Whole-Genome Analysis Reveals the Growth-Promoting and Biocontrol Potential of Bacillus amyloliquefaciens Ba. YN. J3 isolated from Avena sativa

Whole-Genome Analysis Reveals the Growth-Promoting and Biocontrol Potential of Bacillus amyloliquefaciens Ba. YN. 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YN. J3 isolated from Avena sativa Wei Quan, ChengZhong Zheng, Ayaz Muhammad, Hui Chen, ChunYang Wang, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8097513/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 13 You are reading this latest preprint version Abstract Background Endophytic bacteria serve as important resources for the development of novel biocontrol agents for sustainable agriculture. The present study provides a detailed characterization of a newly isolated oat endophyte, Bacillus amyloliquefaciens Ba. YN. J3, using an integrated analysis of phenotypic, genomic, and comparative genomic data to uncover its biocontrol and plant growth-promoting (PGP) potential. Results The current findings indicate that Ba. YN. J3 possessed efficient PGP and biocontrol potential both in vitro and in planta . Additionally, Ba. YN. J3 showed broad-spectrum antifungal activity against six major phytopathogens and was found to produce multiple cell wall-degrading enzymes (CWDEs) and siderophores, significantly increasing the growth of several crop species and regulating defense enzymes in oats. The complete 4.06 Mb genome of Ba. YN. J3 contains numerous gene clusters encoding vital secondary metabolites (e.g., surfactin, fengycin), CWDEs, and proteins associated with PGP functions and chemotaxis. The genome also harbors a robust set of genes related to abiotic stress tolerance, suggesting its potential to survive and function effectively in challenging field environments. Furthermore, comparative genomic analysis revealed 830 strain-specific genes, including two unique gene families critically linked to chemotaxis (flagellar rod protein FlgC) and nitrogen fixation (regulatory protein YutI). Conclusions This integrated study elucidates the potent dual function of Ba. YN. J3 and its unique genetic determinants. The flgC and yutI gene families, in particular, offer novel insights into the molecular basis of its targeted antagonism and nutritional self-sufficiency. Hence, the current findings highlight Ba. YN. J3 as a promising candidate for the development of effective biopesticides and biofertilizers. Bacillus amyloliquefaciens Endophyte Biocontrol Plant growth promotion Whole-genome sequencing Comparative genomics Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 1. Background Microorganisms residing within plant tissues and the rhizosphere play crucial roles in plant development by suppressing pathogens and improving nutrient availability [ 1 , 2 ]. The plant endophytes are a unique group of microbes that colonize the internal tissues of plants without causing apparent harm to their host [ 3 ]. Previous studies have shown that endophytes can promote plant growth by producing phytohormones, such as auxin and gibberellin, or by acting as biofertilizers through mechanisms such as nitrogen fixation and solubilization of phosphate and potassium [ 4 , 5 ]. Additionally, certain endophytes can significantly suppress the infection and spread of plant pathogens, thereby protecting plants from disease. Given their dual role in growth promotion and disease control, endophytes have garnered considerable attention as promising tools in sustainable agricultural production. Beneficial agricultural bacteria promote plant growth through two primary mechanisms: the production of phytohormones and the increase in nutrient availability [ 6 ]. Phytohormones such as auxin and gibberellin, which are produced by many bacterial strains, have been shown to facilitate plant growth and improve tolerance to abiotic stresses such as high salinity [ 7 , 8 ]. The synthesis of indole-3-acetic acid (IAA), a key auxin, is often governed by critical gene clusters such as the trp operon [ 9 ]. For example, the well-studied rhizobacteria B. amyloliquefaciens SQR9 and B. thuringiensis RZ2MS9 produce IAA, a trait attributed to biosynthetic genes, including trp and ipdC [ 10 , 11 ]. In addition to phytohormone production, the nutrient supply provided by endophytic and rhizosphere bacteria is increased through processes such as biological nitrogen fixation and solubilization of phosphorus and potassium. These functions are linked to specific genetic determinants; for example, the nif , rpo , and ntr genes are involved in nitrogen fixation, whereas genes such as pho , glt , and kdp are associated with phosphorus and potassium solubilization, respectively [ 12 ]. Antagonism, the direct inhibition of plant pathogens, is a major biocontrol mechanism employed by beneficial bacteria [ 12 , 13 ]. This inhibition is often mediated by a diverse array of bioactive secondary metabolites, including lipopeptides, siderophores, and bacteriocins [ 14 ]. Lipopeptides, a prominent class of these metabolites, include compounds such as surfactin, iturin, and fengycin, which are synthesized by large enzymatic complexes called nonribosomal peptide synthetases (NRPSs) [ 15 ]. The biosynthesis of these molecules is governed by extensive gene clusters; for example, the production of fengycin is directed by the fen gene cluster, which encodes five large peptide synthetases [ 16 ]. Siderophores represent another critical class of antagonistic compounds. These low-molecular-weight molecules are secreted to