Staphylococcus simulans ZXF-202603 with multiple plant growth promoting functions and application thereof

Abstract

This invention discloses a Staphylococcus mimicking strain ZXF-202603 with multiple plant growth-promoting functions and its applications, belonging to the field of agricultural microbiology technology. The Staphylococcus mimicking strain ZXF-202603 provided by this invention not only possesses the ability to promote plant growth by producing indoleacetic acid (IAA), dissolving inorganic phosphorus, and solubilizing potassium, but also can achieve this at a relatively low cell concentration (cell concentration: 1×10⁻⁶). 6 It exhibits excellent growth performance under simulated drought stress (CFU / mL) with concentrations of 5%–10% PEG-6000, adapts to a pH range of 4.5–10.5, and a temperature range of 12℃–55℃. Furthermore, mimicking Staphylococcus aureus ZXF-202603, it significantly improves soil chemical properties and the activity of related enzymes, enhancing soil fertility. After colonization in the rhizosphere, it effectively promotes crop growth and improves crop quality and yield.

Description

Technical Field

[0001] This invention relates to a Staphylococcus mimicking strain ZXF-202603 with multiple plant growth-promoting functions and its applications, belonging to the field of agricultural microbial technology. Background Technology

[0002] In agriculture, the long-term reliance on chemical fertilizers and pesticides has, while increasing yields, led to a series of problems, including soil fertility decline, soil compaction, increased salinization, and frequent soil-borne diseases. These issues severely restrict the sustainable productivity of farmland and pose a serious threat to food security and green agricultural development. Traditional chemical and physical improvement methods, while providing short-term relief, suffer from high costs, short-lived effects, and the potential for secondary environmental risks, failing to meet the fundamental needs of green and sustainable agricultural development. Against this backdrop, microbial fertilizers centered on plant rhizosphere growth-promoting bacteria have become a key biotechnological approach for achieving green agricultural transformation due to their significant advantages in reducing fertilizer and pesticide use, improving soil, and enhancing crop resistance. Growth-promoting bacteria can directly or indirectly promote plant growth, enhance stress resistance, and improve the rhizosphere microecology through various mechanisms such as phosphorus solubilization, potassium solubilization, and the secretion of plant hormones, thereby improving plant health. However, existing reported and commercialized plant growth-promoting strains still face numerous bottlenecks in practical field applications. Most strains possess only one or two growth-promoting functions, making it difficult to address complex soil degradation and crop growth obstacles. Furthermore, these strains exhibit poor tolerance to stressful environments (such as extreme pH, temperature, drought, and salinity), resulting in low colonization and survival rates and unstable functions under harsh soil conditions, significantly diminishing their effectiveness. Therefore, the core need in the field of agricultural microbiology lies in identifying new PGPR (plant rhizosphere growth-promoting bacteria) germplasm resources that possess multiple highly efficient growth-promoting functions and excellent environmental tolerance, and achieving their low-concentration, high-efficiency field application. This will not only enrich the germplasm resources of beneficial agricultural microorganisms but is also crucial for developing microbial fertilizer products suitable for multiple scenarios, with stable functions and controllable costs, which is of great significance for promoting green agricultural development. Summary of the Invention

[0003] To address the shortcomings of the existing technology, this invention provides a strain that mimics Staphylococcus aureus ZXF-202603, which not only possesses the ability to promote plant growth by producing indoleacetic acid (IAA), dissolving inorganic phosphorus, and solubilizing potassium, but also achieves this effect at a relatively low cell concentration (cell concentration: 1×10⁻⁶). 6It exhibits high CFU / mL tolerance to drought stress simulated by 5%–10% PEG-6000, adapts to a pH range of 4.5–10.5 and a temperature range of 12℃–55℃, and maintains outstanding growth performance under these stress conditions. Furthermore, mimicking Staphylococcus aureus ZXF-202603, it significantly improves soil chemical properties and the activity of related enzymes, enhances soil fertility, and effectively promotes crop growth and improves crop quality after colonization in the rhizosphere.

[0004] The first technical solution provided by this invention is a strain that mimics Staphylococcus aureus ( Staphylococcus simulans ZXF-202603 was deposited at the Guangdong Provincial Center for Microbial Culture Collection on April 30, 2026, with accession number GDMCC No: 68208.

[0005] The second technical solution provided by the present invention is a microbial preparation containing the Staphylococcus aureus ZXF-202603 described in the first technical solution. In one embodiment, the microbial preparation contains live cells, fermentation broth, or metabolites that mimic Staphylococcus ZXF-202603. In one embodiment, the concentration of the mimic Staphylococcus aureus ZXF-202603 in the microbial preparation is not less than 1 × 10⁻⁶. 6 CFU / mL or 1×10 6 CFU / g.

[0006] The third technical solution provided by the present invention is a product containing the Staphylococcus aureus ZXF-202603 mimicking the first technical solution, or the microbial preparation described in the second technical solution. In one embodiment, the product includes microbial fertilizer, microbial soil conditioner, and plant growth promoter. In one embodiment, the amount of the mimicking Staphylococcus aureus ZXF-202603 added to the product is not less than 1×10⁻⁶. 6 CFU / mL. The present invention also provides a fourth technical solution, which is the application of the imitation Staphylococcus ZXF-202603 described in the first technical solution, or the microbial preparation described in the second technical solution, or the product described in the third technical solution in soil improvement. In one implementation, the application has at least one of the following functions: (1) It produces auxin, which promotes plant growth; the auxin is indoleacetic acid; (2) It has the ability to solubilize phosphorus, and can convert insoluble phosphorus in the soil into available phosphorus that can be absorbed by plants; (3) It has the ability to decompose potassium minerals in the soil and release readily available potassium for plant use; (4) Improve soil chemical properties and increase the content of total nitrogen, total phosphorus, total potassium, available potassium, alkaline nitrogen and organic matter in the soil; (5) Enhance soil enzyme activity and increase soil acid phosphatase (ACP) and sucrase (SC) activity.

