14.13%, and 46.51% compared to the CK group, respectively (see [reference]). Figure 2 A, Figure 2 C and Figure 2 E); The available phosphorus and organic carbon contents in the rhizosphere soil of *Phellodendron amurense* seedlings in the BC-15 group were 0.36 mg·g⁻¹. -1 and 19.15 mg·g -1 The available phosphorus and organic carbon contents in the rhizosphere soil of *Phellodendron amurense* seedlings in the control group were 0.30 mg·g⁻¹. -1 and 14.50 mg·g -1The content of BC-15 bacteria in the BC-15 group was increased by 20.00% and 32.07% respectively compared with that in the CK group (see...). Figure 2 B and Figure 2 F); The activities of ALP, LAP, NAG and BG in the rhizosphere soil of *Phellodendron amurense* seedlings in the BC-15 group were 0.10 U·g⁻¹. -1 23.53 U·g -1 6.16 U·g -1 and 6.79 U·g -1 The activities of ALP, LAP, NAG, and BG in the rhizosphere soil of *Phellodendron amurense* seedlings in the CK group were 0.09 U·g⁻¹. -1 7.43 U·g -1 4.72 U·g -1 and 3.60 U·g -1 The activity of the BC-15 bacteria group was increased by 11.11%, 216.69%, 30.51%, and 88.61% respectively compared with the CK group (see [reference]). Figure 2 H, Figure 2 I, Figure 2 J and Figure 2 The BC-15 group showed no significant difference in soluble organic carbon and ammonium nitrogen content compared to the CK group, but significantly improved the rhizosphere soil multifunctionality, indicating that BC-15 phosphate-solubilizing bacteria enhance soil multifunctionality by promoting phosphorus metabolism in the rhizosphere soil of *Phellodendron amurense* (see K). Figure 2 G, Figure 2 D and Figure 3 L).

[0057] 2. After 35 days of culture, the contents of total phosphorus, inorganic phosphorus, total nitrogen, nitrate nitrogen, ammonium nitrogen, and soluble protein in the roots of *Phellodendron amurense* seedlings in the BC-15 group were 2.06 mg·g⁻¹. -1 0.24 mg·g -1 1.97 mg·g -1 51.24 mg·g -1 6.33 mg·g -1 and 1.14 mg·g -1 The contents of total phosphorus, inorganic phosphorus, total nitrogen, nitrate nitrogen, ammonium nitrogen, and soluble protein in the roots of *Phellodendron amurense* seedlings in the control group were 1.66 mg / g. -1 0.19 mg·g -1 1.65 mg / g -1 23.41 mg / g -1 4.56 mg·g -1 and 0.97 mg·g -1 The levels of bacteria in the BC-15 group were increased by 24.10%, 26.32%, 19.39%, 118.89%, 38.82%, and 17.53% respectively compared to the CK group (see [reference]).Figure 3 A, Figure 3 B Figure 3 D、 Figure 3 E, Figure 3 F and Figure 3 In the roots of *Phellodendron amurense* seedlings (G); the activities of ACP, GOGAT, and GS in the BC-15 bacterial group were 0.17 U·g. -1 1.41 U·g -1 and 1.73 mU·g -1 The activities of ACP, GOGAT, and GS in the roots of *Phellodendron amurense* seedlings in the CK group were 0.13 U·g⁻¹. -1 1.18 U·g -1 and 1.64 mU·g -1 The activity of the BC-15 bacteria group was increased by 30.77%, 19.49%, and 5.49% compared with the CK group, respectively (see [reference]). Figure 3 C Figure 4 H and Figure 4 (I) indicates that BC-15 phosphate-solubilizing bacteria enhance the nitrogen and phosphorus metabolism in the roots of Phellodendron amurense seedlings.