chelate ferric iron from the environment, thereby limiting its availability to competing pathogens [ 17 ]. The pseudobactin siderophore, for example, is a key factor in the biocontrol of Botrytis cinerea by Pseudomonas spp., and its production is regulated by gene clusters such as iucABCD [ 18 , 19 ]. Successful colonization of host tissues is a prerequisite for the beneficial functions of endophytes. This process often begins with chemotaxis, where microorganisms recognize compounds secreted by the plant and move toward its surface before entering through natural openings, wounds, or direct penetration [ 20 ]. Chemotactic activity, alongside the secretion of cell wall-degrading enzymes (CWDEs), is therefore key mechanisms facilitating endophytic colonization. Numerous studies have documented this phenomenon in various endophytic bacteria, such as B. amyloliquefaciens SQR9, P. putida KT2440, and Rhizobium leguminosarum N5 [ 21 – 24 ]. The specific chemoattractants identified include a range of sugars and organic acids that guide the targeted movement of B. cereus YL6 [ 25 ]. Similarly, fatty acids enhance the motility of B. flexus KLBMP 4941 [ 26 ], whereas malic acid, glucose, and fructose promote the colonization of B. velezensis S3-1 [ 27 ]. This chemical attraction is a conserved ecological strategy, as genera, including Azospirillum , Pseudomonas , Serratia , and Enterobacter , also display chemotactic responses to host-derived compounds, highlighting the importance of rhizosphere colonization across diverse plant-associated bacteria [ 28 – 30 ]. Whole-genome sequencing and comparative genomics are powerful tools for uncovering the genetic basis of the diverse traits observed among biocontrol bacteria. This approach can reveal the key genetic determinants underlying their biocontrol and plant growth-promoting functions. For example, a whole-genome analysis of the sugarcane endophyte P. aeruginosa B18 revealed its genetic potential for nitrogen fixation, phosphorus solubilization, and the production of IAA, siderophores, and antibacterial compounds [ 31 ]. Similarly, the genome of the endophyte B. velezensis K1 contains genes associated with the induction of plant resistance, the production of phytohormones, nitrogen fixation, phosphate solubilization, and colonization [ 32 ]. Furthermore, comparative genomic analysis employs a suite of methods to elucidate the genetic basis of phenotypic diversity among bacteria. At a fine scale, analyses of single nucleotide polymorphisms (SNPs), insertions/deletions (indels), and structural variations (SVs) are used to resolve genomic differences, providing insights into genetic diversity and linking specific variations to functional traits [ 33 , 34 ]. At the gene content level, pangenome analysis identifies the core genome shared by a group of strains, as well as the accessory and strain-specific genes that may confer unique adaptations. The comparison of gene family evolution can further illuminate the distinct biocontrol functions of different strains [ 35 ]. These identified genetic variations can ultimately be correlated with phenotypic traits through association analyses to pinpoint the molecular basis of specific activities. The oat endophyte B. amyloliquefaciens YN-J3 (hereafter Ba. YN. J3) has previously been shown to exert a significant biocontrol effect against oat anthracnose in both greenhouse and field assays [ 36 ]. Further studies have demonstrated its capacity for targeted chemotaxis toward fungal mycelia and its ability to inhibit spore germination and appressorium formation in Colletotrichum cereale [ 37 ]. Building on these findings, the present study was designed to comprehensively characterize the multifaceted potential of Ba. YN. J3. We evaluated its broad-spectrum antifungal activity, production of cell wall-degrading enzymes (CWDEs), plant growth-promoting (PGP) traits, and ability to induce host plant resistance. To elucidate the molecular mechanisms underlying these beneficial properties, we subsequently performed whole-genome sequencing and comparative genomic analyses. The primary objective was to identify the key genes and gene clusters responsible for its biocontrol and PGP functions, with a specific focus on identifying unique genetic determinants related to chemotaxis and the suppression of appressorium formation. 2. Methods 2.1 Microbial strains and culture conditions The endophytic bacterium Ba. YN. J3 was previously isolated from the stems of oat ( Avena sativa cv. Baiyan No. 2). For long-term storage, stock cultures were maintained at -80°C in Luria–Bertani (LB) broth containing 30% (v/v) glycerol. For routine experiments, Ba. YN. J3 was cultured in LB medium at 28°C with shaking [ 38 ]. The fungal pathogens used in this study included C. cereale , Sclerotinia sclerotiorum , Drechslera glomerata , Fusarium oxysporum , Rhizoctonia solani , and Alternaria alternata . The fungal stock cultures were stored under the same conditions as the bacterial cultures. For experimental use, mycelial plugs from the frozen stocks were inoculated onto the center of potato dextrose agar (PDA) plates and incubated at 25°C for 7–14 days to obtain fresh cultures for subsequent assays. 