[0007] The fifth technical solution provided by the present invention is the application of the imitation Staphylococcus ZXF-202603 described in the first technical solution, or the microbial preparation described in the second technical solution, or the product described in the third technical solution in promoting plant growth and / or improving crop quality.

[0008] In one implementation, improving crop quality specifically refers to enhancing the plant's resistance to environmental stresses; the environmental stresses include drought stress, acid-base stress, and high / low temperature stress.

[0009] The present invention provides a sixth technical solution, which is a method for promoting plant growth, wherein the method is to apply the mimicking Staphylococcus aureus ZXF-202603 described in the first technical solution, or the microbial preparation described in the second technical solution, or the product described in the third technical solution to the roots of the plant.

[0010] In one embodiment, the plant includes sweet corn.

[0011] Compared with the prior art, the beneficial effects of the present invention are as follows: The present invention provides a strain that mimics Staphylococcus aureus ZXF-202603, possessing the ability to promote plant growth by producing indoleacetic acid, solubilizing phosphorus, and releasing potassium, and exhibits outstanding stress resistance. This strain can tolerate drought stress simulated by 5%–10% PEG-6000, and can grow and reproduce normally within a pH range of 4.5–10.5 and a temperature range of 12℃–55℃, demonstrating broad environmental adaptability. The *Staphylococcus aureus* ZXF-202603 strain discovered in this invention significantly promotes the growth of sweet corn and effectively improves its quality, increasing the soluble sugar content of sweet corn by 20.85% ± 2.51%. Furthermore, this strain can exert its growth-promoting and soil-improving effects at relatively low cell concentrations. In practical applications, this bacterium increased the total nitrogen, total phosphorus, total potassium, available potassium, and organic matter in field soils by 13.35% ± 1.21%, 27.87% ± 0.41%, 8.28% ± 1.50%, 58.89% ± 5.39%, and 3.76% ± 14.05%, respectively. It also increased the activities of soil acid phosphatase and sucrase by 18.46% ± 0.54% and 653.82% ± 17.63%, respectively, effectively improving soil vitality. This provides a high-quality microbial resource for reducing fertilizer and pesticide use and promoting green and sustainable agricultural development.

[0012] Preservation of biological materials A strain that mimics Staphylococcus aureus ( Staphylococcus simulans ZXF-202603, categorized and named Staphylococcus simulans It was deposited on April 30, 2026 at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC No: 68208, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou. Attached Figure Description

[0013] Figure 1 This is the phylogenetic tree for mimicking Staphylococcus ZXF-202603 in this invention.

[0014] Figure 2 This invention is a diagram of the lysozyme zone of Staphylococcus aureus ZXF-202603.

[0015] Figure 3 This invention is based on the potassium-solubilizing circle diagram of Staphylococcus aureus ZXF-202603.

[0016] Figure 4 This is a diagram illustrating the drought tolerance of Staphylococcus aureus ZXF-202603, which is a feature of this invention.

[0017] Figure 5 This is a diagram illustrating the acid-base tolerance of Staphylococcus aureus ZXF-202603, as described in this invention.

[0018] Figure 6 This is a diagram illustrating the low-temperature tolerance of Staphylococcus aureus ZXF-202603, as described in this invention.

[0019] Figure 7 This diagram illustrates the high-temperature tolerance of Staphylococcus aureus ZXF-202603, as described in this invention.

[0020] Figure 8 This invention mimics the effect of Staphylococcus aureus ZXF-202603 fermentation broth on the agronomic traits of sweet corn (front view).

[0021] Figure 9 This diagram illustrates the effect of the Staphylococcus aureus ZXF-202603 fermentation broth on the root system of sweet corn seedlings, as described in this invention.

[0022] Figure 10 This diagram illustrates the effect of the Staphylococcus aureus ZXF-202603 fermentation broth on the leaf development during the seedling establishment period of sweet corn.

[0023] Figure 11 This is a concentration standard curve of α-naphthylamine.

[0024] Figure 12 This diagram illustrates the effect of the Staphylococcus aureus ZXF-202603 fermentation broth on the root vigor of sweet corn, as described in this invention.

[0025] Figure 13This diagram illustrates the effect of the Staphylococcus aureus ZXF-202603 fermentation broth on the quality of sweet corn fruits, as described in this invention. Detailed Implementation

[0026] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0027] Raw materials used in the examples: The culture media involved in the following examples are as follows: The formula for LB liquid culture medium is as follows (g / L): 10.0 g tryptone, 10.0 g sodium chloride, 5.0 g yeast extract, and 1000 mL distilled water.

[0028] The NBRIP medium formula is as follows (g / L): glucose 10.0 g, Ca3(PO4)2 5.0 g, MgSO4·7H2O 0.25 g, MgCl2·7H2O 0.5 g, KCl 0.2 g and (NH4)2SO4 0.15 g. For solid medium, add 18.00 g of agar, 1000 mL of deionized water, and pH 6.8–7.0.

[0029] The formula for PKO medium is as follows (g / L): 10g glucose, 0.3g NaCl, 0.3g KCl, 0.3g MgSO4·7H2O, 0.03g FeSO4·7H2O, 0.03g MnSO4·4H2O, 25g Ca3(PO4)2. For solid medium, add 18.00g agar, 1000mL deionized water, and pH 7.2.

[0030] Table 1 Experimental Reagents

[0031] The sweet corn seeds used in the following examples were kindly provided by the sweet corn research team of Foshan University.

