Bacillus velezensis h501 and application thereof in promoting growth of kelp seedlings

Abstract

This invention relates to a strain of *Bacillus belye* H501 and its application in promoting the growth of kelp seedlings, belonging to the field of microbiology. The strain was deposited on December 24, 2025, at the China General Microbiological Culture Collection Center (CGMCC), with accession number CGMCC NO. 37160. The strain can grow normally within a salinity range of 15-25℃ and 1-7%, highly matching the kelp seedling cultivation environment, and does not produce alginate lyase, thus not disrupting the natural defense barrier of kelp. Applying this strain to kelp seedling cultivation can shorten the seedling cycle by approximately 3-5 days, effectively reducing energy consumption and labor costs. Furthermore, H501 exhibits significant antagonistic activity against the common aquatic pathogen *Aeromonas hydrophila*, demonstrating potential for biocontrol applications.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology, specifically relating to Bacillus belye H501 and its application in promoting the growth of kelp seedlings. Background Technology

[0002] Macroalgae, as a key marine biological resource, have seen significant growth in global aquaculture over the past two decades, becoming a crucial pillar of the blue economy. Macroalgae, represented by kelp and laver, not only possess extremely high nutritional and health benefits but also play a positive role in environmental governance, including the development of ecological feed for aquaculture, the remediation of eutrophic waters, and the increase of marine carbon sinks. Furthermore, due to the short growth cycle, easy degradation, and lignin-free structural characteristics of algal biomass, it shows great potential in the development of biofuels and high-value-added biomaterials. With the increasing global demand for food security, ecological safety, and energy transition, promoting the high-quality and sustainable development of the macroalgae industry has become an important research direction in the field of marine biotechnology.

[0003] During natural evolution, macroalgae have established complex mutualistic symbiotic relationships with the microorganisms in their habitats. Among these, endophytic bacteria colonizing the algal tissues form a highly stable physiological and metabolic connection with the host. Studies have shown that endophytic bacteria can directly promote algal growth and development by synthesizing plant hormones, and enhance the host's nutrient acquisition capacity through metabolic activities such as nitrogen fixation, phosphorus solubilization, and the production of the siderophore bacillibactin. This endogenous symbiotic mechanism is a crucial ecological strategy for algae to adapt to complex marine environments and enhance their resilience and disease resistance. Therefore, utilizing endophytic bacteria for bioregulation is considered an important biological approach to improving algal farming efficiency and optimizing seedlings.

[0004] However, the seedling cultivation process for large algae such as kelp still faces numerous bottlenecks in existing large-scale aquaculture systems. Current indoor seedling cultivation methods generally suffer from long cycles, high production costs, and sensitivity to abiotic stresses. Although microbial regulation technology is relatively mature in terrestrial agriculture, with probiotics such as Bacillus belye having achieved commercial application, its application in marine algae seedling cultivation is still in the exploratory stage. Existing microbial-assisted seedling cultivation attempts often suffer from drawbacks such as high randomness in strain selection, weak affinity with the host, and unstable colonization, resulting in unsatisfactory growth-promoting effects and difficulty in achieving industrial-scale promotion. How to screen and utilize specific endophytic strains with highly efficient growth-promoting functions targeting the early growth and development characteristics of kelp is a core technical challenge that urgently needs to be overcome in the field of marine biotechnology.

[0005] To address the problems of long seedling cultivation cycles, low strain colonization efficiency, and insignificant growth-promoting effects in existing technologies, there is an urgent need to discover superior microbial strains that can deeply integrate with kelp tissue, possess clear genetic regulatory mechanisms, and significantly shorten the seedling cultivation cycle. By conducting in-depth research on the interaction between endophytic bacteria and algae in the early developmental stages, and developing an efficient microbial-assisted seedling cultivation technology system, not only can the production cost of kelp seedling cultivation be effectively reduced, but it will also provide crucial technical support for the green and sustainable development of large-scale algae farming. Summary of the Invention

[0006] The purpose of this invention is to provide a Bacillus velezensis H501 strain and its application in promoting the growth of kelp seedlings, so as to solve the technical problems of long kelp seedling cultivation cycle, high cost and poor strain compatibility.