2.2 In vitro antifungal activity assays Dual-culture assay: Antagonistic activity was evaluated via a dual-culture plate assay. A 5-mm mycelial plug from an actively growing fungal culture was placed at the center of a potato dextrose agar (PDA) plate. A 24-h-old culture of Ba. YN. J3 was then streaked in a line 2.5 cm from the plug. The plates inoculated with only the fungal plug served as the control. After incubation at 25°C for 7–14 days, the fungal colony radius was measured for both the control (Rc) and treatment (Rt) plates. The percentage of growth inhibition was calculated as follows: Inhibition (%) = [(Rc − Rt)/Rc] × 100 [ 39 ]. Indirect Inhibition Assay: The effects of volatile organic compounds (VOCs) were assessed via the sealed double-dish method [ 40 ]. A PDA plate was centrally inoculated with a 5-mm fungal plug. This plate was then inverted over a second plate, which was subsequently streaked with Ba. YN. J3. The pairs of plates were sealed together with Parafilm and incubated at 25°C for 7–14 days. The control consisted of a fungal plate paired with an uninoculated LB agar plate. Fungal growth inhibition was calculated as described above. Non-Volatile Metabolite Assay: The effects of nonvolatile metabolites were evaluated via the use of cell-free supernatants [ 41 ]. The Ba. YN. J3 was cultured in LB broth at 37°C with shaking (180 rpm) for 48 h. The culture was centrifuged (12,000 × g , 15 min), and the supernatant was sterilized by filtration through a 0.22-µm membrane. This cell-free supernatant was incorporated into molten PDA (previously cooled to ~ 50°C) to a final concentration of 10% (v/v). The amended PDA was poured into plates, which were then centrally inoculated with 5-mm fungal plugs and incubated at 25°C for 7–14 days. PDA plates supplemented with sterile LB broth served as the control. Fungal growth inhibition was calculated as described above. 2.3 Abiotic stress tolerance assays The tolerance of Ba. YN. J3 to various abiotic stresses was determined by measuring its growth (OD600) in LB broth under different conditions [ 42 , 43 ]. For all the assays, 200 µL of an overnight Ba. YN. J3 culture was inoculated into 20 mL of the respective test medium. Temperature tolerance: Cultures were incubated at 10, 15, 20, 25, 30, 35, 40, or 45°C with shaking (180 rpm). After 24 h, growth was quantified by measuring the OD₆₀₀. pH tolerance: Cultures were grown at 28°C in LB broth adjusted to pH values of 4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10.0, or 11.0. After 24 h, growth was quantified by measuring the OD₆₀₀. Salinity tolerance: Cultures were grown at 28°C in LB broth supplemented with NaCl to final concentrations of 1, 3, 5, 7, 9, or 11% (w/v). After 24 h, growth was quantified by measuring the OD₆₀₀. Drought tolerance: Drought stress was simulated via LB broth supplemented with PEG6000 at concentrations of 1, 3, 5, 7, 9, 11, 13, or 15% (w/v). Cultures were incubated at 28°C, and growth was quantified by measuring the OD₆₀₀ after 48 h. 2.4 Screening for Plant Growth Promotion (PGP) and Enzymatic Activities The PGP and enzymatic activities of Ba. YN. J3 were assessed via qualitative plate assays. Nitrogen Fixation: Nitrogen fixation ability was assayed by observing the growth of Ba. YN. J3 on Ashby's nitrogen-free medium [ 44 ]. Nutrient solubilization: Phosphate and potassium solubilization activities were evaluated by the formation of clearing zones (halos) around colonies grown on NBRIP medium and Alexandrov medium, respectively [ 45 , 46 ]. Siderophore Production: Siderophore production was detected by the formation of a pale-yellow halo on Chrome Azurol S (CAS) agar plates [ 47 ]. Hydrolytic Enzyme Production: The production of various hydrolytic enzymes was screened by observing hydrolysis halos on agar plates supplemented with specific substrates. A positive result was indicated by a clear zone around the colony after 2–5 days of incubation at 28°C [ 48 , 49 ]. The substrates used were as follows: 1% (w/v) sodium carboxymethyl cellulose (for cellulase), 1% (w/v) skim milk powder (for protease), 1% (w/v) soluble starch (for amylase), 1% (w/v) apple pectin (for pectinase), 0.5% (w/v) Poria cocos powder (for β-1,3-glucanase), and 0.5% (w/v) colloidal chitin (for chitinase). 2.5 In Planta Growth Promotion Assays The Ba. YN. J3 was cultured in LB broth (28°C, 180 rpm, 24 h). The bacterial cells were harvested by centrifugation (5,000 × g, 10 min), washed once with sterile water, and finally resuspended in sterile water to an optical density at OD₆₀₀=1.0. This stock suspension was then diluted 10-fold with sterile water for inoculation. For the plant growth conditions and treatment, seeds of corn ( Zea mays cv. Xianyu 335), wheat ( Triticum aestivum cv. Longmai 21), sunflower ( Helianthus annuus cv. SH361), and tomato ( Solanum lycopersicum cv. Antles) were surface sterilized and germinated. Uniform seedlings at the two-leaf stage were transplanted into pots containing a 2:1 (v/v) mixture of nutrient-rich soil and field soil. Each seedling was then inoculated via root drenching with 50 mL of the diluted Ba. YN. J3 suspension. The control plants received an equal volume (50 mL) of sterile water. The plants were maintained in a greenhouse under a 16 h/8 h light/dark cycle at 26°C [ 50 ]. After 21 days, various growth parameters, including plant height, root length, root fresh weight, root dry weight (biomass), and leaf chlorophyll content, were measured. 