[0032] Example 1: Determination and identification of the growth-promoting characteristics of the strain 1. Screening and identification of strains (1) Screening of growth-promoting strains: Using fermented soybean paste as a sample, after pretreatment and mixing, 0.5 mL was added to 4.5 mL of 0.9% physiological saline for serial dilution. The serially diluted solutions were plated on LB solid medium and incubated at 37℃ for 48 h. Colonies were picked and streaked onto LB plates for purification. Single colonies were then transferred to LB liquid medium for further culture, resulting in 120 isolated and purified bacterial strains, which were preserved in 50% glycerol. Subsequently, these 120 strains were screened for their growth-promoting effects, including the production of alpha-1, phosphate solubilization, potassium solubilization, nitrogen fixation, iron production, and ACC deaminase production. Figures 1-3 As shown, the optimal growth-promoting strain with the above-mentioned characteristics was finally obtained, mimicking Staphylococcus aureus ZXF-202603.

[0033] 2. Determination of strain characteristics Staphylococcus mimicry strain ZXF-202603, preserved in a laboratory freezer at -80°C, was transferred to a 4°C freezer for thawing. In a sterile environment, 200 μL of the bacterial culture was inoculated into 5 mL of LB liquid medium and cultured at 37°C and 160 r / min with shaking for 12 h to obtain the first-generation activated bacterial culture. Subsequently, 200 μL of the first-generation activated bacterial culture was transferred to a fresh 5 mL LB liquid medium and cultured under the same conditions with shaking until OD (dose elapsed). 600nm A concentration of 0.6–0.8 indicates a second-generation activated bacterial solution, used for determining the growth-promoting properties of subsequent strains, with a viable cell count reaching 1 × 10⁻⁸. 7 CFU / mL.

[0034] (1) Determination of auxin-indoleacetic acid (IAA) content: Second-generation activated bacterial suspension was inoculated at a 2% inoculum into LB liquid medium containing a final concentration of 0.5 mg / mL tryptophan, and cultured at 37℃ and 160 r / min for 48 h with shaking. After culture, the bacterial suspension was centrifuged at 3500 r / min for 10 min, and the supernatant was collected. 2 mL of the supernatant was accurately transferred and mixed with an equal volume of Salkowski colorimetric solution, thoroughly shaken, and allowed to stand in the dark for 30 min for colorimetric reaction. Subsequently, the absorbance of the reaction solution was measured at 530 nm using a UV-Vis spectrophotometer. An equal volume of blank medium without bacterial inoculation was used as a negative control. Simultaneously, a standard curve was plotted using IAA standards at gradient concentrations, and the IAA yield (mg / L) in the sample was calculated using a linear regression equation.

[0035] The supernatant of the mimicking Staphylococcus aureus ZXF-202603 fermentation strain in this invention reacts with Salkowski colorimetric solution to produce a pink color. The IAA yield in the fermentation broth was measured to be 47.38 ± 0.80 mg / L. In existing patents, the IAA yield of Staphylococcus warwickii is 16.25 ± 0.18 mg / L.

[0036] (2) Determination of inorganic phosphorus content: 2 mL of the second-generation activated bacterial suspension was inoculated into 50 mL of inorganic phosphorus liquid culture medium and cultured with shaking at 30℃ and 160 r / min for 72 h. After the culture, the fermentation broth was centrifuged at 4℃ and 5000 r / min for 10 min, and the supernatant was collected. 2.5 mL of the supernatant was accurately transferred to a 25 mL volumetric flask, 2-3 drops of 2,6-dinitrophenol indicator (0.1% w / v) were added, and 10% (w / v) Na2CO3 solution was added dropwise to adjust the pH of the system until it was just slightly yellow. Then, 5 mL of molybdenum antimony anti-coloring agent was accurately added, and the volume was adjusted to the mark with distilled water. The mixture was shaken well and allowed to stand at room temperature for 30 min. The absorbance of the reaction solution was measured at 700 nm using a UV-Vis spectrophotometer, and the soluble phosphorus content in the culture medium was calculated according to the phosphorus standard curve. The experiment was set up in three biological replicates, with an equal volume of sterile water replacing the bacterial suspension as a blank control.

[0037] The phosphorus solubility of the simulant Staphylococcus aureus ZXF-202603 was measured to be 1.64 ± 0.05 mg / L.

[0038] (3) Determination of potassium content: The second-generation activated bacterial suspension was inoculated into PKO liquid medium at an inoculation rate of 2% and cultured with shaking at 37℃ and 160 r / min for 7 days. After the culture was completed, 10 mL of fermentation broth was placed in a sterile centrifuge tube, filtered with filter paper to remove impurities, and the filtrate was collected and centrifuged at 8500 r / min for 10 min. The supernatant was collected for later use. The content of available potassium in the supernatant was determined according to the method in the National Food Safety Standard "Determination of Potassium and Sodium in Food" (GB5009.91–2017). The experiment was set up in three biological replicates, and an equal volume of sterile water was used as a blank control instead of bacterial suspension.

[0039] The potassium solubility of the simulant Staphylococcus aureus ZXF-202603 was measured to be 21.42 ± 0.06 mg / L.

[0040] Example 2: Drought tolerance test of strains Take the second-generation activated bacterial culture in the logarithmic growth phase and adjust the bacterial concentration to 1×10⁻⁶. 7 CFU / mL. Subsequently, at an inoculum volume of 2%, the cells were transferred to LB liquid medium containing different mass concentrations of PEG-6000 (5%, 10%, 20%, 30%, and 40%), and cultured at 37°C and 160 r / min for 48 h with shaking before measuring OD. 600 nm, with OD 600The nm value represents the growth and reproduction status under different drought levels. Before measurement, the instrument was zeroed using PEG-6000 culture medium of the corresponding mass concentration. The optical density (OD) of drought-resistant strains... 600nm The values ​​are used for classification: Highly sensitive: OD < 0.3; Sensitive: 0.3 ≤ OD ≤ 0.39; Tolerant: 0.4 ≤ OD ≤ 0.5; Fully tolerant: OD > 0.5.

[0041] Table 2. Results of drought tolerance of strains

[0042] Note: Different lowercase letters indicate significant differences. p <0.05), the same applies below.