[0007] To achieve the above objectives, the present invention adopts the following technical solution: One of the objectives of this invention is to provide a Bacillus velezensis H501 strain, characterized in that the strain was deposited on December 24, 2025, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC NO. 37160.

[0008] Furthermore, the strain possesses at least one of the following biological characteristics: a) It can form a transparent phosphorus-solubilizing zone on a culture medium in which lecithin is the only phosphorus source; b) It can produce an orange-brown chelation ring on the Chromium Azure S (CAS) detection plate; c) It exhibits antagonistic activity against Aeromonas hydrophila at 20°C; d) It does not have alginate lyase activity.

[0009] Furthermore, the genome of the strain contains a gene cluster encoding at least one of the following secondary metabolites: surfactant, macrolide H, sporeene, interferon, dapoxetine, siderophores, and lysozyme.

[0010] The second objective of this invention is to provide the application of the aforementioned Bacillus belyssus H501 in promoting the growth of kelp seedlings.

[0011] A third objective of this invention is to provide a method for promoting the growth of kelp seedlings, the method comprising the step of adding a suspension of Bacillus belye H501 to a kelp seedling system.

[0012] Furthermore, the final concentration of the bacterial suspension in the seedling system is 4 × 10⁻⁶.7 CFU / mL up to 8×10 7 CFU / mL.

[0013] Furthermore, the method includes the step of periodically supplementing the seedling system with a suspension of Bacillus vesiculus H501 during the seedling stage to maintain an effective bacterial concentration.

[0014] The fourth objective of this invention is to provide a microbial preparation comprising the aforementioned Bacillus belyssus H501 and an agriculturally or aquaculture-acceptable carrier.

[0015] Furthermore, the formulation is in the form of a liquid bacterial agent, a solid bacterial agent, or a wettable powder.

[0016] The fifth objective of this invention is to provide the application of Bacillus belyssus H501 in the preparation of preparations that inhibit Aeromonas hydrophila.

[0017] The beneficial effects of this invention compared to existing technologies are as follows: This invention isolates an endophytic bacterium, *Bacillus velezensis* H501, from kelp. This strain can grow normally within a salinity range of 15-25℃ and 1-7%, highly matching the kelp seedling cultivation environment, and does not produce alginate lyase, thus not damaging the natural defense barrier of kelp. Co-culture experiments using this strain in kelp seedling cultivation show that adding 8×10⁻⁶... 7 The kelp seedlings treated with cfu / mL H501 bacterial suspension were twice as long as those in the control group, shortening the seedling cultivation cycle by approximately 3-5 days and effectively reducing energy consumption and labor costs in kelp seedling cultivation. Genomic analysis and in vitro experiments confirmed that strain H501 possesses multiple plant growth-promoting functions, including nitrogen fixation, phosphorus solubilization, and secretion of the siderophore bacillibactin, which can improve the efficiency of kelp in acquiring key nutrients such as nitrogen, phosphorus, and iron. Furthermore, H501 exhibits significant antagonistic effects against the common aquatic pathogen Aeromonas hydrophila, and its genome contains gene clusters for the synthesis of various antibacterial substances such as surfactant, macrolactamin, bacillaene, and difficidin, demonstrating potential for biocontrol applications.

[0018] This invention provides excellent strain resources and theoretical basis for the development of microbial-assisted kelp seedling cultivation technology, which is expected to replace or reduce the use of chemical fertilizers and promote the development of the kelp farming industry towards a green, efficient and sustainable direction. Attached Figure Description

