Phosphorus-solubilizing strain XY04 and application thereof in activation of soil selenium element

Abstract

The invention belongs to the technical field of microorganisms, and particularly relates to a phosphorus-solubilizing strain XY04 and application thereof in activation of soil selenium. The invention discloses a strain XY04, the taxonomic name of the strain XY04 is Lysinibacillus capsici, the strain XY04 is preserved in Guangdong Microbial Culture Collection Center on July 01, 2025, and the preservation number of the strain XY04 is GDMCC No: 66618. The strain XY04 disclosed by the invention is subjected to morphological observation and molecular biological identification, and it is determined that the strain XY04 is lysinibacillus capsici. In addition, tests find that the strain XY04 has the effects of dissolving phosphorus, resisting selenium and cadmium and promoting growth, can improve the content of available phosphorus and effective selenium in soil and reduce the content of effective cadmium, and is beneficial to improving the utilization rate of the available phosphorus in the soil and realizing selenium enrichment and cadmium reduction of crops; meanwhile, the growth of crops can be promoted, the selenium and cadmium stress resistance of the crops is improved, and the method has important significance for producing selenium-rich green products.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology, specifically relating to a phosphate-solubilizing strain XY04 and its use in activating soil selenium. Background Technology

[0002] Phosphorus (P) is one of the three essential nutrients for plants and a crucial component of organic compounds in crops, playing a vital role in plant growth and development. To promote crop growth and increase yield, growers apply soluble phosphate fertilizers to the soil. However, in practice, most of the phosphorus in these fertilizers reacts with calcium in the soil. 2+ Fe 2 + Fe 3+ Al 3+ Phosphorus ions combine with phosphorus to form insoluble phosphates, resulting in low utilization rates of this phosphorus fertilizer by crops. This not only wastes phosphorus resources but also impacts the surrounding environment. Therefore, improving soil phosphorus utilization is of great significance for achieving sustainable agricultural development.

[0003] Soil microbial phosphorus solubilization technology has attracted widespread attention due to its environmental friendliness and sustainability. Several genera of microorganisms have been reported to possess phosphorus-solerating capabilities. Phosphate-solving microorganisms (PSMs), also known as phosphate-solubilizing bacteria, are a class of microorganisms that can convert insoluble phosphorus, which is difficult for plants to absorb, into soluble phosphates, thereby improving the bioavailability of phosphorus in the soil and contributing to increased soil phosphorus utilization efficiency. Therefore, screening for phosphorus-solubilizing strains is of great significance.

[0004] In addition, selenium (Se) is one of the essential trace elements for human health, possessing antioxidant, anti-aging, and anti-cancer properties. 72% of China's regions are selenium-deficient, with 30% being severely deficient, making it a globally recognized selenium-deficient country. Insufficient selenium intake leads to selenium deficiency, weakened immunity, and can trigger various diseases such as diabetes, Keshan disease, arthritis, cardiovascular disease, and even cancer. Rice ( Oryza sativa Selenium (L.) is a staple food crop for Chinese residents. Consuming selenium-enriched rice to supplement selenium has advantages such as safety and economy, and is considered the best way to increase the selenium intake of the population.

[0005] Applying inorganic selenium to the soil or spraying it with inorganic selenium solution on the leaves are common methods for increasing the selenium content of crops. However, applying inorganic selenium to the soil carries the risk of excessive selenium content in the soil, and most of the selenium is adsorbed and deposited by soil particles, easily leaching out through soil runoff, polluting groundwater, and causing potential environmental risks and waste of selenium resources. In addition, crops have poor absorption of foliar-sprayed inorganic selenium solutions, and the utilization rate of selenium transferred to crop fruits is low. To achieve the ideal selenium enrichment effect, it is often necessary to increase the amount of inorganic selenium solution sprayed, which exacerbates the aforementioned risks.

[0006] Secondly, the heavy metal cadmium can cause various human diseases such as osteoarthritis, headaches, hypertension, kidney disease, lung cancer, and prostate cancer. Rice is a staple food crop consumed daily by urban and rural residents. Rice is highly tolerant of cadmium but also readily absorbs and accumulates it. Cadmium-contaminated rice enters the human body through daily consumption, harming human health. Chemical remediation methods (such as adding chemical passivating agents to the soil) can quickly reduce the content of available cadmium in the soil, but this also reduces the activity of selenium in the soil, producing inorganic precipitates, leading to soil compaction, damaging soil structure, degrading the original functions of the soil, and causing secondary pollution. Therefore, more and more researchers are turning their attention to microbial bioremediation.

[0007] The inventors screened several indigenous microorganisms, including strain XY04, from soils in Guangxi Zhuang Autonomous Region that are normally used for crop cultivation. These microorganisms have the ability to activate phosphorus, are resistant to selenium, activate selenium, produce IAA, and inhibit cadmium activity. They help improve the utilization rate of soil phosphorus, increase the content of available selenium in the soil, and inhibit the activity of heavy metal cadmium in the soil, thereby helping to achieve selenium enrichment and cadmium reduction in crops.

[0008] There are currently no reports on the phosphorus solubilization and activation of soil selenium by strain XY04. Summary of the Invention

[0009] The purpose of this invention is to provide a phosphate-solubilizing strain XY04 and its use in activating soil selenium.

[0010] To achieve the above objectives, the present invention provides the following technical solution: The first objective of this invention is to provide a strain XY04, characterized in that the taxonomic name of strain XY04 is *Bacillus capsicisae*. Lysinibacillus capsici It was deposited at the Guangdong Provincial Center for Microbial Culture Collection on July 1, 2025, with accession number GDMCC No: 66618.

[0011] A second object of the present invention is to provide the use of the strain XY04 in phosphorus solubilization.

[0012] A third object of the present invention is to provide the use of the strain XY04 in increasing the available phosphorus content in soil.

[0013] The fourth object of the present invention is the use of the strain XY04 in activating soil selenium and thus increasing the effective selenium content.

[0014] Compared with the prior art, the present invention has the following beneficial effects: Based on morphological observation and molecular biological identification, strain XY04 of this invention was identified as *Bacillus capsicisae*. Lysinibacillus capsici Furthermore, experiments have revealed that strain XY04 of this invention possesses phosphorus-solubilizing, selenium- and cadmium-tolerant, and growth-promoting effects. It can increase the available phosphorus and selenium content in the soil, reduce the available cadmium content, and help improve the utilization rate of available phosphorus in the soil, as well as achieve selenium enrichment and cadmium reduction in crops. At the same time, it can promote crop growth and improve their tolerance to selenium and cadmium stress, which is of great significance for the production of selenium-enriched green products.

[0015] Preservation Information Strain XY04, taxonomically named *Bacillus capsicisae* (… Lysinibacillus capsici It was deposited on July 1, 2025 at the Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangdong Province, Institute of Microbiology, Guangdong Academy of Sciences, with accession number GDMCC No: 66618.