[0043] The results of drought tolerance mimicking Staphylococcus ZXF-202603 are shown in Table 2 and Figure 4 As shown, it exhibits complete tolerance to 5% and 10% concentrations of PEG-6000, and high sensitivity to 20%–40% concentrations of PEG-6000.

[0044] Example 3: Acid and alkali tolerance test of the strain This embodiment measures the OD of the bacterial culture. 600nm The acid-base tolerance of *Staphylococcus aureus* ZXF-202603 was assessed. Logarithmic growth phase bacterial suspensions were inoculated at 2% in LB broth at pH 2.5, 3.5, 4.5, 6, 7, 9.5, and 10.5, respectively, and cultured at 37°C with shaking at 160 r / min. OD values ​​were measured at 12, 24, 36, 48, 60, and 72 hours. 600 nm value, with 3 replicates per group.

[0045] Table 3 Results of strain acid and alkali tolerance

[0046] The results of mimicking the acid-base tolerance of Staphylococcus aureus ZXF-202603 are shown in Table 3 and Figure 5 As shown, this bacterium is extremely sensitive to acidic environments (pH ≤ 3.5), and cannot survive or proliferate in such conditions. Its optimal growth pH range is weakly acidic to strongly alkaline (pH 4.5–10.5), with vigorous growth in the pH range of 4.5–7.0, and the optimal growth pH being 7.0. While the strain can still grow under alkaline conditions (pH 9.5–10.5), its OD... 600nm The value decreased significantly, indicating that the strongly alkaline environment has a significant inhibitory effect on its proliferation.

[0047] Example 4: Low-temperature tolerance test of the strain This embodiment measures the OD of the bacterial culture. 600The nm value was used to assess the low-temperature tolerance of *Staphylococcus aureus* ZXF-202603. Bacterial cultures in the logarithmic growth phase were placed in constant-temperature shakers at 0℃, 4℃, 8℃, 12℃, and 16℃ with shaking at 160 r / min. Ods per minute (OD) values ​​were measured at 12, 24, 36, 48, 60, 72, and 84 hours. 600 nm value, each temperature setting was repeated 3 times.

[0048] Table 4 Results of strains' low-temperature tolerance

[0049] The results of low-temperature tolerance mimicking Staphylococcus aureus ZXF-202603 are shown in Table 4 and Figure 6 As shown, the minimum growth temperature of this bacterium is 8℃; it cannot proliferate at ≤4℃; and although the strain can grow slowly at 8℃, its proliferation efficiency is extremely low (OD). 600nm ≤0.800); when the temperature rises to the range of 12~16℃, the strain grows well, and a high cell concentration (OD) can be achieved at 12℃. 600nm =2.644). In summary, this strain exhibits excellent low-temperature tolerance and proliferation ability, and its suitable low-temperature range for growth is 12~16℃.

[0050] Example 5: High-temperature tolerance test of the strain This embodiment uses the plate count method to evaluate the high-temperature tolerance of mimicking Staphylococcus aureus ZXF-202603. Logarithmic growth phase bacterial cultures were incubated at 45°C, 50°C, 55°C, 60°C, and 65°C with shaking at 160 r / min, and samples were taken at 5, 10, 20, 30, and 60 min, with each temperature treatment repeated in triplicate. The samples were serially diluted and plated onto LB agar plates, and the viable cell count was performed after incubation.

[0051] Table 5. Results of strains' high-temperature tolerance

[0052] Note: The values ​​in the table represent the number of viable bacteria after treatment at various temperatures and times, in CFU / mL.

[0053] The results of the high-temperature tolerance of mimicking Staphylococcus aureus ZXF-202603 are shown in Table 5 and Figure 7 As shown, this bacterium exhibits good heat resistance in the temperature range of 45℃ to 55℃, and can still maintain a molecular weight of 3.71×10⁻⁶ after treatment at 55℃ for 5 min. 7 The strain exhibited a high viable count of CFU / mL, and the survival curves at various temperatures tended to flatten over time. However, when the temperature reached 60℃, a significant critical change in the strain's tolerance occurred, with a sharp increase in the inactivation rate; after 60 min of treatment, the viable count dropped to 6.47 × 10⁻⁶.3 CFU / mL. At 65℃, although the measured values ​​fluctuated at individual time points, the overall survival level was extremely low, making it difficult to maintain a viable bacterial count. In summary, the maximum tolerable temperature for this strain is approximately 55℃.

[0054] Example 6: Effects of fermentation broth of bacterial strains on agronomic traits of vegetables (1) Preparation of fermentation broth: After activating the ZXF-202603 strain obtained in Example 1, the second-generation bacterial suspension was inoculated into LB liquid medium at a 2% inoculum and cultured with shaking at 37°C and 160 r / min for 48 h. After the culture, the bacterial suspension was centrifuged at 5000 r / min for 10 min, the supernatant was discarded, and the bacterial precipitate was repeatedly washed with sterile water and resuspended in sterile water. The optical density (OD) of the bacterial suspension at 600 nm was determined by turbidimetric assay. 600 nm), and adjust OD with sterile water. 600 The fermentation broth is obtained by adjusting the concentration of the culture medium (nm) to 0.8–1.0, and the viable cell count in the fermentation broth is approximately 1 × 10⁻⁶. 7 CFU / mL.

[0055] (2) Sweet corn growth experiment: The strain stored at -80℃ in glycerol was taken out, thawed at 4℃, and inoculated into 5 mL LB liquid medium at a 2% inoculum. The culture was then incubated at 37℃ and 160 r / min with shaking for 12 h to obtain the first-generation culture. This second-generation culture was then transferred to 50 mL LB liquid medium at a 2% inoculum and incubated under the same conditions for 4-6 h with shaking to obtain the second-generation culture. The second-generation culture was then inoculated into 2 L LB liquid medium at a 2% inoculum and incubated at 37℃ and 160 r / min with shaking for 12 h. The resulting culture is the fermentation broth of this strain, which can be used as a microbial fertilizer. Before application, the fermentation broth should be diluted with sterile water to a viable count of 1×10⁻⁶ cells / mL, according to experimental requirements. 6 CFU / mL, with the bacterial concentration in each treatment group remaining consistent.