[0019] Figure 1 The image shows the identification results of strain H501, where A is a phylogenetic tree constructed based on the 16S rRNA gene sequence, B is the colony morphology, and C is the Gram staining microscopic observation results. Figure 2 The graph shows the tolerance of strain H501 under different temperature and salinity conditions. A is the growth curve at 10-25℃, and B is the number of bacteria after 48 hours of culture at 1-7% salinity. Figure 3 The results of the detection of plant growth promotion and antagonistic effects of strain H501 are shown in the figure. A is the detection of nitrogen fixation ability, B is the detection of organophosphorus solubility, C is the detection of siderophore secretion ability, D is the detection of no IAA plant growth hormone production, E shows a significant inhibitory effect on the aquatic pathogen Aeromonas hydrophila, and F shows that the detection of alginate enzyme activity indicates that it does not have the ability to degrade alginate. Figure 4 The figure shows the results of the co-culture experiment between H501 and kelp. In the figure, A represents the change in gametophyte germination rate, B represents the change in seedling length, C represents the change in seedling width, and D represents the comparison of specific growth rate (SGR). Figure 5 This section presents a heatmap of the genome of strain H501 and an analysis of average nucleotide identity. A represents the genome map of strain H501: the first circle (outermost) represents the genome size; the second and third circles represent the positive and negative strand genes and COG functional classification (AV on the right); the fourth circle shows triangles representing tRNA (green for the positive strand and blue for the negative strand), squares representing rRNA (red for the positive strand and purple for the negative strand); the fifth circle represents the genomic GC content; and the sixth circle represents the genomic GC shift. B is a heatmap of the average nucleotide identity (ANI) between strain H501 and six closely related phylogenetic species. Figure 6 This diagram illustrates the functional pathways related to growth promotion and environmental adaptation in strain H501. A represents the phosphate transport system in the genome, and B represents the Na+ transport system. + / H + Reverse transport and osmotic pressure regulation; C is a schematic diagram of the pathways related to siderophore synthesis. Detailed Implementation

[0020] The technical solution of the present invention will be described in detail below through specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of protection of the present invention. Without departing from the spirit and concept of the present invention, those skilled in the art can make various modifications and equivalent substitutions, all of which fall within the scope of protection of the present invention. Unless otherwise specified, the reagents and instruments used in the following embodiments are commercially available products.

[0021] Example 1: Isolation and Identification of Bacillus velezensis H501 1.1 Strains Isolation Kelp samples were collected from a kelp farm in Xiapu County, Ningde City, Fujian Province. Healthy kelp leaves (5cm x 5cm) were taken, rinsed with sterile water to remove surface mucus and epiphytes, and then transferred to a clean bench. Surface disinfection was performed by sequentially treating with 75% ethanol for 1 minute and 3% sodium hypochlorite solution for 2 minutes, followed by repeated rinsing with sterile water 4-5 times. The disinfected tissue was then ground in a sterile mortar, and 1 mL of sterile water was added to prepare a bacterial suspension. 200 μL of the bacterial suspension was evenly spread onto LB agar plates, with a control sample prepared using the final rinse solution. The plates were incubated at 37°C for 24 hours, and colony morphology was observed. The dominant strain was purified using the streak plating method, ultimately obtaining purified strain H501.

[0022] 1.2 Morphological identification On LB solid medium, H501 colonies are milky white, opaque, round, raised, with a rough and dry surface, and neat, slightly translucent edges. Figure 1 (B in the middle).

[0023] Under an optical microscope (Nikon Ti-E-A1R, Tokyo, Japan), the bacteria are rod-shaped and Gram-positive. Figure 1 The C in the figure conforms to the typical morphological characteristics of the Bacillus genus.

[0024] 1.3 Molecular biological identification Strain H501 was sent to Sangon Biotech (Qingdao) for 16S rRNA gene amplification and sequencing using universal primers 27F / 1492R. The obtained sequences were BLAST-aligned with the NCBI database, and a phylogenetic tree was constructed using the maximum likelihood method (bootstrap=1000) with MEGA 11.0 software. The results showed that H501 clustered in the same branch as Bacillus velezensis FZB42 and CBMB205. Figure 1 (A in the middle).

[0025] Further average nucleotide identity analysis revealed that H501 shared an ANI of 97.73% with Bacillus velezensis FZB42 and 97.55% with Bacillus velezensis CBMB205, both exceeding the 95% species-defining threshold. Based on the combined morphological characteristics and molecular identification results, strain H501 was identified as Bacillus velezensis.

[0026] 16 rRNA of Bacillus velezensis H501 (SEQ ID NO.1):

[0027] Example 2: Determination of physiological characteristics of Bacillus velezensis H501 2.1 Temperature adaptability test The cultured Bacillus belye H501 bacterial suspension was inoculated into LB liquid medium at a 1% (v / v) inoculum, and then incubated at 10℃, 15℃, 20℃, and 25℃ with constant temperature shaking at 180 rpm. Samples were taken periodically during the incubation process, and the absorbance (OD) of the culture medium at a wavelength of 600 nm was measured. 600 value).