[0016] Strain XY10, taxonomically named *Bacillus longiformis* (Lysine Bacillus) Lysinibacillus macroides It was deposited on July 1, 2025 at the Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangdong Province, Institute of Microbiology, Guangdong Academy of Sciences, with accession number GDMCC No: 66619.

[0017] strain XY11, taxonomic name Lysinibacillus sp . It was deposited on July 1, 2025 at the Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangdong Province, Institute of Microbiology, Guangdong Academy of Sciences, with accession number GDMCC No: 66620. Attached Figure Description

[0018] Figure 1 This is a diagram showing the colony morphology of the bacterial strain. Figure 2 Phylogenetic tree of strain XY04; Figure 3 Phylogenetic tree of strain XY10; Figure 4 Phylogenetic tree of strain XY11; Figure 5 To assess the strain's tolerance to selenium and cadmium; Figure 6 The effect of different pH values ​​on the growth of the strain; Figure 7 The effect of different culture times on the phosphorus-solubilizing ability of the strain; Figure 8 The effect of different inoculum amounts on the phosphate-solubilizing ability of the strain; Figure 9 The effect of different temperatures on the phosphorus-solubilizing ability of the strains; Figure 10 The effect of different pH values ​​on the phosphate-solubilizing ability of the strain; Figure 11 The effect of different initial cadmium concentrations on the phosphorus-solubilizing ability of the strains; Figure 12 The effect of different initial selenium concentrations on the phosphorus-solubilizing ability of the strains; Figure 13 The effect of the strain on the available phosphorus content in the soil; Figure 14 The effect of the strain on the available cadmium content in the soil; Figure 15 The effect of the strain on the available selenium content in the soil. Detailed Implementation

[0019] The technical solution of this invention patent will be clearly and completely described below. Obviously, the described embodiments are only some, not all, of the embodiments of this invention. Based on the embodiments of this invention, all other embodiments obtained by those skilled in the art without inventive effort are within the scope of protection of this invention.

[0020] 1. Materials and Methods 1.1 Screening soil for bacteria Soil samples were collected from Nanning, Jingxi, Liuzhou and Guiping cities in Guangxi Zhuang Autonomous Region.

[0021] 1.2 Culture medium formulation LB liquid medium: 10g tryptone, 5g yeast extract, 10g sodium chloride, add 1000mL deionized water, adjust pH to 7.0, boil to dissolve, and then autoclave at 121℃ for 20min.

[0022] LB solid medium: 10g tryptone, 5g yeast extract, 10g sodium chloride, 15g agar powder, add 1000mL deionized water, adjust pH to 7.0, boil to dissolve, and then autoclave at 121℃ for 20min.

[0023] Isolation medium: 10g tryptone, 5g yeast extract, 10g sodium chloride, 15g agar powder, add 1000mL deionized water, adjust pH to 7.0, boil to dissolve, and then autoclave at 121℃ for 20min. After sterilization, while still hot, add cadmium chloride (CdCl2) and sodium selenite (Na2SeO3) solution to neutralize the Cd in the medium. 2+ and Se 4+ The concentration of each medium was 1 mg / L (relative to the unsolidified medium). After mixing, the medium was poured into sterile petri dishes to make plates.

[0024] PVK medium: 15g glucose, 5.0g Ca3(PO4)2, 0.2g NaCl, 0.5g (NH4)2SO4, 0.1g MgSO4, pH 6.8-7.0, dissolved in 1000mL deionized water, autoclaved at 121℃ for 20min.

[0025] NBRIP medium (phosphate-solubilizing medium): (NH4)2SO4 0.5g, NaCl 0.3g, MgSO4·7H2O 0.3g, glucose 10.0g, MnSO4·2H2O 0.03g, Ca3(PO4)2 5.0g, FeSO4·7H2O 0.03g, KCl 0.3g, pH 7.2-7.4, dissolved in 1000mL deionized water, autoclaved at 121℃ for 20min.

[0026] Nitrogenous medium (YN): 10.0g sucrose, 1.0g ammonium sulfate, 2.0g dipotassium hydrogen phosphate, 0.5g magnesium sulfate heptahydrate, 0.1g sodium chloride, 0.5g yeast extract, and 0.5g calcium carbonate were dissolved in distilled water in sequence, and the volume was adjusted to 1000mL. The pH was adjusted to 7.2, and the medium was autoclaved at 121℃ for 20min.

[0027] 1.3 Strain screening Weigh 10g of soil sample and add it to an Erlenmeyer flask containing 90mL of sterile water. Shake the flask at room temperature for 1 hour to disperse the microorganisms evenly. After standing for 30 minutes, take 5mL of the supernatant and add it to 150mL of bacterial liquid culture medium. Incubate at 30℃ and 120r / min for 5 days and observe the growth of the microorganisms.

[0028] 1.4 Strain purification Dilute the bacterial suspension cultured for 5 days (1.3): First, take 1 mL of the bacterial suspension and add it to 9 mL of sterile water; this makes a 10⁻⁶ solution. -1 After mixing the bacterial suspension, take 1 mL of 10 -1 Add the bacterial culture to 9 mL of sterile water; this is 10. -2 Bacterial solution, and so on, yielding 10 -3 10 -4 10-5 Diluted bacterial suspensions; take 10... -3 10 -4 10 -5 Spread 0.5 mL of diluted bacterial solution evenly on the isolation medium and incubate at 30°C inverted position for 48 h.

[0029] Visually inspect and select colonies of different shapes and colors, then continue dilution using the above method. Further isolate and purify using the spread plate method until no abnormal colonies remain on the plate. Inoculate the purified strain onto LB agar slants and store at 4°C.

[0030] 1.5 Strain identification and growth characteristics 1.5.1 Colony morphology characteristics After sterilizing LB solid medium, pour it into sterile petri dishes to prepare plates. Use an inoculation loop to pick up one loopful of the preserved strain and place it in 9 mL of sterile water. Dilute to 10⁻⁶ using the steps in 1.4. -3 10 -4 10 -5 Each of the three dilutions of bacterial culture was taken and spread onto LB solid medium. After incubation at 30°C for 48 hours, the size, shape and color of the colonies were observed.

[0031] 1.5.2 Identification of 16S rRNA in the strain DNA was extracted from the bacterial strain using the Ezup column-based bacterial genomic DNA extraction kit (B518255). Universal primers 27F and 1492R were selected for PCR amplification. The primer sequences of the PCR amplification products are shown in Table 1, and the PCR reaction system is shown in Table 2.

[0032] Table 1 16S rRNA primers

[0033] Table 2 PCR reaction system

[0034] The PCR program was as follows: 95℃, 5 min; 94℃, 30 s; 57℃, 30 s; 72℃, 1 min, 30 cycles; 72℃, 10 min.

[0035] The purified PCR products were sequenced, and the sequencing results are shown in SEQ ID No. 1-SEQ ID No. 3 of the sequence listing. The obtained sequences were compared with nucleic acid data in GenBank using the Blast program for homology analysis, and a phylogenetic tree was constructed.