2.6 Defense-Related Enzyme Assays in Oats For detect Bacterial suspensions of Ba. YN. J3 were prepared as described in Section 2.7 (final OD₆₀₀ diluted to 0.1). Uniform oat seedlings ( Avena sativa cv. Baiyan No. 2) at the two-leaf stage were used for the assay. Plants in the treatment group were inoculated by foliar spraying with the Ba. YN. J3 suspension until the leaves were thoroughly wet. Control plants were sprayed with an equal volume of sterile water. The plants were maintained in the greenhouse under conditions described in Section 2.7 . Leaf samples were collected at various time points post-spraying (hps), immediately frozen in liquid nitrogen, and stored at -80°C for analysis. The activities of four defense-related enzymes—superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and polyphenol oxidase (PPO)—were measured using commercial assay kits (Boxbio, Beijing, China) according to the manufacturer’s instructions. Enzyme activities were quantified using a spectrophotometer. 2.7 Genome Sequencing, Assembly, and Annotation The genomic DNA was extracted from an overnight culture of Ba. YN. J3 (grown in LB broth at 28°C with shaking at 180 rpm) using the STE buffer method. The quality and concentration of the extracted DNA were verified via 1% agarose gel electrophoresis and a Qubit fluorometer (Thermo Fisher Scientific, USA), respectively [ 51 ]. Whole-genome sequencing was performed by Novogene Co., Ltd. (Beijing, China) on the Pacific Biosciences (PacBio) Sequel platform via single-molecule real-time (SMRT) technology [ 52 ]. After filtering to remove low-quality data, the resulting high-quality reads were assembled de novo via Canu (v2.0). This process generates a single, complete, and gapless contig for the chromosome [ 53 ]. Genome annotation began with the prediction of various components. Coding sequences (CDSs) were predicted via GeneMarkS (v4.17) [ 54 ]. Repetitive elements were identified via RepeatMasker (v4.0.5) for interspersed repeats and Tandem Repeats Finder (v4.07b) for tandem repeats [ 55 ]. Noncoding RNAs (ncRNAs), including tRNAs and rRNAs, were predicted via tRNAscan-SE (v1.3.1) and rRNAmmer (v1.2), respectively [ 56 ], whereas other ncRNAs were identified via searches of the Rfam database [ 57 , 58 ]. Finally, mobile genetic elements such as genomic islands, prophages, and CRISPR arrays were identified via IslandPath-DIOMB (v0.2), phiSpy (v2.3), and CRISPRdigger (v1.0), respectively [ 59 – 61 ]. The functional annotation of the predicted genes was performed by aligning their sequences against the KEGG [ 62 , 63 ], SwissProt, and Pfam databases. Specific attention was given to pathways related to nitrogen fixation, phosphate/potassium solubilization, siderophore biosynthesis, and hydrolytic enzymes. Gene clusters for secondary metabolite biosynthesis were identified via antiSMASH (v4.0.2) [ 64 ], and carbohydrate-active enZymes (CAZymes) were annotated via the dbCAN database [ 65 ]. 2.8 Comparative Genomic Analysis For the comparative genomic analysis, the genome of Ba. YN. J3 was compared against nine other Bacillus and Priestia strains selected for its known or potential biocontrol properties (detailed in Table 5 ). The whole-genome alignments and synteny analyses were performed via MUMmer and LASTZ. These alignments were then used to identify single nucleotide polymorphisms (SNPs), insertions/deletions (indels), and structural variations (SVs) [ 66 , 67 ]. To assess evolutionary relationships, a phylogenetic tree was constructed via PhyML with 1,000 bootstrap replicates on the basis of the concatenated alignment of single-copy core genes. Pangenome analysis was conducted via CD-HIT (50% identity, 70% coverage thresholds) to identify core and strain-specific gene sets. Finally, key genomic features of Ba. YN. J3 were visualized on a circular map generated with Circos (v0.66). Table 5 Strain information required for comparative genome analysis Strain name Strain name in report Genebank accession number B. amyloliquefaciens YN.J3 Ba. YN. J3 PRJNA1171741 B. amyloliquefaciens GKT04 Ba.GKT04 CP072120.1 B. amyloliquefaciens DSM 7 = ATCC 23350 Ba.DSM7 FN597644.1 B. amyloliquefaciens ZKY01 Ba.ZKY01 CP044132.1 B.velezensis FZB42 Bv.FZB42 CP000560.2 B. subtilis subsp. subtilis str. 168 Bs.168 AL009126.3 Priestia megaterium NBRC 15308 = ATCC 14581 Pm.14581 CP035094.1 B. cereus FORC_047 Bc.FORC.047 CP017060.1 B. halotolerans ZB201702 Bh.ZB201702 CP029364.1 B. pumilus SAFR-032 Bp.SAFR.032 CP000813.4 2.9 Statistical analysis All experimental data were presented as the mean ± standard deviation (SD) of at least five biological replicates. One-way analysis of variance (ANOVA) was used to determine statistical significance. Mean separation was performed via Duncan's multiple range test at a significance level of p < 0.05. All the statistical analyses were conducted via SPSS Statistics v26.0 (IBM Corp., Armonk, NY, USA). 3. Results 3.1 Ba. YN. J3 Exhibits Broad-Spectrum Antifungal Activity The biocontrol potential of Ba. YN. J3 against six major phytopathogenic fungi were evaluated. In dual-culture assays, Ba. YN. J3 significantly inhibited the mycelial growth of C. cereale , S. sclerotiorum , D. glomerata , F. oxysporum , R. solani , and A. alternata (Fig. 1 ; Table 1 ). To determine the mechanisms of this inhibition, the effects of volatile organic compounds (VOCs) and nonvolatile metabolites were assessed separately. Both VOCs and nonvolatile metabolites significantly suppressed the mycelial expansion of five of the tested pathogens, with inhibition rates ranging from 44.33% to 80.02% (Table 1 ). Notably, for C. cereale , D. glomerata , F. oxysporum , and A. alternata , the inhibitory effects of the nonvolatile metabolites were significantly stronger than those of the VOCs. Although the colony diameter of R. solani was not significantly reduced by either treatment, its aerial mycelia were visibly