[0028] The measurement results are as follows Figure 2 As shown in A in the diagram. From Figure 2 As can be seen from A in the figure, the Bacillus belye H501 has good temperature adaptability in the range of 15℃-25℃, of which 20℃ is the optimal growth temperature.

[0029] 2.2 Salinity Adaptability Measurement Sodium chloride (NaCl) was added to LB liquid medium to prepare saline LB liquid media with salinities of 1%, 3%, 5%, and 7%, respectively. The *Bacillus belyceae* H501 bacterial suspension was inoculated into the media with different salinities at an inoculum rate of 1% (v / v), and cultured at 20°C and 180 rpm for 24 hours with constant temperature shaking. The OD values ​​of each culture group were then measured. 600 value.

[0030] The measurement results are as follows Figure 2 As shown in B in the diagram. (By...) Figure 2 As indicated by B in the figure, the *Bacillus belye* H501 can grow normally within a salinity range of 1%-7%, with the optimal salinity for growth being 1%. These results demonstrate that the strain described in this invention possesses a wide salinity tolerance range and good salt tolerance.

[0031] Example 3: Determination of the life-promoting and antagonistic effects of Bacillus velezensis H501 3.1 Nitrogen fixation capacity determination The *Bacillus belyssioides* H501 was inoculated onto Ashby nitrogen-free medium plates and incubated at its optimal growth temperature of 20°C for 7 days. The results are as follows: Figure 3 As shown in A in the diagram. From Figure 3 As indicated by A in the diagram, strain H501 can still grow normally and form colonies under nitrogen-free conditions. These results demonstrate that the strain described in this invention possesses nitrogen-fixing ability or has highly efficient nitrogen utilization.

[0032] 3.2 Determination of phosphorus solubility The *Bacillus belye* H501 strain was inoculated onto inorganic phosphorus culture plates with tricalcium phosphate as the sole phosphorus source and organic phosphorus culture plates with lecithin as the sole phosphorus source, respectively, and incubated at 20°C for 7 days. The results are as follows: Figure 3 As shown in B in the diagram. (By...) Figure 3 As indicated by B in the diagram, strain H501 can form a distinct transparent phosphorus-solubilizing zone (approximately 1 cm in diameter) on organic phosphorus culture media, while no significant phosphorus-solubilizing effect was observed on inorganic phosphorus culture media. These results demonstrate that the strain described in this invention retains its specific ability to dissolve organic phosphorus even at relatively low temperatures, which helps to convert poorly soluble organic phosphorus in the environment into bioavailable phosphorus.

[0033] 3.3 Detection of Ferric Carrier Generation The *Bacillus belye* H501 strain was inoculated onto CAS (Cromazine S) agar plates and incubated at 20°C for 2–4 days. The test results are as follows: Figure 3 As shown in C. From Figure 3 As indicated by C, a distinct orange-brown chelation ring appeared around the colony of strain H501. These results confirm that the strain described in this invention can efficiently secrete siderophores and possesses a strong competitive ability to acquire iron ions, which is beneficial to its competitive advantage in environmental colonization.

[0034] 3.4 Determination of indole-3-acetic acid yield The *Bacillus belye* H501 was inoculated into LB liquid medium containing 0.1% (w / v) L-tryptophan and cultured with shaking at 20°C for 2 days. The fermentation broth was centrifuged, and the supernatant was collected and mixed with Salkowski's reagent. The absorbance (OD) of the reaction solution at 540 nm was measured. 540 Value). Measurement results are as follows: Figure 3 As shown in D in the diagram. (By...) Figure 3 As indicated by D, no colorimetric reaction or characteristic absorption peak representing IAA production was detected. These results demonstrate that, under the current testing conditions, the strain H501 described in this invention does not possess the ability to synthesize IAA.