[0036] 1.5.3 Colony physiological and biochemical characteristics Referring to "Microbiology Experiments", Gram staining, starch hydrolysis test, indole reaction, methyl red test, glucose fermentation test, VP reaction, salt tolerance test and ammonia production test were performed on the target colony.

[0037] 1.6 Experiment on the strain's tolerance to selenium-cadmium One loopful of the isolated and purified bacterial strain, preserved at 4℃, was inoculated onto an LB agar slant and activated at 35℃ for 48 hours. The activated strain was then inoculated onto LB agar plates using the streak plate method and incubated upside down at 35℃ for 48 hours. Colony growth was observed and photographed.

[0038] To investigate the tolerance of each strain to cadmium selenide, LB solid medium was sterilized, and cadmium chloride (CdCl2) and sodium selenite (Na2SeO3) solution was added before the medium solidified, so that the Cd concentration in the medium increased. 2+ Se 4+ Cd 2+ +Se 4+ Concentrations of 1 mg / L, 5 mg / L, and 1 mg / L + 5 mg / L were prepared, mixed thoroughly, and poured onto plates. Using the streak plating method, the strains grown on the aforementioned LB solid medium plates (without added selenium and cadmium) were inoculated onto these LB solid medium plates (first round with added selenium and cadmium). The plates were incubated upside down at 35°C for 48 hours, and colony growth was observed and photographed. This process was repeated for a second round of LB solid medium plates with added selenium and cadmium (Cd). 2+ Se 4+ Cd 2+ +Se 4+ Concentrations of 2 mg / L, 10 mg / L, and 2 mg / L + 10 mg / L were added, and the third round of LB solid culture medium plates with added selenium and cadmium (Cd) were used. 2+ Se 4+ Cd 2+ +Se 4+ Concentrations of 5 mg / L, 20 mg / L, and 5 mg / L + 20 mg / L were added in the fourth round of LB solid culture medium plates with added selenium and cadmium (Cd). 2+ Se 4+ Cd 2+ +Se 4+ Concentrations of 10 mg / L, 50 mg / L, and 10 mg / L + 50 mg / L were added in the fifth round of LB solid culture medium plates with added selenium and cadmium (Cd). 2+ Se 4+ Cd 2+ +Se 4+ Concentrations of 20 mg / L, 100 mg / L, and 20 mg / L + 100 mg / L were added, until the ninth round of adding cadmium selenide (Cd) LB solid medium plates. 2+ Se 4+ Cd2+ +Se 4+ The concentrations were 300 mg / L, 2000 mg / L, and 300 mg / L + 2000 mg / L, respectively.

[0039] 1.7 Effects of different environmental conditions on strain growth and phosphate solubility 1.7.1 Effect of different pH values ​​on the growth of the strain The bacterial strain was inoculated onto LB solid medium and activated at 30°C for 24 hours. Then, under aseptic conditions, the activated strain was inoculated into LB liquid medium and incubated at 30°C and 120 rpm for 24 hours. Finally, the culture solution was diluted with sterile water to the required OD for the experiment. 600 Bacterial suspension.

[0040] bacterial suspension (OD) 600 The culture medium was inoculated with a pH of 1.0 at a rate of 5% (v / v) into LB liquid medium and incubated at 33°C with constant temperature shaking at 160 rpm. An uninoculated culture medium served as a blank control, and each treatment was repeated three times. The pH of the LB liquid medium was adjusted using 1 mol / L HCl or 1 mol / L NaOH solution to establish six pH gradients: 5.0, 6.0, 7.0, 8.0, 9.0, and 10.0. Samples were taken after 36 hours of incubation, and the absorbance (OD) of the bacterial culture at 600 nm was measured using a spectrophotometer. 600 value.

[0041] 1.7.2 Effects of different environmental conditions on phosphorus solubility 1.7.2.1 Effect of different culture times on phosphorus solubility bacterial suspension (OD) 600 The culture medium (pH 6.0) was inoculated into 100 mL of NBRIP medium (pH 6.0), with an inoculum size of 10%, and cultured at 31℃ with shaking at 180 rpm. Culture times were set at 12, 24, 36, 48, 60, 72, 84, 96, 108, and 120 h. At each time point, the cultured bacterial solution was shaken well, 10 mL was transferred to a centrifuge tube, centrifuged at 400 rpm for 10 min, and 5 mL of the supernatant was transferred to a 50 mL volumetric flask. The phosphorus content in the solution was determined using the molybdenum antimony colorimetric method.

[0042] 1.7.2.2 Effect of different inoculum amounts on phosphorus solubility bacterial suspension (OD) 600The culture medium (pH 6.0) was inoculated into 100 mL of NBRIP medium with an inoculum concentration (v / v) of 0.5%, 1%, 2%, 5%, and 10%. The culture was incubated at 31°C with shaking at 180 rpm for 96 h. After incubation, 10 mL of the culture was transferred to a centrifuge tube and centrifuged at 400 rpm for 10 min. 5 mL of the supernatant was transferred to a 50 mL volumetric flask, and the phosphorus content was determined using the molybdenum antimony colorimetric method.

[0043] 1.7.2.3 Effect of different culture temperatures on phosphorus solubility bacterial suspension (OD) 600 The culture medium (pH 6.0) was inoculated into 100 mL of NBRIP medium (pH 6.0), with an inoculum size of 10%. Five temperature gradients were set at 28℃, 31℃, 34℃, 37℃, and 40℃, and the culture was incubated with shaking at 180 rpm for 96 h. After incubation, the culture was shaken well, and 10 mL was transferred to a centrifuge tube and centrifuged at 4000 rpm for 10 min. 5 mL of the supernatant was transferred to a 50 mL volumetric flask, and the phosphorus content in the solution was determined using the molybdenum antimony colorimetric method.

[0044] 1.7.2.4 Effect of different pH values ​​on phosphorus solubility bacterial suspension (OD) 600 The culture medium (value 0.8) was inoculated into 100 mL of NBRIP medium (inoculum size 10%), and cultured at 31℃ with shaking at 180 rpm for 96 h. After incubation, the culture was shaken well, and 10 mL was transferred to a centrifuge tube and centrifuged at 4000 rpm for 10 min. 5 mL of the supernatant was transferred to a 50 mL volumetric flask, and the phosphorus content was determined using the molybdenum antimony colorimetric method. The NBRIP medium was adjusted to five pH gradients (5.0, 6.0, 7.0, 8.0, and 9.0) using 1 mol / L HCl or 1 mol / L NaOH solution.