[0056] The experiment included a blank control group and experimental treatment groups, with three replicates per group (one tray per replicate). The seedling substrate used in the experiment consisted of imported coconut coir, imported peat moss, carbonized rice husks, and perlite, purchased from Foshan Haomiao Wangtu Agricultural Co., Ltd. The fermentation broth of strain ZXF-202603 (with a viable count of 1×10⁻⁶) was used. 6 After thoroughly mixing the CFU / mL solution with the seedling substrate, the solution was placed in a tray, and the substrate moisture content was adjusted to 60%–70%. The treatment that was irrigated with an equal amount of sterile water served as the blank group, while the treatment that was irrigated with the fermentation broth of the bacterial strain served as the experimental group.

[0057] After sowing sweet corn seeds, the experimental group received a second root irrigation when the plants had 2-3 leaves (the first irrigation was the fermentation liquid mixed with the substrate at sowing), with 500 mL of the fermentation liquid per tray. When the plants had 3-4 leaves, the seedlings were transplanted to the field, and 4 L of the fermentation liquid was applied to each tray after transplanting. The control group used an equal amount of sterile water instead of the fermentation liquid during the same period, and the other cultivation and management conditions were the same as the experimental group.

[0058] At each key growth stage of maize, five representative plants were selected from each plot to measure plant height (from the ground to the top of the plant) and stem diameter (from the base of the stem) using a vernier caliper. All data were taken as the average of three replicate measurements, and the results are expressed as mean ± standard deviation.

[0059] The effects of the fermentation broth of the strains on the agronomic traits of sweet corn are shown in Tables 6-10. Figures 8-10 As shown, the seedling height, above-ground fresh weight, below-ground fresh weight, above-ground dry weight, below-ground dry weight, and chlorophyll content of the fourth leaf from the base of sweet corn treated with Staphylococcus aureus ZXF-202603 were increased by 12.59%±18.85%, 33.70%±58.92%, 90.84%±67.85%, 29.13%±65.75%, 33.08%±62.28%, and 22.71%±70.56% respectively compared with the control group; the number of leaves expanded during the seedling establishment period was increased by 32.79%±20 compared with the control group. 28%; before tasseling, plant height, stem diameter, and unfolded leaf length increased by 23.73%±14.09%, 9.75%±8.84%, and 20.40%±9.99% respectively compared with the control group; during tasseling, plant height, stem diameter, and ear height increased by 23.84%±11.57%, 6.12%±12.20%, and 4.73%±10.46% respectively compared with the control group; before harvest, plant height, ear height, and stem diameter increased by 8.53%±5.75%, 3.23%±7.41%, and 10.28%±13.22% respectively compared with the control group.

[0060] Table 6 Effects of fermentation broth from strains on agronomic traits of sweet corn seedlings

[0061] Note: Data in the same column marked with *, ** and *** indicate significant differences at the 0.05, 0.01 and 0.001 levels, respectively, and the same applies below.

[0062] Table 7. Effects of fermentation broth from the strain on chlorophyll in sweet corn seedlings.

[0063] Table 8 Effects of fermentation broth from strains on the seedling establishment period of sweet corn

[0064] Table 9 Effects of fermentation broth of the strain on agronomic traits of sweet corn before and during tasseling.

[0065] Table 10 Effects of fermentation broth from strains on agronomic traits of sweet corn before harvest.

[0066] Example 7: Effects of the fermentation broth of the strain on the yield and quality of sweet corn The effects of the fermentation broth ZXF-202603 on sweet corn harvest and yield are shown in Tables 11 and 12. Compared with the control group, the net weight per ear, ear diameter, number of rows per ear, number of kernels per row, number of kernels per ear, and 100-kernel weight of sweet corn treated with the mimicry of Staphylococcus aureus JQZ74 increased by 13.20%±7.01%, 5.34%±2.35%, 27.27%±2.06%, 12.15%±7.73%, 5.02%±15.28%, and 25.51%±2.22%, respectively.

[0067] The anthrone colorimetric method was used to determine the soluble sugar content in sweet corn kernels to evaluate the effect of treatment with Staphylococcus aureus ZXF-202603 fermentation broth on sweet corn quality. The effects of this strain's fermentation broth on sweet corn quality are shown in Table 13. Figure 13 As shown, the soluble sugar content of sweet corn treated with the styrogenic fermentation broth of Staphylococcus aureus ZXF-202603 was significantly increased, by 20.85% ± 2.51% compared with the control group.

[0068] Table 11 Effects of fermentation broth from strains on sweet corn yield

[0069] Table 12 Effects of fermentation broth from strains on sweet corn yield

[0070] Table 13 Effects of fermentation broth from strains on the quality of sweet corn

[0071] Example 8: Effect of strain fermentation broth on enzyme activity in maize roots Root activity of sweet maize was determined by the α-naphthylamine oxidation method (standard curve as shown). Figure 11 (As shown). The measurement results are shown in Table 14 and... Figure 12 As shown, sweet corn treated with a fermentation broth mimicking Staphylococcus aureus ZXF-202603 exhibited a significantly higher root α-naphthylamine oxidation capacity (root activity) compared to the control group, with an average growth rate of 83.51% ± 7.16%. This result indicates that applying the fermentation broth of this strain can effectively enhance the oxidative metabolic capacity of sweet corn roots and significantly improve root activity.

[0072] Table 14 Effects of fermentation broth from four strains on enzyme activity in sweet corn roots.