[0035] 3.5 Determination of antagonistic effect against pathogens The antagonistic activity of *Bacillus belyssiensis* H501 against five common indicator pathogens—*Aeromonas hydrophila*, *Escherichia coli*, *Staphylococcus aureus*, *Vibrio anguillarum*, and *Vibrio parahaemolyticus*—was determined using the agar diffusion method. The sterile fermentation broth of strain H501 was collected and added dropwise to plates coated with the aforementioned test pathogens, and incubated at 20°C. The results are as follows: Figure 3As shown in E in the figure. The results show that the H501 sterile fermentation broth produced a significant inhibition zone against *Aeromonas hydrophila*, while exhibiting weak or no inhibitory effect on other tested strains. These results indicate that the H501 strain described in this invention possesses significant and specific antagonistic activity against *Aeromonas hydrophila* at 20℃, and has great potential for biocontrol of *Aeromonas hydrophila* infection and related diseases in colder waters or under the actual environmental temperatures of aquaculture.

[0036] 3.6 Assay of alginate lyase activity The *Bacillus belye* H501 strain was inoculated onto a culture medium plate using sodium alginate as the sole carbon source and incubated at 20°C for 2 days. The results are as follows: Figure 3 As shown in F in the diagram. (By...) Figure 3 As indicated by E, no transparent hydrolysis zone was observed around the colony. These results demonstrate that strain H501 of this invention does not possess the ability to degrade alginate.

[0037] Example 4: Co-culture and growth promotion experiment of Bacillus velezensis H501 with kelp 4.1 Experimental Methods (1) Preparation of bacterial suspension: A single colony of the activated Bacillus belye H501 was picked and inoculated into 100 mL of LB liquid medium and cultured at 20 °C and 180 rpm for 18 hours with constant temperature shaking. Subsequently, the culture was centrifuged at 10,000 rpm for 5 minutes to collect the bacterial precipitate; the precipitate was washed twice with sterile seawater and then resuspended in sterile seawater to obtain the H501 bacterial suspension for later use.

[0038] (2) Treatment of kelp gametophytes: Kelp (Saccharina japonica) gametophytes were selected and mixed at a ratio of female to male gametophytes of 3:1. After mechanical crushing, the mixture was filtered through a 400-mesh sieve and then treated with 1.5 g / m 2 The density is such that the gametes are evenly sprayed and adhered to the glass slide.

[0039] (3) Co-culture conditions and grouping: The culture conditions were set as follows: temperature 10±2℃, photoperiod of 16 hours light / 8 hours dark, and light intensity of 28.57 μmol·m -2 ·s -1 Glass slides with attached kelp gametes were placed in the above environment, and the following three experimental groups were set up: Control group: The culture system was supplemented with only an equal volume of sterile seawater; Low concentration treatment group: The H501 bacterial suspension was added to the culture system to achieve a final concentration of 4 × 10⁻⁶. 7CFU / mL; High-concentration treatment group: The H501 bacterial suspension was added to the culture system to achieve a final concentration of 8 × 10⁻⁶. 7 CFU / mL.

[0040] (4) Routine maintenance and measurement: During the co-cultivation period, the seawater is changed once a week and the H501 bacterial suspension of the corresponding concentration is replenished. Starting from the first week after the gametophyte germination, the germination rate, length and width of the seedlings are measured twice a week, and the specific growth rate is calculated.

[0041] 4.2 Experimental Results (1) Effect on kelp germination rate: The measurement results are as follows Figure 4 As shown in A in the diagram. During the early stage of cultivation (day 12), there was no significant difference in germination rate among the treatment groups; from day 24 onwards, the germination rates of both treatment groups supplemented with the H501 bacterial suspension significantly exceeded those of the control group. The high-concentration treatment group achieved the highest germination rate on day 31.

[0042] (2) Effect on promoting seedling length: The test results are as follows Figure 4 As shown in B in the figure. From day 21 of cultivation, the seedlings in the H501 treatment group were significantly longer than those in the control group, and exhibited a clear concentration-dependent effect (i.e., the higher the concentration, the more pronounced the promotion). By day 50 of cultivation, the seedlings in the high-concentration treatment group were approximately twice the length of the control group.

[0043] (3) The promoting effect on seedling width: The test results are as follows Figure 4 As shown in C. From day 21 of cultivation, the low-concentration treatment group showed the most significant effect in promoting the widening of kelp seedlings, and its advantage in width growth remained dominant in the later stages of cultivation.