[0045] 1.7.2.5 Effects of different initial concentrations of selenium and cadmium stress on phosphorus solubility Add cadmium chloride (CdCl2) and sodium selenite (Na2SeO3) solutions to 100 mL of NBRIP medium (pH 5.0) to establish five concentration gradients of selenium and cadmium: CdCl2 + Na2SeO3 + Na2SeO3 + Na2SeO3. 2+ Se 4+ The concentrations were 1 mg / L, 3 mg / L, 5 mg / L, 7 mg / L, and 9 mg / L, respectively. Then, the bacterial suspension (OD200) was... 600The culture medium (value 0.8) was inoculated into NBRIP medium and cultured at 31℃ with shaking at 180 rpm for 96 h. After the culture was well mixed, 10 mL of the culture was placed into a centrifuge tube and centrifuged at 4000 rpm for 10 min. 5 mL of the supernatant was transferred to a 50 mL volumetric flask, and the phosphorus content in the solution was determined by the molybdenum antimony colorimetric method.

[0046] 1.8 Soil Sample Pot Experiment To evaluate the regulatory effect of the strain on the available phosphorus, cadmium and selenium content in rice soil.

[0047] Select a round plastic basin 5cm high and 5cm in diameter, and fill each basin with 300g of soil sample (soil sample collected from paddy fields in Shilong Town, Guiping City) that has been air-dried through a 2mm nylon sieve. Completely saturate the soil and place it in a dry environment for 2 weeks to equilibrate, replenishing water periodically until the moisture content is maintained at 60% of the field's maximum water holding capacity.

[0048] Original bacterial culture of the strain (OD) 600 The bacterial solution was applied at two concentration gradients (5% and 10% v / m, i.e., 5 mL or 10 mL of bacterial solution added to 100 g of soil sample) to plastic pots containing 100 g of paddy soil. Six treatments were set up: XY10-5%, XY10-10%, XY11-5%, XY11-10%, XY04-5%, and XY04-10% (the "-" indicates the strain number, and the "-" indicates the inoculum size). The original soil without bacterial solution served as a blank control. Each treatment was replicated in triplicate. All plastic pots were placed in an artificial climate chamber with a temperature of 31℃ and a relative humidity of 60%, simulating the daylight hours of paddy soil (14 h light / 10 h dark) to ensure uniform and sufficient light exposure for each pot.

[0049] On days 7, 14, and 21, 50g of mixed soil samples were collected from each plastic basin using a diagonal sampling method. After natural air drying, grinding, and passing through a 2mm nylon sieve, the available phosphorus, available selenium, and available cadmium contents in the soil were determined.

[0050] The determination of available phosphorus in soil was performed using the sodium bicarbonate extraction-molybdenum antimony spectrophotometric method (reference standard: Determination of Available Phosphorus in Soil - Sodium Bicarbonate Extraction-Molybdenum Antimony Spectrophotometric Method HJ704-2014): 2.5000 g of air-dried soil sample was taken, and 50 mL of 0.5 mol / L NaHCO3 extraction solution (pH 8.5) was added. The mixture was shaken at 25℃ and 180 r / min for 30 min. The sample was filtered through phosphorus-free filter paper, and 10 mL of the filtrate was transferred to a 50 mL colorimetric tube. Water was added to a final volume of 15-20 mL, followed by one drop of 2,4-dinitrophenol indicator. Sulfuric acid solution was then added dropwise until the solution was nearly colorless. 0.75 mL of ascorbic acid solution was added, and the mixture was stirred until dissolved. After 30 seconds, 5 mL of molybdate solution was added, and the volume was adjusted to 50 mL with water. The mixture was stirred until dissolved. The volumetric flask was then placed at room temperature for 30 min. Colorimetric measurements were performed using a 10mm cuvette at a wavelength of 880nm under ambient conditions above 20°C, with deionized water as a reference. The absorbance was measured, and the available phosphorus content in the soil was calculated based on a pre-established standard curve.

[0051] The available selenium and available cadmium contents in the soil were determined using DTPA extraction-atomic fluorescence spectrometry and DTPA extraction-inductively coupled plasma atomic emission spectrometry. 10.00 g of air-dried soil sample (passed through a 2.00 mm sieve) was weighed and placed in a 100 mL plastic bottle. 20.0 mL of diethylenetriaminepentaacetic acid-calcium chloride-triethanolamine (DTPA-CaCl2-TEA) buffer solution was accurately added. The bottle was capped, shaken well, and oscillated at 180 r / min for 2 h at a constant temperature of 20 ± 2 °C. After settling and standing for 30 min, the extract was slowly filtered through medium-speed filter paper and then gravity filtered. The extract was measured within 48 h. Available cadmium content was determined using inductively coupled plasma atomic emission spectrometry, and available selenium content was determined using atomic fluorescence spectrometry.

[0052] 1.9 Hydroponic Experiment To investigate the promoting effect of bacterial solution on rice seed germination and its alleviating effect on selenium and cadmium stress, a hydroponic experiment was conducted.

[0053] S1. Soak rice seeds in 1% CuSO4 solution for 30 min, then soak them in 0.5% ZnSO4 solution for 15 min. After disinfection, rinse them repeatedly with sterile water 3-5 times. The volume ratio of rice seeds to sterile water during rinsing is 1:20. S2. In a sterile operating room, inoculate the bacterial strain at a 5% (v / v) inoculation rate into LB liquid medium (pH 8.0), and incubate at 34°C for 2 days. Afterward, remove the cultured bacterial solution, centrifuge at 4000 rpm for 10 minutes, collect the bacterial cells, and adjust the OD ratio with sterile water. 600 Adjust the pH to 0.8 to obtain a bacterial suspension for later use; S3. Place sterile filter paper in a sterilized petri dish (9cm in diameter). Divide the rice seeds treated in S1 into two groups: one group is soaked in the bacterial suspension for 2 hours, and the other group is not soaked in the bacterial suspension. Then, spread 50 seeds per dish evenly on the filter paper and set the following treatments for the rice seeds: Treatment 1: Do not soak the bacterial suspension, add 5 mL of sterile water to moisten; Treatment 2: Soak in bacterial suspension with a selenium-cadmium concentration of 0, then add 5 mL of sterile water to moisten; Treatment 3: Without soaking the bacterial suspension, add 5 mL of Se at concentrations of 5 mg / L, 10 mg / L, and 15 mg / L, respectively. 4+ Solution and Cd 2+ Solution (prepared with cadmium chloride and sodium selenite solution, concentration expressed as Se) 4+ and Cd 2+ express); Treatment 4: Soak the bacterial suspension and add 5 mL of Se at concentrations of 5 mg / L, 10 mg / L, and 15 mg / L, respectively. 4+ Solution and Cd 2 + Solution (prepared with cadmium chloride and sodium selenite solution, concentration expressed as Se) 4+ and Cd 2+ express); Each of the above treatments was repeated 3 times. The seeds were cultured for 7 days under the conditions of 26±1℃, 70% relative humidity, 12h photoperiod / 12h dark period, and seed germination was observed every 24 hours. Seedling length and root length were measured on the 7th day.