[0073] Example 9: Effects of bacterial strain fermentation broth on soil chemical properties (1) Soil pH determination: Weigh a soil sample and place it in a 50 mL beaker. Use distilled water (with CO2 removed) as the extraction solvent and add the solvent at a water-to-soil ratio (volume to mass) of 2.5:1. Stir for 1 min to fully disperse the soil particles, then let it stand for 30 min and complete the determination within 1 h. Immerse the indicator electrode and reference electrode (or use a pH composite electrode) in the soil suspension to form a galvanic cell. At a certain temperature, the electromotive force of the galvanic cell is a function of the pH value of the suspension. By measuring this electromotive force, the pH value of the soil can be obtained.

[0074] (2) Determination of total nitrogen content in soil (Kjeldahl method): Weigh 0.40-1.00 g of the sample and place it at the bottom of a Kjeldahl flask. Rinse the sample adhering to the flask wall with a small amount of water. Add 5.0 mL of sulfuric acid and 2 g of accelerator to the flask, shake carefully, and place a small curved-neck funnel at the mouth of the flask. First, heat at a low temperature for about 10 min, then increase the temperature and continue heating until the digestion solution and soil particles all turn grayish-white and slightly greenish, then continue digestion for 1 h. After digestion, cool and prepare for distillation. Before distillation, prepare the prepared sodium hydroxide solution, sulfuric acid standard solution, and mixed indicator. Preheat the nitrogen analyzer thoroughly and perform empty distillation to clean the pipeline until the reading is stable.

[0075] Soil total nitrogen content, expressed as nitrogen in grams per kilogram (g / kg), is calculated using the following formula: Soil total nitrogen content =

[0076] In the formula: c —The concentration of the sulfuric acid standard solution, expressed in moles per liter (mol / L). V —The volume of sulfuric acid standard solution consumed in titrating the sample, in milliliters (mL); V 0 — Volume of sulfuric acid standard solution consumed in the titration of blank, in milliliters (mL); 14.01 — Molar mass of nitrogen, in grams per mole (g / mol). m —Weigh the sample, in grams (g).

[0077] (3) Determination of total phosphorus in soil: Accurately weigh 0.2 g of sieved, air-dried sample, accurate to 0.0001 g, and carefully place it at the bottom of a nickel crucible, avoiding adhesion to the inner wall of the crucible. Add 5 drops of anhydrous ethanol to the sample to moisten it, and then evenly spread 2 g of sodium hydroxide on the sample surface. Place the nickel crucible in a high-temperature electric furnace, heat it to about 400℃, then cut off the power and keep it at that temperature for 15 min; then continue to heat it to 720℃ and keep it at that temperature for 15 min, and remove the crucible to cool naturally. Add 10 mL of hot water at about 80℃ to the cooled crucible, let it stand and cool, and after the molten material disperses, transfer the dispersion to a 50 mL volumetric flask with deionized water. Wash the crucible with 10 mL of 6 mol / L hydrochloric acid, and then wash the crucible several times with deionized water. All washing liquid is transferred to the volumetric flask mentioned above. After cooling, dilute to the mark with deionized water, mix well, and let it stand to clarify, thus obtaining the soil molten liquid. Prepare a reagent blank solution using the same method, and set aside for later use.

[0078] Pipette 0, 1, 2, 4, 6, 8, and 10 mL of 5 mg / L phosphorus standard solution into 25 mL volumetric flasks. Add an equal volume of blank solution to each flask, the same volume as the sample solution used in the subsequent colorimetric determination, and add 2–3 drops of dinitrophenol indicator. Adjust the solution to a slightly yellow color with 10% sodium carbonate solution or 5% sulfuric acid solution. Add 2.5 mL of molybdenum antimony colorimetric reagent, shake well, and dilute to the mark with deionized water. After shaking well, incubate at 15°C or above for 30 min, and measure the absorbance of each solution at a wavelength of 700 nm. Plot a standard curve with absorbance on the ordinate and phosphorus concentration (mg / L) on the abscissa.

[0079] Pipette the molten soil sample into a 25 mL volumetric flask and dilute with deionized water to approximately 3 / 5 of the total volume. Add 2–3 drops of dinitrophenol indicator to adjust the solution to a slightly yellow color. Add 2.5 mL of molybdenum-antimony colorimetric reagent, shake well, and dilute to the mark with deionized water. Incubate at room temperature (above 15°C) for 30 min. Measure the absorbance of the sample at 700 nm, using a reagent blank solution as a reference. Determine the corresponding phosphorus content from the standard curve based on the measured absorbance.

[0080] The total phosphorus content in soil, expressed as P in grams per kilogram (g / kg), is calculated using the following formula: Total phosphorus (P) content =

[0081] In the formula: C —The phosphorus content in the colorimetric solution obtained from the standard curve or regression equation, in milligrams per liter (mg / L). m —Weigh the sample, in grams (g); V 1 — The final volume of the sample after melting, in milliliters (mL); V 2—The final volume of the solution during color development, in milliliters (mL); V 3 — The volume of sample solution taken from the melt sample after it has been brought to volume, in milliliters (mL).

[0082] (4) Determination of total potassium in soil: Accurately weigh 0.2 g of sieved, air-dried soil sample, accurate to 0.0001 g, and carefully place it at the bottom of a nickel crucible, avoiding adhesion to the inner wall of the crucible. Add 5 drops of anhydrous ethanol to the sample to moisten it, and then evenly spread 2 g of sodium hydroxide on the sample surface. Place the nickel crucible in a high-temperature electric furnace, heat it to about 400℃, then cut off the power and keep it at that temperature for 15 min; then continue to heat it to 720℃ and keep it at that temperature for 15 min, then remove the crucible and let it cool naturally. Add 10 mL of hot water at about 80℃ to the cooled crucible, let it stand and cool, and after the molten material disperses, transfer the dispersion to a 50 mL volumetric flask with deionized water. Wash the crucible with 10 mL of 6 mol / L hydrochloric acid, and then wash the crucible several times with deionized water. All washing liquid is transferred to the volumetric flask mentioned above. After cooling, dilute to the mark with deionized water, mix well, and let it stand to clarify, thus obtaining the soil molten liquid. Prepare a reagent blank solution using the same method, and set aside for later use.