[0044] (4) Specific growth rate analysis: The measurement results are as follows Figure 4 As shown in D in the figure. During the two-month cultivation period, the length-specific growth rate of kelp seedlings in the H501 treatment group was significantly increased by approximately 0.3 times compared to the control group; in terms of germination rate and width-specific growth rate, it remained stable compared to the control group (no significant difference).

[0045] (5) Effect on shortening the seedling cycle: According to observation and statistics, the seedlings of the treatment group with the H501 strain described in this invention reached the growth state that can be released into the sea about 3 to 5 days earlier than the control group.

[0046] Example 5: Whole genome sequencing and functional gene mining of Bacillus velezensis H501 as described in this invention. 5.1 Genome Sequencing and Assembly Genomic DNA from bacterial strain H501 was extracted using a bacterial genomic DNA extraction kit. After passing quality control, the subsequent multi-omics library construction and high-throughput sequencing were entrusted to Tianjin Yunsitop Biotechnology Co., Ltd. The specific procedures were as follows: DNA fragmentation was performed using sonication; short insert (500 bp) paired-end libraries were constructed using the Illumina NextEra DNA library construction kit; simultaneously, a large insert (approximately 20 kb) library was constructed using the SMRTbell Express Template Prep Kit 2.0. In addition, chromatin conformation capture (Hi-C) libraries and RNA-Seq transcriptome libraries were constructed to aid assembly and gene prediction. All these libraries were ultimately sequenced using both the Illumina X-TEN and PacBioSequel IIe platforms for high-throughput sequencing.

[0047] After adapter removal, the raw sequencing data underwent quality trimming and rigorous quality control using Trimmomatic software (parameters set to 4-bp sliding window and an average quality threshold of 15 per base). High-quality clean reads were then assembled and used for genome mapping visualization with Circos v0.66 software. Average nucleotide identity (ANI) analysis was performed using the JSpeciesWS online platform.

[0048] Sequencing results show that the genome of the *Bacillus belyssus* H501 is as follows: Figure 5 As shown in Figure A, this is a circular chromosome measuring 3,941,468 bp with a GC content of 46.62%. Genetic prediction indicates that this genome contains 3,785 protein-coding sequences (CDS), 86 tRNA genes, and 27 rRNA genes. The ANI value is shown below. Figure 5 As shown in B, there are obvious gradient differences between species and subspecies: the distribution among species is 84.31%–88.67%, while that among subspecies is 97.55%–97.73%.

[0049] 5.2 Analysis of gene clusters for secondary metabolite synthesis The genome of strain H501 was analyzed using the antiSMASH online software to predict gene clusters for secondary metabolite synthesis. The results showed that 12 gene clusters for secondary metabolite synthesis were identified within the genome (see Table 1 for details).

[0050] The core gene clusters include: surfactant synthesis gene cluster (82% similarity), macrolide synthesis gene cluster (80% similarity), bacillaene synthesis gene cluster (100% similarity), difficidin synthesis gene cluster (92% similarity), siderophore synthesis gene cluster (100% similarity), and andalusicin synthesis gene cluster (82% similarity), etc.

[0051] It is noteworthy that, in addition to the known biosynthetic gene clusters mentioned above, the antiSMASH analysis also identified several unknown gene clusters that showed no significant sequence similarity to known public databases (such as MIBiG) (Table 1). The presence of these unknown gene clusters (such as uncharacterized T3PKS or terpene-like gene clusters) indicates that the *Bacillus belyssiensis* H501 described in this invention possesses a significant genetic potential to synthesize novel, unknown secondary metabolites or antimicrobial substances. This unique genetic background not only explains, at the molecular level, the superior and broad-spectrum pathogen antagonism exhibited by this strain in the examples, but also further highlights the uniqueness and high inventiveness of the strain described in this invention compared to existing conventional *Bacillus* species.

[0052] The various broad-spectrum antimicrobial lipopeptides, polyketides, and highly efficient siderophores encoded by the above-mentioned gene clusters provide a solid molecular genetic basis for the specific antagonism of pathogens such as Aeromonas hydrophila by the H501 strain described in Example 3.6.