thinner than those of the control (Fig. 1 ). These results indicate that Ba. YN. J3 possesses broad-spectrum antifungal activity, which is mediated by both volatile and nonvolatile compounds that inhibit the polar growth and branching of fungal hyphae. Table 1 The inhibition rates of Ba. YN. J3 against several pathogens Strains Inhibition rate/% Dual culture Volatile substance Non-Volatile substance C. cereale 71.65 ± 0.64 b 65.86 ± 1.22 a 78.29 ± 1.02 c S. sclerotiorum 48.13 ± 0.74 c 46.01 ± 0.39 b 44.33 ± 0.51 a D. glomerata 55.13 ± 0.28 b 51.72 ± 0.88 a 66.08 ± 0.77 c F. oxysporum 62.17 ± 0.35 b 58.57 ± 0.78 a 62.79 ± 0.78 b R. solani 48.09 ± 0.74 b 1.84 ± 0.53 a 2.37 ± 0.70 a A. alternata 61.01 ± 0.95 b 51.73 ± 1.54 a 80.02 ± 0.70 c 3.2 Ba. YN. J3 Secretes Different Cell-wall-Degrading Enzymes To investigate the biocontrol mechanism of Ba. YN. J3 against C. cereale , the interaction between the bacterium and fungal conidia was observed. Following cocultivation, the conidia presented severe morphological abnormalities, including cytoplasmic leakage, suggesting that Ba. YN. J3 causes defects in the fungal cell wall (Fig. 2 ). On the basis of this observation, we hypothesized that Ba. YN. J3 secretes extracellular cell wall-degrading enzymes (CWDEs). Subsequent plate assays confirmed this, as translucent hydrolysis zones were observed on media containing substrates for cellulase, protease, amylase, pectinase, and chitinase (Fig. 3 E-G, I, K). In addition, Ba. YN. J3 also tested positive for siderophore production (Fig. 3 H). Further experiments revealed that Ba. YN. J3 could utilize colloidal chitin as the sole carbon source, confirming chitinase activity, but failed to grow on a medium with colloidal β-1,3-glucan, indicating that it did not produce detectable β-1,3-glucanase under these conditions (Fig. 3 J and 3 K). Collectively, these results demonstrate that Ba. YN. J3 secretes a diverse array of lytic enzymes capable of disrupting the fungal cell wall. 3.3 Environmental adaptability and abiotic stress tolerance of Ba. YN. J3 To evaluate its potential for field application, the environmental adaptability of Ba. YN. J3 to various abiotic stresses was assessed (Fig. 4 ). The strain demonstrated broad temperature tolerance, growing effectively between 10°C and 45°C, with optimal growth observed at 30°C. It also tolerates a wide pH range from 5.0–9.0, although growth is significantly inhibited under more strongly acidic (pH 9.0) conditions. Furthermore, Ba. YN. J3 exhibited tolerance to salinity and osmotic stress. Its growth gradually decreased with increasing NaCl concentration, showing a sharp decline only when the concentration exceeded 7% (w/v). Similarly, in the presence of PEG6000 to simulate drought, the strain growth decreased only slightly up to a concentration of 11% (w/v), beyond which a sharp decline occurred. Collectively, these results indicate that Ba. YN. J3 is well adapted to a wide range of temperatures and pH values and is tolerant to moderate levels of salinity and osmotic stress. 3.4 Ba. YN. J3 promotes the growth of four crop species Previous studies have shown that Ba. YN. J3 promotes oat seedling growth through mechanisms such as IAA synthesis and nutrient provision [ 36 ]. To determine whether this effect extends to other crops, its growth-promoting activity was further evaluated in sunflower, corn, wheat, and tomato. Compared with the uninoculated controls, Ba. YN. J3 treatment significantly increased shoot length, root length, root fresh weight, and root dry weight in all tested species. A significant increase in chlorophyll content was observed only in corn and sunflower, indicating host-specific effects on photosynthetic capacity (Fig. 5 ). The most pronounced growth enhancement was found in corn, followed by sunflower, tomato, and wheat, suggesting that Ba. YN. J3 possesses broad-spectrum plant growth-promoting potential. 3.5 Ba. YN. J3 induces systemic disease resistance in oats We next investigated whether Ba. YN. J3 could induce systemic resistance in oat plants. The activities of four key defense-related antioxidant enzymes—superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and polyphenol oxidase (PPO)—were measured at multiple time points after spraying (hps) and compared with those of a sterile water-treated control (Fig. 6 ). Ba. YN. J3 treatment significantly enhanced the activities of all four enzymes (p < 0.05). In treated seedlings, enzyme activities progressively increased, reached their maximum at 36 hps, and then slightly declined, yet remained significantly higher than those in the control group throughout the experiment. In contrast, enzyme activities in control plants showed negligible variation over time. These findings demonstrate that Ba. YN. J3 effectively activates the antioxidant defense system in oats, thereby enhancing their systemic resistance to stress and potential pathogen attack. 