[0083] Accurately pipette 10 mL of a 1000 mg / L potassium standard solution into a 100 mL volumetric flask, dilute to the mark with deionized water, and mix well to obtain a 100 mg / L potassium standard solution. Dilute the above 100 mg / L potassium standard solution with deionized water to prepare a series of standard solutions with concentrations of 0 mg / L, 6 mg / L, 12 mg / L, 18 mg / L, 24 mg / L, and 30 mg / L. Measure the response values ​​of the series of standard solutions sequentially. Plot a standard curve with potassium concentration on the x-axis and response values ​​on the y-axis, and calculate the linear regression equation. Perform sample determination according to the instrument's operating instructions. Adjust the instrument zero point using a solution with zero potassium concentration in the series of standard solutions, measure the response value of the test solution, and substitute it into the linear regression equation to calculate the potassium concentration in the test solution.

[0084] Soil total potassium content, expressed as potassium (K) in grams per kilogram (g / kg), is calculated using the following formula: Soil total potassium content =

[0085] In the formula: C —The potassium content in the soil test solution obtained from the standard curve or regression equation, in milligrams per liter (mg / L). m —Weigh the sample, in grams (g); V 1 — The final volume of the sample after melting, in milliliters (mL).

[0086] (5) Determination of available potassium content in soil: Reagent preparation: Prepare a 1 mol / L neutral ammonium acetate solution (pH 7). The potassium standard series solution is prepared as follows: accurately weigh 0.19 g of potassium chloride dried at 110℃ for 2 h, dissolve and dilute with 1 mol / L neutral ammonium acetate solution to prepare a 100 mg / L potassium standard stock solution; accurately pipette 0 mL, 1.5 mL, 3 mL, 6 mL, 9 mL, 12 mL and 15 mL of the above 100 mg / L potassium standard solution into seven 50 mL volumetric flasks, dilute each with 1 mol / L neutral ammonium acetate solution to the mark, shake well, and obtain potassium standard series solutions with concentrations of 0 mg / L, 3 mg / L, 6 mg / L, 12 mg / L, 18 mg / L, 24 mg / L and 30 mg / L respectively.

[0087] Accurately weigh 2.0000 g of the sieved, air-dried soil sample (accurate to 0.0001 g) and place it in a 100 mL Erlenmeyer flask. Add 20 mL of 1 mol / L neutral ammonium acetate solution, seal tightly with a rubber stopper, shake for 30 min, and filter immediately. Collect the filtrate in a small Erlenmeyer flask. Measure the filtrate together with the aforementioned potassium standard solutions on a flame photometer. Adjust the instrument zero point using a solution where the potassium concentration in the standard series is zero. Measure the emission intensity of each standard solution and the sample filtrate. Plot a standard curve with potassium concentration on the x-axis and emission intensity on the y-axis, or directly read the potassium concentration in the sample filtrate from the instrument.

[0088] Results Calculation: The available potassium content in the soil, expressed as potassium (K) in milligrams per kilogram (mg / kg), is calculated using the following formula: Available potassium content =

[0089] In the formula: C – The concentration of potassium in the sample filtrate, expressed in milligrams per liter (mg / L). V – Volume of extractant, in milliliters (mL); m — Soil sample mass, in grams (g).

[0090] (6) Determination of soil alkaline nitrogen content: Weigh 2.000 g of sieved, air-dried soil sample and spread it evenly in the outer chamber of the diffusion dish. Add 1 g of zinc-ferrous sulfate to the outer chamber of the soil sample and spread it evenly on the soil sample. At the same time, prepare a reagent blank as a reference. Add 3 mL of 20 g / L boric acid indicator solution to the inner chamber of the diffusion dish. Apply alkaline adhesive, cover with frosted glass and rotate several times to ensure that the frosted glass is completely adhered to the edge of the diffusion dish. Then slowly rotate one side of the frosted glass to expose a narrow slit in the diffusion dish. Add 10 mL of 1.8 mol / L sodium hydroxide solution to the outer chamber of the diffusion dish through this slit and immediately cover it tightly with frosted glass. Gently rotate the diffusion dish horizontally to fully mix the solution in the outer chamber with the soil sample. Secure the frosted glass with two rubber bands crossed in a cross shape. Place the diffusion dish in a constant temperature incubator and keep it at 40℃ for 24 h. Titrate the amount of ammonia absorbed by the boric acid in the inner chamber with 0.01 mol / L hydrochloric acid standard solution. The endpoint is reached when the solution color changes from blue-purple to red. Perform a reagent blank determination simultaneously with the sample determination.

[0091] The content of available nitrogen in soil, expressed in milligrams per kilogram (mg / kg), is calculated using the following formula: Alkaline nitrogen content =

[0092] In the formula: V0—The volume of hydrochloric acid standard titration solution consumed in the blank test, in milliliters (mL); V—The volume of hydrochloric acid standard titration solution consumed in the sample determination, in milliliters (mL). C – Concentration of the hydrochloric acid standard solution, in moles per liter (mol / L). 14 — Molar mass of nitrogen, in milligrams per millimole (mg / mmol). m — Sample mass, in grams (g).