[0054] The methods for measuring each indicator are as follows: (1) Germination potential (GE) and germination rate (GR) refer to the percentage of all normally germinated seeds out of the total number of tested seeds at the beginning and end of the germination stages. Germination potential was measured on day 3 after sowing, and germination rate was measured on day 7. The formulas for calculating germination potential and germination rate at this time are: Germination potential (%) = (Number of seeds germinated within 3 days / Total number of seeds tested) × 100; Germination rate (%) = (Number of seeds germinated within 7 days / Total number of seeds tested) × 100; (2) Germination index is a seed vigor indicator. A high germination index means high vigor.

[0055] Germination Index (GI) = ∑Gt / Dt, where Gt represents the number of seeds that germinate on day t, and Dt represents the corresponding number of germination days; (3) Vitality Index (VI) = GI × S, where S represents the average root length; (4) Seedling length and root length: Take 10 seedlings with uniform growth in each dish and measure their seedling length and root length. Take the average value of the results.

[0056] 2. Results and Analysis 2.1 Isolation and purification of strains During the process of strain isolation and purification, strain XY04 was screened from soil samples collected in Nanning City. Strains XY10 and XY11 were screened from soil samples collected from Guiping City. 2.2 Colony morphology characteristics The colony morphology of strains XY04, XY10, and XY11 after incubation on inverted plates at 30°C for 48 hours is shown in the figure. Figure 1 .

[0057] Depend on Figure 1 It can be seen that the single colonies formed by strain XY04 on LB medium plates are regular round, white, with the surface and bottom of the colony being the same color. The surface of the colony is moist and smooth, with flat edges without serrations. The colony is slightly raised, relatively thin, and has a soft texture, making it easy to pick up.

[0058] The single colonies formed by strain XY10 on LB agar plates are regular round, white, with the surface and bottom of the colony being the same color. The surface of the colony is moist and smooth, with flat, smooth and thick edges. The center of the colony is slightly concave, and the thickness is very thin. The colony texture is relatively soft and easy to pick up.

[0059] The single colonies formed by strain XY11 on LB agar plates are irregularly round, white, with the surface and bottom of the colony being the same color. The surface of the colony is moist and smooth, with smooth and even edges without serrations. The center of the colony is slightly convex, and the colony is relatively thin. The texture of the colony is soft and sticky, similar to jelly. It is easy to pick up but not easy to stick to the inoculation loop.

[0060] 2.3 Colony Biochemical Characteristics The results are shown in Table 3.

[0061] Table 3. Colony biochemical characteristics

[0062] Note: In Gram staining, "+" represents positive bacteria and "-" represents negative bacteria; the salt tolerance test is set with sodium chloride solution concentrations of 5%, 9%, and 13%. In the table, "5%" and "9%" represent that the strain grows well at these concentrations, and "-" indicates that it does not grow at the lowest concentration of 5%; in other test items, "+" represents positive, "++" represents a stronger positive reaction, and "-" represents negative.

[0063] 2.4 Phylogenetic Tree See results Figure 2-4 .

[0064] Combining colony morphological characteristics, colony biochemical characteristics and Figure 2-4The strain XY04 was identified as *Bacillus capsicisae*. Lysinibacillus capsici Strain XY10 is a long-shaped lysine-containing Bacillus. Lysinibacillus macroides Strain XY11 is Lysinibacillus sp . .

[0065] 2.5 Strain tolerance to cadmium selenide See results Figure 5 .

[0066] Depend on Figure 5 It can be seen that in the ninth round of adding LB solid medium plates (Cd) to the strain to improve its tolerance to cadmium selenide, the strain's tolerance to cadmium selenide was significantly improved. 2+ Se 4+ Cd 2+ +Se 4+ All three strains were able to grow at concentrations of 300 mg / L, 2000 mg / L, and 300 mg / L + 2000 mg / L.

[0067] XY04 strain in Cd 2+ It grew normally and well on LB solid medium plates with a concentration of 300 mg / L. In Se... 4+ The strain XY04 grew normally and vigorously on LB solid medium plates at a concentration of 2000 mg / L, showing a tendency to expand beyond the streaked area. It also reacted with selenium in the medium, changing the valence state of selenium and thus exhibiting a deep orange-red color, indicating that strain XY04 has high tolerance to selenium. When treated with a combination of selenium and cadmium, strain XY04 grew well, better than with cadmium alone, but worse than with selenium alone, indicating that 300 mg / kg Cd... 2+ Concentration had a certain inhibitory effect on its growth, while selenium may have played a role in alleviating the inhibition of cadmium.

[0068] XY10 strain in Cd 2+ The sample could grow on LB solid medium plates with a concentration of 300 mg / L, but the growth was not very good, indicating that this concentration of Cd... 2+ It had a strong inhibitory effect on its growth. In Se 4+ The XY10 strain grew normally and vigorously on LB solid medium plates at a concentration of 2000 mg / L, exhibiting a tendency to expand beyond the streaked area. It also reacted with selenium in the medium, altering its valence state and producing a deep orange-red color, indicating that the XY10 strain also has high tolerance to selenium. When treated with a combination of selenium and cadmium, the XY10 strain grew well, better than with cadmium alone, but worse than with selenium alone, indicating that 300 mg / kg Cd... 2+ Concentration had a certain inhibitory effect on its growth, while selenium may have played a role in alleviating the inhibition of cadmium.

[0069] XY11 strain in Cd2+ The sample could barely grow on LB solid medium plates with a concentration of 300 mg / L, and the growth was extremely poor, indicating that this concentration of Cd... 2+ It had a strong inhibitory effect on its growth. In Se 4+ The XY11 strain grew normally and vigorously on LB solid medium plates at a concentration of 2000 mg / L, exhibiting divergent growth. It also reacted with selenium in the medium, changing its valence state and producing a deep orange-red color, indicating that the XY11 strain also has high selenium tolerance. When treated with a combination of selenium and cadmium, the XY11 strain grew well and vigorously, significantly better than with cadmium alone, but worse than with selenium alone. This suggests that 300 mg / kg Cd... 2+ Concentration had a strong inhibitory effect on its growth, while selenium also played a role in alleviating the inhibition of cadmium.

[0070] Overall, the three strains were effective against Cd. 2+ The growth of the three strains on LB solid medium plates with a concentration of 300 mg / L (or mg / kg) was in the following order: XY04 > XY10 > XY11. However, the concentration of selenium was far from reaching the tolerance limit of the three strains. The addition of selenium also greatly alleviated the inhibitory effect of cadmium on the growth of the three strains.

[0071] 2.6 Effects of different pH values ​​on the growth of the strain The effects of inoculum size of 5%, culture temperature of 33℃, and culture time of 36 h on the growth of the strain are shown in the figure. Figure 6 .