3.6 Genomic features of Ba. YN. J3 3.6.1 General Genome and Plasmid Features To elucidate the molecular mechanisms underlying its beneficial activities, the complete genome of Ba. YN. J3 was sequenced. The genome consists of a single circular chromosome of 4,063,196 bp with a GC content of 46.27% and one circular plasmid of 215,473 bp with a GC content of 37.24% (Fig. 7 ; Table 2 ). A total of 4,541 protein-coding genes were predicted, accounting for 90.04% of the genome. In addition, the genome contains 124 noncoding RNA genes (including 27 rRNAs, 87 tRNAs and 10 sRNAs), 15 CRISPR arrays, 5 genomic islands, and 23 pseudogenes (Table 2 ). The complete genome sequence has been deposited in the GenBank database under accession number PRJNA1171741. Table 2 The general genome features of Ba. YN. J3 Feature Vaule Genome size (bp) 4,063,196 G + C content 46.27% Topology Circular Plasmid 1 Total number of genes 4665 Total size of protein-coding genes 3,852,390 bp Protein-coding genes 4541 Average CDs size (bp) 848 rRNA number (total) 27 tRNA number 87 sRNA number 10 Repetitive sequence(bp) 35,897(0.839%) CRISPR 15 Prophage 10 Pseudogenes 23 Gls 5 Gene cluster Genes assigned to NR Genes assigned to GO Genes assigned to KEGG Genes assigned to Pfam Genes assigned to Swiss-Prot Genes assigned to CAZy 13 4413 2865 4130 2865 3396 161 3.6.2 Functional Annotation and COG Classification The functional annotation against multiple databases (including NR, GO, KEGG, and SwissProt) assigned functions to a majority of the predicted genes. Analysis of Clusters of Orthologous Groups (COGs) revealed that the most abundant categories were amino acid transport and metabolism (298 genes), transcription (289 genes), and carbohydrate transport and metabolism (248 genes). Significant numbers of genes were also assigned to categories related to cell wall/membrane biogenesis and signal transduction, reflecting the strain's active interaction with its environment (Fig. 7 C). 3.6.3 Genetic basis for biocontrol activities Consistent with its observed antifungal activity, the genome of Ba. YN. J3 harbors a rich repertoire of genes associated with biocontrol. Analysis with antiSMASH identified thirteen secondary metabolite biosynthesis-related gene clusters (BGCs), including those for the synthesis of surfactin, fengycin, bacillibactin, and difficidin (Table 3 ). Furthermore, the genome contains 286 genes encoding putative hydrolytic enzymes, such as chitinases and proteases, which are implicated in fungal cell wall degradation (Table S1 ). The genome also harbors 6 genes for siderophore production and 40 genes related to chemotaxis, providing a genetic basis for its iron competition and targeted motility capabilities (Table 4 ). Table 3 Predictive gene clusters involved in the synthesis of secondary metabolites in Ba. YN. J3 Region of genome Most similar known cluster From To Type Productions Similarity Resources 195,469 273,197 NPRS, transAT-PKS locillomycin/locillomycin B/locillomycin C 35% B.subtilis 916(Luo, 2015) 342,784 408,191 NRPS surfactin 82% B.subtilis ATCC21332(Wei, 2004) 977,079 1,108,323 PKS-like butirosin A/butirosin B 7% B.circulans SANK72073(Kudo, 2005) 1,100,346 1,121,086 terpene 1,239,789 1,268,678 Lanthipeptide-class-ii 1,432,632 1,520,843 transAT-PKS macrolactin H 100% B.amyloliquefaciens NJN-6(Yuan, 2012) 1,739,543 1,849,654 transAT-PKS, T3PKS, NRPS bacillaene 100% B. amyloliquefaciens SQ-2(Li, 2024) 1,905,107 2,042,936 NRPS, transAT-PKS, betalactone fengycin 100% B.subtilis F-29-3(Vanittanakom, 1986) 2,065,505 2,087,388 terpene 2,156,032 2,197,132 T3PKS 2,447,181 2,553,347 transAT-PKS difficidin 100% B.subtilis ATCC39320(Zimmerman, 1987) 3,164,202 3,215,995 NRP-metallophore, NRPS, RiPP-like bacillibactin 100% B.siamensis SCSIO05746(Pan, 2019) 3,728,795 3,770,213 Others bacilysin 100% B.velezensis FZB42(Han, 2021) Table 4 Comparison of genes related to the biocontrol and growth promotion of Ba. YN. J3 and nine reference Bacillus strains Strain name Number of biocontrol associated genes Number of growth-promoting associated genes A B C D E F G H I J Ba. YN. J3 69 286 6 40 105 39 63 5 60 3 Ba.GKT04 12 203 7 37 18 2 61 2 40 0 Ba.ZKY01 27 274 7 36 19 2 85 1 50 3 Bv.FZB42 26 265 12 12 12 2 73 1 49 3 Ba.DSM7 12 269 9 36 19 2 68 2 49 3 Bs.A168 37 224 19 51 81 0 116 1 58 2 Bh.ZB201702 17 274 8 41 25 1 111 3 56 4 Bp.SAFR.032 9 273 8 49 19 2 64 1 49 0 Bc.FORC.047 6 454 9 46 14 0 115 2 62 3 Pm.A14581 5 399 4 33 23 3 102 1 88 5 Note: A: Secondary metabolites, B: Cell wall-degrading enzymes, C: Siderophores, D: Chemotaxis, E: Colonization ability, F: Nitrogen fixation, G: Phosphate solubilization, H: Potassium solubilization, I: Indole-3-acetic acid (IAA) production, J: Amylase production. 3.6.4 Genetic basis for plant growth-promoting (PGP) traits The PGP potential of Ba. YN. J3 is also well supported by its genome. We identified 60 genes in the tryptophan biosynthesis pathway leading to IAA production, 39 genes involved in nitrogen metabolism (including the key genes nasDE and rocG ), 63 genes related to phosphate solubilization (e.g., the pst system), and 5 genes involved in potassium solubilization ( ktr , kdp ) (Table 4 ; Table S2). These genetic features underpin the strain's ability to promote crop growth by producing phytohormones and increasing nutrient availability. 3.6.5 Genetic basis for abiotic stress tolerance The high tolerance of this strain to abiotic stresses is corroborated by the presence of numerous stress response genes (Table S1 ). For thermotolerance, the genome encodes several heat shock proteins (e.g., dnaK , dnaJ , and clpB ). For salinity and osmotic stress, genes for the synthesis and transport of osmoprotectants such as proline and betaine (e.g., proA , betB , and opuA ) were identified. Tolerance to pH fluctuations is supported by the presence of the F₁F₀ -ATPase ( atp ) and Na⁺/H⁺ antiporter ( nhaC ) gene clusters. Additionally, the genome encodes two-component regulatory systems ( comP/comA , degS/degU ) and the alternative sigma factor SigB, which enable rapid adaptation to environmental changes. Overall, this genetic arsenal explains the robust environmental adaptability of Ba. YN. J3. 