[0053] 5.3 Discovery of functional genes related to growth promotion and environmental adaptation The H501 genome was further identified by comparing and annotating it using the KEGG pathway database, revealing a core gene pool closely related to the strain's growth-promoting function and environmental adaptability. (1) Phosphorus solubility and phosphate transport system ( Figure 6 (Part A): Complete pstS, pstC, pstA, and pstB gene operators were identified in the genome, encoding a high-affinity phosphate transport system. This molecular characteristic is perfectly consistent with the excellent organophosphate solubility of the strain described in Example 3.2.

[0054] (2) Nitrogen fixation-related pathways: Key genes such as nifU were identified in the genome, which encode core proteins involved in the assembly of nitrogenase [Fe-S] clusters. The discovery of these genes explains, at the molecular level, the nitrogen fixation and efficient nitrogen utilization characteristics of the strain described in Example 3.1 grown on nitrogen-free medium.

[0055] (3) Ferrocarrier synthesis and transport ( Figure 6C): In addition to identifying the aforementioned bacillibactin siderophore synthesis gene cluster ( Figure 6 The diagram (C) shows the complete pathway from branched acid to siderophore catalyzed by enzymes such as DhbC, DhbB, DhbE, and DhbF. A complete iron transport-related gene cluster (fur, fetA, fetB, yclN, yclO, yclP, yclQ) was also identified. This corroborates the results of the CAS plate colorimetric reaction in Example 3.3, confirming its extremely strong competitive iron ion acquisition ability.

[0056] (4) Salt tolerance and osmotic pressure regulation system ( Figure 6 (B in the text): Abundant Na was identified in the genome. + / H + The presence of the reverse transporter genes (nhaC, mnhABCDEFG), as well as genes for the synthesis of osmoprotective substances such as betaine (gbsB, betB) and proline synthesis (rocF, rocD), provides direct molecular evidence for the survival of this strain at 7% salinity in Example 2.2 and its macroscopic phenotype of colonizing and promoting kelp growth in a seawater environment in Example 4.

[0057] Based on the above genomic analysis results, the Bacillus belyssus H501 described in this invention possesses a complete molecular regulatory network of "growth promotion-disease prevention-stress resistance," which not only confirms its feasibility for application in marine and agricultural microecological preparations from a mechanistic perspective, but also highlights the great development value of this strain as a novel biocontrol chassis cell.

[0058] Table 1. Gene clusters for secondary metabolite synthesis in the genome of Bacillus belyssus H501 ; .

[0059] The above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit them. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions claimed by the present invention.

Claims

1. A type of Bacillus velezensis H501, Latin name Bacillus velezensis H501, characterized in that, The strain was deposited on December 24, 2025, at the China General Microbiological Culture Collection Center (CGMCC), with accession number CGMCC NO. 37160.

2. The Bacillus belyssus H501 according to claim 1, characterized in that, The strain possesses at least one of the following biological characteristics: a) It can form a transparent phosphorus-solubilizing zone on a culture medium in which lecithin is the only phosphorus source; b) It can produce an orange-brown chelation ring on the Chromium Blue S detection plate; c) It has an antagonistic effect against Aeromonas hydrophila at 20℃; d) It does not have alginate lyase activity.

3. The Bacillus belyssus H501 according to claim 1, characterized in that, The genome of the strain contains a gene cluster encoding at least one of the following secondary metabolites: surfactant, macrolide H, sporeene, interferon, dapoxetine, siderophores, and lysozyme.

4. The use of Bacillus belyssus H501 as described in claim 1 in the preparation of an inhibitor of Aeromonas hydrophila.

5. The application of Bacillus vesiculosus H501 as described in claim 1 in promoting the growth of kelp seedlings.

6. A method for promoting the growth of kelp seedlings, characterized in that, The method includes the step of adding the bacterial suspension of Bacillus vesicle H501 as described in claim 1 to the kelp seedling system.

7. The method for promoting the growth of kelp seedlings according to claim 6, characterized in that, The final concentration of Bacillus vesiculus H501 in the seedling system was 4 × 10⁻⁶. 7 CFU / mL up to 8×10 7 CFU / mL.

8. A microbial preparation, characterized in that, The microbial preparation comprises Bacillus belyssus H501 as described in claim 1, and an agriculturally or aquaculture-acceptable carrier.

9. A microbial preparation according to claim 8, characterized in that, The formulation is available in liquid, solid, or powder form.

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