[0058] 3. After 35 days of culture, the contents of chlorophyll a, chlorophyll b, carotenoids, and total chlorophyll in the leaves of *Phellodendron amurense* seedlings in the BC-15 group were 0.62 mg / g. -1 0.28 mg·g -1 0.10 mg / g -1 and 0.89 mg·g -1 The contents of chlorophyll a, chlorophyll b, carotenoids, and total chlorophyll in the leaves of *Phellodendron amurense* seedlings in the control group were 0.33 mg / g. -1 0.19 mg·g -1 0.05 mg·g -1 and 0.52 mg·g -1 The levels of bacteria in the BC-15 group were increased by 87.88%, 47.37%, 100.00%, and 71.15% respectively compared to the CK group (see [reference]). Figure 4 A- Figure 4 D); The transpiration rate, net photosynthetic rate, and stomatal conductance of *Phellodendron amurense* seedlings in group BC-15 were 1.80 mmol H₂O·m⁻¹. -2 ·s -1 3.02 µmol CO2·m -2 ·s -1 and 0.05 mol H2O·m -2 ·s -1 The transpiration rate, net photosynthetic rate, and stomatal conductance of the leaves of *Phellodendron amurense* seedlings in the CK group were 0.85 mmol H2O·m. -2 ·s -1 1.28µmol CO2·m-2 ·s -1 and 0.03 mol H2O·m -2 ·s -1 Compared with the CK group, the BC-15 group showed increases of 111.76%, 135.94%, and 66.67%, respectively, while the intercellular CO2 concentration in the leaves of *Phellodendron amurense* seedlings in the BC-15 group was 88.21 µmol CO2·mol⁻¹. -1 The CK group had a CO2 concentration of 106.96 µmol·mol⁻¹. -1 The BC-15 group showed a 17.53% decrease compared to the CK group (see [reference]). Figure 5 E- Figure 6 (H) indicates that BC-15 phosphate-solubilizing bacteria improves the photosynthetic efficiency of Phellodendron amurense seedlings by promoting pigment accumulation.

[0059] 4. After 35 days of culture, the plant height, taproot length, total fresh weight, root fresh weight, leaf fresh weight, and stem fresh weight of *Phellodendron amurense* seedlings in the BC-15 group were 18.17 cm, 27.57 cm, 10.54 g, 4.66 g, 4.79 g, and 1.09 g, respectively. In the control group (CK), these figures were 13.07 cm, 20.10 cm, 4.77 g, 1.81 g, 2.41 g, and 0.56 g, respectively. The BC-15 group showed increases of 39.02%, 37.16%, 120.96%, 157.46%, 98.76%, and 94.64% compared to the CK group, indicating that the rhizosphere BC-15 phosphate-solubilizing bacteria significantly promoted the growth of *Phellodendron amurense* seedlings (see [link to relevant documentation]). ​ ).

[0060] 5. Correlation analysis showed that the rhizosphere soil multifunctionality (SMF) of *Phellodendron amurense* was significantly positively correlated with total fresh weight, root fresh weight, taproot length, total nitrogen content in roots, and APC activity, indicating that BC-15 phosphate-solubilizing bacteria promote the growth of *Phellodendron amurense* seedlings by enhancing the rhizosphere soil multifunctionality (see [link to article]). ​ ).

[0061] Although the principles of the present invention have been described in detail above with reference to preferred embodiments, those skilled in the art should understand that the above embodiments are merely illustrative explanations of the implementation of the present invention and are not intended to limit the scope of the present invention. The details in the embodiments do not constitute a limitation on the scope of the present invention. Any obvious changes, such as equivalent transformations or simple substitutions, based on the technical solutions of the present invention without departing from the spirit and scope of the present invention fall within the protection scope of the present invention.

Claims

1. A Bacillus cereus-15, characterized in that, The preservation number of the Bacillus cereus-15 is GDMCC No:67524.

2. The Bacillus cereus-15 according to claim 1, characterized in that, The nucleotide sequence of the Bacillus cereus-15 is shown in SEQ ID NO.

1.

3. A Bacillus cereus-15 inoculant, characterized in that, It is prepared using Bacillus cereus-15 as described in claim 1 or 2.

4. The use of Bacillus cereus-15 as described in claim 1 or 2 in increasing the available phosphorus content, ammonium nitrogen content and / or organic carbon content in the rhizosphere soil of Phellodendron amurense seedlings.

5. The application of Bacillus cereus-15 as described in claim 1 or 2 in improving ALP activity, LAP activity, NAG activity and / or SMF index in the rhizosphere soil of Phellodendron amurense seedlings.

6. The use of Bacillus cereus-15 as described in claim 1 or 2 in increasing the total phosphorus content, inorganic phosphorus content, total nitrogen content, nitrate nitrogen content, ammonium nitrogen content, GOGAT activity and / or GS activity in the roots of Phellodendron amurense seedlings.

7. The use of Bacillus cereus-15 as described in claim 1 or 2 in regulating the content of soluble protein in the roots of Phellodendron amurense.

8. The use of Bacillus cereus-15 as described in claim 1 or 2 in increasing pigment content and / or photosynthetic rate in the leaves of Phellodendron amurense seedlings.

9. The use of Bacillus cereus-15 as described in claim 1 or 2 in improving the plant height, root length, total fresh weight, root fresh weight, leaf fresh weight and / or stem fresh weight of Phellodendron amurense seedlings.

10. The application of Bacillus cereus-15 as described in claim 1 or 2 in improving the multifunctionality of rhizosphere soil of Phellodendron amurense.

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