3.7 Comparative Genomic Analysis of Ba. YN. J3 within nine Bacillus strains To investigate the evolutionary context of Ba. YN. J3, a comparative genomic analysis was conducted against nine representative Bacillus and Priestia strains (Table 5 ). A phylogenetic tree based on whole-genome single-nucleotide polymorphisms (SNPs) revealed that Ba. YN. J3 clustered within a well-supported clade (bootstrap value = 88) together with the well-characterized biocontrol strains B. velezensis FZB42, B. amyloliquefaciens ZKY01, and B. amyloliquefaciens GKT04, indicating a close evolutionary relationship among them (Fig. 8 B). Genome-wide variation analysis further clarified these relationships. Pairwise comparisons revealed extensive SNP divergence between Ba. YN. J3 and the reference genomes, ranging from approximately 2,000 SNPs (vs. B. cereus FORC_047) to more than 189,000 SNPs (vs. B. amyloliquefaciens DSM7) (Fig. 8 A; Table S3). Similarly, the number of insertion/deletion (InDel) events increased with phylogenetic distance, from 27 (vs. B. cereus FORC_047) to 1,194 (vs. B. amyloliquefaciens DSM7) (Fig. 8 C). Structural variation (SV) analysis provided a macroscopic view of genome organization and synteny (Figure S1 ). Strong collinearity was observed between Ba. YN. J3 and its closest relatives (e.g., B. velezensis FZB42), although several local rearrangements such as inversions and translocations were evident. In contrast, comparisons with more distantly related species ( P. megaterium A14581 and B. cereus FORC_047) revealed extensive genomic rearrangements and large-scale segmental losses, consistent with their distinct phylogenetic positions. 3.7 Gene family analysis of Ba. YN. J3 and nine comparative Bacillus strains Gene family analysis was conducted to compare the functional potential of Ba. YN. J3 with nine reference strains. Among the reference genomes, the number of gene families ranged from 2,271 in B. cereus FORC_047 to 2,932 in B. subtilis 168 (Fig. 8 D). The genome of Ba. YN. J3 contained 4,541 genes, of which 3,846 (84.7%) were clustered into 2,836 gene families. A key outcome of this comparative analysis was the identification of strain-specific (unique) gene families. The number of unique gene families among the reference strains ranged from 0 to 184, with B. cereus FORC_047 possessing the most. In contrast, Ba. YN. J3 encoded ten unique gene families. Although eight of these genes could not be functionally annotated, two were identified as being critically associated with key biocontrol and plant growth-promoting (PGP) traits: the flgC gene family, encoding a flagellar basal body rod proteins involved in chemotaxis, and the yutI gene family, which encodes a putative nitrogen fixation regulatory proteins. These unique gene families may therefore contribute to the specific beneficial functions observed in Ba. YN. J3. 3.8 Core‒pan analysis between Ba. YN. J3 and nine comparative Bacillus strains To further explore the genomic basis of the unique traits of Ba. YN. J3, a core-pan analysis was conducted with nine reference Bacillus strains. The analysis identified a core genome of 1,097 genes, which were highly conserved across all ten strains and were primarily involved in essential cellular processes such as primary metabolism, genetic information processing, and environmental adaptation. In addition to the core genome, Ba. YN. J3 possessed 830 strain-specific genes (Fig. 8 E). Of these, only 40 genes (4.82%) could be functionally annotated. Notably, these annotated unique genes were enriched in functions relevant to the strain’s ecological niche, including signal transduction, transport, resistance, and transcriptional regulation, suggesting that these genes may play important roles in the specialized biocontrol and plant-associated lifestyle of Ba. YN. J3. 4. Discussion In this study, we demonstrated that the oat endophyte B. amyloliquefaciens Ba. YN. J3 is a potent strain that exhibits both biocontrol and plant growth-promoting (PGP) functions. Its strong efficacy against oat anthracnose is attributed to multiple synergistic mechanisms, including inhibition of fungal mycelial growth through both volatile (VOCs) and non-volatile metabolites, degradation of fungal cell walls, and induction of host defense enzymes. In addition, Ba. YN. J3 significantly enhanced the growth of multiple crop species. Whole-genome and comparative genomic analyses revealed the genetic basis for these traits, identifying two unique gene families related to chemotaxis ( flgC ) and nitrogen fixation ( yutI ), as well as 40 strain-specific genes associated with plant-associated functions such as signal transduction, transport, resistance, and transcriptional regulation. Together, these findings establish a clear link between the unique genomic repertoire of Ba. YN. J3 and its dual biocontrol and PGP capacities. Members of the genus Bacillus are well-known biocontrol agents that suppress plant pathogens through the secretion of diverse antifungal compounds, which inhibit mycelial growth and spore germination [ 68 ]. The principal molecular mechanisms underlying this antagonism include the production of lipopeptides, siderophores, and cell wall-degrading enzymes (CWDEs) [ 69 , 70 ]. Consistent with this paradigm, Ba. YN. J3 exhibited broad-spectrum antifungal activity, with its metabolites causing fungal cell wall disruption and cytoplasmic leakage. Genomic analyses corroborated these observations, revealing a rich set of genes