[0072] Depend on Figure 6 It can be seen that the OD of the bacterial culture of strain XY04 is... 600 The OD value decreases slightly at pH 5.0-6.0, rises slowly at pH 6.0-9.0, reaches its maximum at pH 9.0, and then decreases again. At pH 9.0, the OD value... 600 Maximum value (OD) 600 =1.558), which is only slightly better than OD. 600 The lowest pH value is 10.0 (OD). 600 =1.509) when the value was 3.24%, showing a narrow fluctuation trend overall, indicating that strain XY04 is not sensitive to changes in pH value and can grow stably in all pH ranges examined.

[0073] OD of bacterial culture of strains XY10 and XY11 600 The values ​​were highest at pH 5.0, then decreased rapidly. XY10 approached equilibrium at pH 6.0; XY11 only began to decline at pH 9.0. At pH 5.0, the OD value of the XY10 strain was... 600The value was 1.582 for XY10 and 1.662 for XY11, indicating that these two strains are more suitable for growth in acidic environments, with XY11 showing slightly better growth performance than XY10.

[0074] 2.7 Effect of different culture times on the phosphate-solubilizing ability of the strain The effects of pH 6.0, culture temperature 31℃, and inoculum size 10% on the phosphate-solubilizing ability of the strain are shown in the figure. Figure 7 .

[0075] Depend on Figure 7 It can be seen that the phosphorus solubility of the three strains gradually increased with the passage of time. The increase rate of strain XY10 was slightly lower than that of the other two strains. Strain XY11 reached its highest phosphorus solubility at 84 hours and then tended to stabilize. Strain XY04 had the highest phosphorus solubility at 96 hours and then fluctuated slightly. Strain XY10 showed a continued slight upward trend after 96 hours. Based on the performance of the three strains, the subsequent culture time was set at 96 hours.

[0076] 2.8 Effect of different inoculum amounts on the phosphate-solubilizing ability of the strain The effects of pH 6.0 and culture temperature 31℃ on the phosphate-solubilizing ability of the strain are shown in the figure. Figure 8 .

[0077] Depend on Figure 8 It can be seen that the phosphorus solubility of strain XY04 shows a decreasing trend in the low inoculum range of 0.5-2%, with a small fluctuation range, and then gradually increases, with a significant increase at 5% inoculum and 149.89 mg / L at 10% inoculum. The phosphorus solubility of strain XY10 showed a spiral increase, with a significant increase at a 5% inoculum amount, reaching 149.49 mg / L at a 10% inoculum amount; The phosphorus solubility of strain XY11 fluctuated little within the inoculum size range of 0.5-2%, gradually increasing at a 5% inoculum size, reaching 139.96 mg / L at a 10% inoculum size. The phosphorus solubility of all three strains showed a significant increase at 5% inoculum size, reaching its maximum at 10%.

[0078] 2.9 Effect of different temperatures on the phosphate-solubilizing ability of the strain The effect of pH 6.0 and inoculum size of 10% on the phosphate-solubilizing ability of the strain is shown in the figure. Figure 9 .

[0079] Depend on Figure 9 It can be seen that the phosphorus solubility of strain XY04 is the highest at a culture temperature of 31℃, and the phosphorus solubility is significantly lower at other temperatures, indicating that its optimal culture temperature is 31℃. Strain XY10 showed similar behavior to XY04, with the highest phosphorus solubility at 31℃ and little difference in phosphorus solubility at other temperatures. Strain XY11 exhibited good phosphorus solubility under all temperature conditions, demonstrating that it has a wide temperature adaptability range. It performed best at 28℃, with a phosphorus solubility of 163.12 mg / L.

[0080] 2.10 Effect of different pH values ​​on the phosphate-solubilizing ability of the strain The effects of inoculum size of 10%, culture temperature of 31℃, and different pH values ​​on the phosphorus solubility and phosphorus solubility of the strain are shown in the figure. Figure 10 .

[0081] Depend on Figure 10 It was found that all three strains exhibited better phosphorus solubility under slightly acidic conditions (pH 5.0-6.0), and their phosphorus solubility gradually decreased with increasing pH. The phosphorus solubility of all three strains was optimal at pH 5.0, with the following order: XY11 (176.35 mg / L) > XY10 (154.98 mg / L) > XY04 (148.80 mg / L). No significant change was observed at pH 6.0, but all strains showed varying degrees of decrease at pH 7.0. Strain XY10 showed a smaller decrease in phosphorus solubility under neutral and alkaline conditions compared to the other two strains. The pH of paddy fields in southern China is generally low, often acidic, typically between 5.0 and 6.5. Therefore, these three strains are well-suited for application in southern paddy fields.

[0082] 2.11 Effect of different initial cadmium concentrations on the phosphorus-solubilizing ability of the strain See results Figure 11 .

[0083] Depend on Figure 11 It can be seen that strain XY04 exhibited a certain degree of phosphorus solubility promotion at a cadmium concentration of 3 mg / L, with its phosphorus solubility increasing by 14.68% compared to 1 mg / L. However, as the cadmium concentration further increased, the phosphorus solubility was inhibited, but the phosphorus solubility increased significantly again at 9 mg / L.

[0084] Strain XY10 exhibits unique cadmium tolerance; at cadmium concentrations of 3-5 mg / L, its phosphorus-solubilizing effect is not only not inhibited but also shows a stimulating effect, with phosphorus solubility exceeding that of Cd. 2+ At concentrations of 3 mg / L and 5 mg / L, the relative phosphorus solubility increased by 5.79% and 16.41% compared to a concentration of 1 mg / L, respectively. However, at high concentrations of 7-9 mg / L, the phosphorus solubility showed a decreasing trend. The phosphate-solubilizing ability of strain XY11 is affected by Cd 2+ The inhibitory effect is significant; even a low concentration of cadmium (1 mg / L) has an inhibitory effect.

[0085] 2.12 Effect of different initial selenium concentrations on the phosphorus-solubilizing ability of strains See results Figure 12 .

[0086] Depend on Figure 12 It can be seen that the phosphorus solubility of strain XY04 generally fluctuates within a small range, and its phosphorus solubility is not high under all treatments, indicating that the strain is relatively sensitive to selenium metal, and its phosphorus solubility is significantly inhibited.

[0087] Strain XY10 exhibited better selenium tolerance, showing higher phosphorus solubility than other strains at all concentrations. 4+ The highest level of phosphorus solubility was observed at a concentration of 1 mg / L, after which the degree of inhibition gradually increased with increasing selenium concentration. Strain XY11 is most sensitive to selenium, and its phosphorus solubility is in the range of Se. 4+ The highest concentration was observed at 1 mg / L, followed by a sharp drop to 80.75% of the 1 mg / L concentration at 3 mg / L, after which the effect on Se... 4+ The reaction was minimal with increasing concentration, and remained stable at a low level.

[0088] 2.13 Effects of bacterial strains on soil available phosphorus content See results Figure 13 .