[0093] (7) Determination of soil organic matter content: Accurately weigh 0.02-0.50 g (accurate to 0.0001 g) of sieved, air-dried soil sample and place it in a hard glass test tube. Then add 10.00 mL of 0.4 mol / L potassium dichromate-sulfuric acid solution. Insert the test tubes one by one into the wire cage, and then submerge the wire cage in an oil bath at 185-190℃, so that the liquid level in the test tube is lower than the oil level. After submerging, lower the oil bath temperature to 170-180℃. Start timing when the solution in the test tube begins to boil. During this time, lift the wire cage and shake it several times in the oil bath to make the liquid temperature uniform. After 5 min ± 0.5 min, remove the wire cage from the oil bath and cool it. Transfer the digestion solution and soil residue in the test tube into a 100 mL Erlenmeyer flask. Rinse the test tube and small funnel with water to control the total volume of the solution in the Erlenmeyer flask to 50-60 mL. Add 3 drops of o-phenanthroline indicator and titrate the remaining potassium dichromate with ferrous sulfate standard solution. Perform 2-3 blank tests for each batch of analysis. Soil organic matter content, expressed in grams per kilogram (g / kg), is calculated using the following formula: Organic matter content =

[0094] In the formula: V 0 — The volume of ferrous sulfate standard solution consumed in the blank test, in milliliters (mL); V —The volume of ferrous sulfate standard solution consumed in the sample determination, in milliliters (mL). c —The concentration of the ferrous sulfate standard solution, in moles per liter (mol / L). 3 – 1 / 4 of the molar mass of a carbon atom, expressed in grams per mole (g / mol). 1.724 — the coefficient for converting organic carbon to organic matter; 1.10 — Oxidation correction factor; m —Sample mass, in grams (g); 1000 – a coefficient for converting soil content per kilogram.

[0095] The effects of the fermentation broth of the strain on the physicochemical properties of seedling soil and field soil are shown in Tables 15 and 16. Compared with the control group, the total nitrogen, total phosphorus, total potassium, and available nitrogen in the seedling soil treated with the simulant Staphylococcus aureus ZXF-202603 increased by 99.64%±6.29%, 78.43%±5.77%, 48.88%±2.39%, and 42.55%±1.03%, respectively. Compared with the control group, the total nitrogen, total phosphorus, total potassium, available potassium, and organic matter in the field soil treated with the simulant Staphylococcus aureus ZXF-202603 increased by 13.35%±1.21%, 27.87%±0.41%, 8.28%±1.50%, 58.89%±5.39%, and 3.76%±14.05%, respectively.

[0096] Table 15 Effects of bacterial strain fermentation broth on the physicochemical properties of seedling soil

[0097] Table 16 Effects of fermentation broth of the strain on the physicochemical properties of field soil

[0098] Example 10: Effects of bacterial strain fermentation broth on soil enzyme activity Plant rhizosphere soil samples treated with bacterial fermentation broth were collected using the root shaking method. After being dried at a constant temperature of 37℃ or naturally air-dried, the samples were sieved through a 30-50 mesh screen. The resulting fine soil was used to determine soil enzyme activity.

[0099] Soil acid phosphatase (ACP) and soil sucrase (SC) were measured using a kit method (Isehisa (Jiangsu Lianyungang) Biotechnology Co., Ltd.), following the instructions in the kit's manual.

[0100] The results are shown in Tables 17 and 18. After treatment with the fermentation broth of strain ZXF-202603, the activities of soil acid phosphatase (ACP) and sucrase (SC) were significantly increased compared with the control group. The ACP and SC activities in the seedling soil increased by 19.15% ± 2.25% and 88.76% ± 0.19%, respectively; while the ACP and SC activities in the field soil increased by 18.46% ± 0.54% and 653.82% ± 17.63%, respectively.

[0101] Table 17 Effects of bacterial strain fermentation broth on enzyme activity in seedling soil

[0102] Table 18 Effects of fermentation broth from strains on enzyme activity in field soil

[0103] 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. A strain that mimics Staphylococcus aureus ( Staphylococcus simulans ZXF-202603, characterized in that, It was deposited at the Guangdong Provincial Center for Microbial Culture Collection on April 30, 2026, with accession number GDMCC No: 68208.

2. A microbial preparation containing the mimicking Staphylococcus ZXF-202603 as described in claim 1.

3. The microbial preparation according to claim 2, characterized in that, In the microbial preparation, the concentration of the mimic Staphylococcus aureus ZXF-202603 is not less than 1×10⁻⁶. 6 CFU / mL or 1×10 6 CFU / g.

4. A product characterized in that, The product contains the mimicry Staphylococcus ZXF-202603 as described in claim 1, or the microbial preparation as described in any one of claims 2 to 3.

5. The product according to claim 4, characterized in that, The products include microbial fertilizers, microbial soil conditioners, and plant growth promoters.

6. The application of the Staphylococcus aureus ZXF-202603 as described in claim 1, or the microbial preparation as described in claims 2-3, or the product as described in any one of claims 4-5, in soil improvement.

7. The application according to claim 6, characterized in that, The application has at least one of the following functions: (1) It produces auxin, which promotes plant growth; the auxin is indoleacetic acid; (2) It has the ability to dissolve phosphorus, converting insoluble phosphorus in the soil into available phosphorus that can be absorbed by plants; (3) It has the ability to decompose potassium minerals in the soil and release readily available potassium for plant use; (4) Improve soil chemical properties and increase the content of total nitrogen, total phosphorus, total potassium, available phosphorus, available potassium, alkaline nitrogen and organic matter in the soil; (5) Enhance soil enzyme activity and increase soil phosphatase and sucrase activity.

8. The application of the Staphylococcus aureus ZXF-202603 as described in claim 1, or the microbial preparation as described in claims 2-3, or the product as described in any one of claims 4-5 in promoting plant growth and / or improving crop quality.

9. The application according to claim 8, characterized in that, Improving crop quality specifically refers to enhancing the plant's resistance to stressful environments; these stressful environments include drought stress, acid-base stress, and high / low temperature stress.

10. A method for promoting plant growth, characterized in that, The method is to apply the Staphylococcus aureus ZXF-202603 as described in claim 1, or the microbial preparation as described in claims 2-3, or the product as described in any one of claims 4-5 to the roots of a plant.

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