[0089] Depend on Figure 13 It was found that after 7 days of treatment, the XY04 treatment group showed the most significant phosphorus solubility effect. The available phosphorus content in the soil of the XY04-10% treatment group reached 29.68 mg / kg, which was 94.13% higher than that of the control group. The XY04-5% treatment group was the second most effective, with an increase of 48.42% in available phosphorus content. The XY10-10% and XY11-5% treatment groups also showed significant phosphorus solubility effects, which were 27.45% and 24.76% higher than that of the control group, respectively. However, the available phosphorus content in the soil of the XY10-5% and XY11-10% treatment groups was actually lower than that of the control group.

[0090] After 14 days of treatment, the available phosphorus content in the soil of all treatment groups was higher than that of the control group, indicating that all strains played a phosphorus-solubilizing role. Among them, the XY04-5% treatment group showed the most outstanding performance, with the available phosphorus content in the soil reaching 38.07 mg / kg, an increase of 123.68% compared with the control group; the XY04-10% treatment group was the second best, with an increase of 96.14%. The phosphorus-solubilizing effect of all treatment groups at 14 days was generally better than that at 7 days, indicating that the strains need a certain amount of time to adapt to the soil environment and fully exert their phosphorus-solubilizing function.

[0091] After 21 days of treatment, the phosphorus-solubilizing effect in half of the treatment groups decreased compared to 14 days, but still maintained a positive effect. The XY10-5% treatment group showed the most significant phosphorus-solubilizing effect, with the soil available phosphorus content increasing by 112.86%, far exceeding other treatment groups; the XY10-10% treatment group was second, with an increase of 62.31%. The available phosphorus content in the XY11-10% and XY04-5% treatment groups increased by more than 50% compared to the control group. The outstanding phosphorus-solubilizing effect shown by the XY10-5% treatment group at 21 days indicates that this strain may have long-lasting phosphorus-solubilizing characteristics, making it suitable for agricultural production environments requiring continuous phosphorus supply.

[0092] 2.14 Effects of bacterial strains on available cadmium content in soil See results Figure 14 .

[0093] Depend on Figure 14 It was found that all three strains effectively reduced the content of available cadmium in the soil, demonstrating a significant inhibitory effect on cadmium activity. After 7 days of treatment, the XY11-5% treatment group showed the most significant cadmium inhibition effect, with an available cadmium content reduced by 63.53% compared to the control group, while the XY11-10% treatment group reduced it by 62.34%. Even though the cadmium inhibition effect of strain XY10 was relatively weaker, it still reduced the content by 38.23% (5% inoculum) and 44.72% (10% inoculum), respectively. Among the three strains, the available cadmium content was lowest in the XY11 treatment group, followed by XY04 and XY10.

[0094] After 14 days of treatment, the cadmium inhibition effect of each treatment group was weaker than that after 7 days, but it still remained at a high level overall. Among them, the XY11-10% treatment group performed the best, with the effective cadmium content reduced by 39.05% compared with the control group. Among the three strains, strain XY11 showed the strongest cadmium inhibition ability in the short to medium term, followed by XY04 and XY10.

[0095] After 21 days of treatment, all treatment groups continued to maintain good cadmium inhibition effects. The XY11-10% and XY10-10% treatment groups performed the best, with effective cadmium content decreasing by 30.20% and 28.90% respectively compared with the control group, indicating that these two strains have good long-term cadmium inhibition ability under high inoculum conditions.

[0096] The differences in cadmium inhibition effects among different strains are related to their metabolic characteristics and mechanisms of action. In the long term, the cadmium inhibition effect of strain XY11 is more persistent, which may be related to their ability to continuously secrete cadmium-inhibiting substances or alter the soil microenvironment.

[0097] 2.15 Effects of bacterial strains on soil available selenium content See results Figure 15 .

[0098] Depend on Figure 15It was found that after 7 days of treatment, except for the XY10-5% treatment group, all other treatment groups reduced the content of available selenium in the soil. The available selenium content of the XY10-5% treatment group was 0.149 mg / kg, an increase of 42.96% compared to the control group. The available selenium content of the XY04-10% treatment group decreased by 42.08%, and the XY11-10% treatment group decreased by 35.87%. The available selenium content of the other treatment groups was basically the same as that of the control group. Overall, the 5% inoculum amount reduced the amount of available selenium less than that of the 10% inoculum. This phenomenon may be due to the strain consuming soluble selenium in the environment during the initial growth process, or the secretion of certain substances temporarily reducing the solubility of selenium. Further investigation is needed. Among the three strains, the XY10 treatment group showed the smallest decrease in available selenium content, and even showed an increase.

[0099] After 14 days of treatment, the available selenium content in the XY10-5% treatment group was 0.137 mg / kg, an increase of 1.31% compared to the control group. The available selenium content in all other treatment groups was lower than that in the control group to varying degrees. However, the decrease under high inoculum conditions was generally less than that under 7 days, indicating that the inhibitory effect of high inoculum strains on selenium weakened with prolonged treatment. Among them, the XY04-5% and XY04-10% treatment groups showed the largest decreases in available selenium content compared to the control group, decreasing by 19.91% and 19.40%, respectively. The decreases in other treatment groups were smaller. Among the three strains, strain XY10 showed the lowest decrease in available selenium, even showing a slight increase. The available selenium content was not significantly different under the two inoculum conditions.

[0100] After 21 days of treatment, most treatment groups continued to show selenium inhibition compared to the control group. The XY04-10% treatment group showed selenium activation, with the effective selenium content increasing by 0.23% compared to the control group. Over three time periods, the effective selenium content in the XY04-10% treatment group increased from 0.060 mg / kg (7 days), 0.109 mg / kg (14 days), and 0.147 mg / kg (21 days) to -42.08% (7 days), 19.40% (14 days), and 0.23% (21 days) respectively compared to the control group. This result suggests that strain XY04 may have long-term selenium activation potential. Furthermore, the effect of the XY10-5% treatment group on soil available selenium content was the opposite of that of the XY04-10% treatment group. Over three time periods, the available selenium content in the XY10-5% treatment group increased from 0.149 mg / kg (7d), 0.137 mg / kg (14d), and 0.120 mg / kg (21d) to 42.96% (7d), 1.31% (14d), and 18.46% (21d) in the control group, respectively. This indicates that strain XY10 has the ability to activate soil selenium in the short term and its activation performance is relatively stable.

[0101] Therefore, among these three strains, there is strain XY10, which has the short-term activation potential for soil selenium, and strain XY04, which has the long-term activation potential for soil selenium. In actual agricultural production activities, the appropriate strain can be applied according to actual needs.

[0102] 2.16 Effects of bacterial strains under selenium and cadmium stress on rice seed germination indices The results are shown in Tables 5 and 6.

[0103] Table 5. Effects of strains on rice seed germination potential under selenium-cadmium stress (%)

[0104] Note: All data in the table are mean ± standard deviation. Different lowercase letters in each column indicate significant differences between treatments of different strains (p<0.05).

[0105] Table 6. Effects of strains on rice seed germination rate under selenium-cadmium stress (%)

[0106] Note: All data in the table are mean ± standard deviation. Different lowercase letters in each column indicate significant differences between treatments of different strains (p<0.05).

[0107] As shown in Tables 5 and 6, under selenium- and cadmium-free stress conditions, the germination potential and germination rate of rice seeds soaked by the four bacterial strains were higher than those of the control group (CK, unsoaked bacterial suspension). The germination potential of the groups soaked in bacterial suspensions of strains XY10, XY11, and XY04 increased by 21.82%, 55.76%, and 50.91%, respectively, and the germination rate increased by 5.91%, 11.41%, and 9.84%, respectively, compared with the control group. This indicates that all three strains have a certain ability to promote growth.

[0108] Under selenium-cadmium stress, the germination potential and germination rate of the CK group and the group soaked with the three bacterial strains were lower than those under no selenium-cadmium stress, indicating that the selenium-cadmium solution exerted a stress effect on rice seed germination and inhibited it. Compared with the control group (CK) that was not soaked in bacterial solution, the seeds soaked in the bacterial suspension of the three bacterial strains had greater germination potential and germination rate at the same concentration of selenium-cadmium, indicating that the three bacterial strains could alleviate the inhibitory effect of selenium-cadmium stress on seed germination to some extent. Among them, the seeds soaked in the bacterial suspension of strain XY11 showed the highest germination potential and germination rate under all treatments, demonstrating its strong tolerance to selenium and cadmium stress and its ability to promote growth.

[0109] 2.17 Effects of strains under selenium and cadmium stress on germination index and vigor index of rice seeds The results are shown in Tables 7 and 8.

[0110] Table 7. Effects of bacterial strains on rice seed germination index under selenium-cadmium stress.

[0111] Note: All data in the table are mean ± standard deviation. Different lowercase letters in each column indicate significant differences between treatments of different strains (p<0.05).

[0112] Table 8. Effects of strains on rice seed vigor index under selenium-cadmium stress.

[0113] Note: All data in the table are mean ± standard deviation. Different lowercase letters in each column indicate significant differences between treatments of different strains (p<0.05).

[0114] As shown in Tables 7 and 8, under selenium- and cadmium-free stress, the germination indices of strains XY10, XY11, and XY04 were 23.67%, 45.40%, and 47.54% higher than those of the control group, respectively, and the vigor indices were 62.54%, 298.27%, and 203.19% higher, respectively. This further confirms that the three strains have a positive promoting effect on rice seed germination, with strains XY11 and XY04 showing particularly significant growth-promoting effects.

[0115] Under selenium and cadmium stress, the germination index and vigor index of rice seeds soaked in the four bacterial strains were lower than those in the selenium and cadmium-free stress environment. However, compared with the control group without bacterial suspension, the germination index and vigor index of seeds soaked in bacterial suspension were still higher than those of the control group (CK). This indicates that even under selenium and cadmium stress, the three bacterial strains can still alleviate the inhibitory effect of selenium and cadmium stress on seed germination to a certain extent, maintaining and improving the germination ability and vigor of seeds. Among them, the seeds soaked in the suspension of strain XY11 showed the highest germination index and vigor index under all selenium and cadmium stress treatments, which again proves that strain XY11 has strong tolerance and growth-promoting ability to selenium and cadmium stress. XY04 was the second highest, while the seeds soaked in XY10 also showed a certain growth-promoting effect under selenium and cadmium stress, but the increase in germination index and vigor index was relatively small.

[0116] 2.18 Effects of strains under selenium and cadmium stress on root length and seedling growth in rice The results are shown in Tables 9 and 10.

[0117] Table 9. Effects of strains on rice root length (cm) under selenium and cadmium stress.

[0118] Table 10 Effects of strains on rice seedling growth (cm) under selenium and cadmium stress

[0119] As shown in Tables 9 and 10, under selenium- and cadmium-free stress, the root length and seedling length of rice treated with strains XY10, XY11, and XY04A4 were significantly higher than those of the control group (CK). Root length increased by 32.69%, 174.04%, and 98.08%, respectively, and seedling length increased by 12.11%, 65.79%, and 43.16%, respectively. This indicates that these three strains had a greater effect on promoting root growth than on promoting seedling growth. Among them, the XY11 treatment group showed the best performance in both root length and seedling length, increasing by 174.04% and 65.79% compared to the control group, respectively. This was followed by the XY04A treatment group, which increased by 98.08% and 43.16%, respectively. Although the XY10 treatment group also showed some growth-promoting effect, the increase was smaller compared to the XY11 and XY04A treatment groups. This further confirms the positive promoting effect of these three strains on rice growth.

[0120] Under selenium-cadmium stress, except for Cd 2+ Except for a concentration of 5 mg / L, the root length and seedling length of rice in the three strain treatment groups were lower than those in the selenium- and cadmium-free stress environment. However, compared with the control group that was not soaked in bacterial solution, the root length and seedling length of rice soaked in bacterial suspension were still higher than those of the control group. 2+ At a concentration of 5 mg / L, the root length of all treatment groups increased to some extent. Except for the XY04 treatment group, the seedling length of other treatment groups also increased to some extent, but the increase was less than that of the root length.

[0121] In summary, based on morphological observation and molecular biological identification, strain XY04 of this invention has been identified as *Bacillus capsicisae*. Lysinibacillus capsici Furthermore, experiments have revealed that strain XY04 of this invention possesses phosphorus-solubilizing, selenium- and cadmium-tolerant, and growth-promoting effects. It can increase the available phosphorus and selenium content in the soil, reduce the available cadmium content, and help improve the utilization rate of available phosphorus in the soil, as well as achieve selenium enrichment and cadmium reduction in crops. At the same time, it can promote crop growth and improve their tolerance to selenium and cadmium stress, which is of great significance for the production of selenium-enriched green products.

[0122] The foregoing description of specific exemplary embodiments of the invention is for illustrative and explanatory purposes. These descriptions are not intended to limit the invention to the precise forms disclosed, and it will be apparent that many changes and variations can be made in accordance with the foregoing teachings. The exemplary embodiments were chosen and described in order to explain the specific principles of the invention and its practical application, thereby enabling those skilled in the art to implement and utilize various different exemplary embodiments of the invention, as well as various different choices and variations. The scope of the invention is intended to be defined by the claims and their equivalents.

Claims

1. A strain XY04, characterized in that, The strain XY04 is taxonomically named *Bacillus capsicis-lysine*. Lysinibacillus capsici It was deposited at the Guangdong Provincial Center for Microbial Culture Collection on July 1, 2025, with accession number GDMCC No: 66618.

2. The use of strain XY04 according to claim 1 in phosphorus solubilization.

3. The use of strain XY04 of claim 1 in increasing the available phosphorus content in soil.

4. The use of strain XY04 of claim 1 in activating soil selenium and thus increasing effective selenium content.

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