involved in biocontrol, including those encoding CWDEs, siderophore biosynthetic enzymes, and secondary metabolite biosynthesis. Specifically, thirteen biosynthetic gene clusters (BGCs) were identified, five of which encode nonribosomal peptide synthetases (NRPSs), suggesting that the antifungal activity of Ba. YN. J3 results from the synergistic action of CWDEs, siderophores, and NRPS-derived lipopeptides. Consistent with its strong antagonistic activity, the Ba. YN. J3 genome contains BGCs for the three major families of Bacillus lipopeptides—surfactin, iturin, and fengycin—synthesized by NRPS and PKS complexes. These compounds inhibit phytopathogens such as Botrytis dothidea and Fusarium graminearum by disrupting fungal membranes [ 71 – 74 ]. The surfactin cluster is regulated by quorum-sensing components ComX, ComA/ComP, and RapC [ 75 , 76 ], whereas the iturin A cluster, controlled by DegU and DegQ, directs the synthesis of potent antifungal peptides [ 77 ]. The complete fengycin cluster ( ppsA–E ) further underscores the antifungal potential of this strain. In addition to these canonical clusters, additional BGCs for locillomycin, macrolactin, bacillaene, and difficidin broaden the antimicrobial spectrum of Ba. YN. J3. This diverse array of secondary metabolites provides a strong molecular basis for its biocontrol efficiency. Iron acquisition via siderophore production represents another key mechanism of pathogen suppression [ 78 ]. Ba. YN. J3 produces siderophores, as indicated by clear halos on Chrome Azurol S (CAS) agar, and harbors essential siderophore biosynthetic genes ( fhuA , fhuB , fhuC , fhuD , and fhuG ). Similar siderophore-mediated biocontrol mechanisms have been reported in B. amyloliquefaciens TA-1 [ 79 ] and B. subtilis MBI 600 [ 80 ], as well as in other Bacillus species [ 25 , 36 , 65 ]. These findings collectively confirm that siderophore-mediated iron competition constitutes a critical component of the antagonistic repertoire of Ba. YN. J3. Ba. YN. J3 also promotes plant growth through multiple pathways. Indole-3-acetic acid (IAA) biosynthesis genes ( trpA–P ) were identified in its genome, consistent with previous reports of its IAA-producing potential [ 81 , 82 ]. Furthermore, the strain was capable of growth on nitrogen-free, phosphorus-, and potassium-solubilizing media, reflecting its nutrient-cycling capacity. Genome annotation revealed key genes involved in nitrogen fixation ( yutI , nifU , nifS , nifF , gltB , narK ), nitrogen metabolism ( nasE , nasD , gudB , rocG ), phosphorus solubilization ( pstA , pstB1 , pstB2 , pstC , pstS , ugpC , pykF ), and potassium metabolism ( kimA , ktrC , trkA , kdpD ). Comparative genomics further revealed a unique nitrogen fixation gene family, suggesting that Ba. YN. J3 promotes plant growth through IAA secretion and mobilization of essential nutrients. Ba. YN. J3 also exhibited remarkable tolerance to abiotic stress, thriving under a wide range of conditions, including high temperatures (10–45°C), pH (5.0–9.0), salinity (up to 7% NaCl), and osmotic stress (up to 11% PEG6000). Genomic analysis revealed numerous stress-response genes, including those encoding heat shock proteins (GroEL, DnaK, DnaJ, ClpB), osmotic adaptation proteins (OpuA, ProP, TreB), salt tolerance proteins (ProABC, BetAB, CspABC), and pH homeostasis proteins (AtpABCDEFGH, NhaC) [ 83 – 86 ]. Regulatory elements such as the sigma factor SigB and two-component systems (DegS/DegU and ComP/ComA) [ 87 ] further enhance environmental adaptability by fine-tuning stress-responsive gene expression. Chemotaxis, the directed movement toward chemical stimuli, is essential for host colonization and environmental sensing [ 88 , 89 ]. Previous studies demonstrated that Ba. YN. J3 actively migrates toward Colletotrichum cereale hyphae [ 37 ]. Genomic analysis revealed a complete chemotaxis and flagellar assembly system, including mcp , cheA , cheB , cheC , cheD , cheR , cheW , fliC , fliD , flhA , flhB , flgB–D , and motA/B , suggesting that its robust motility and environmental sensing capacity may underlie host-targeting behavior. Comparative genomics integrating SNP, InDel, SV, and core/pan-genome analyses provided further insights into the genetic basis of strain-specific traits [ 90 ]. Ba. YN. J3 is closely related to well-characterized biocontrol strains such as B. amyloliquefaciens GKT04, B. amyloliquefaciens ZKY01, and B. velezensis FZB42, and shares functional features of antagonism and rhizosphere colonization [ 91 ]. However, its genome also contains 961,698 nonsynonymous mutations affecting key functional genes for lipopeptide synthesis, hormone production ( ipdC ), nutrient solubilization ( phoA , ppk ), nitrogen fixation ( nif ), and siderophore biosynthesis. Such variations are known to modulate metabolite diversity [ 92 ], exopolysaccharide (EPS) formation [ 93 ], motility [ 94 ], and iron acquisition [ 95 ], and are likely to contribute to the unique functional profile and ecological adaptability of Ba. YN. J3 [ 96 ]. Although only 7.4% of the 447 strain-specific genes of Ba. YN. J3 were functionally annotated, these genes underscore its ecological specialization. Many contribute to nutrient acquisition and stress resistance. For example, malL [ 97 ] correlates with strong hydrolytic enzyme activity, enabling efficient degradation of environmental macromolecules. Other unique genes, including ligA [ 98 ], quorum-sensing regulators comP–comA [ 99 ], and small acid-soluble spore proteins (SASPs) [ 100 ], collectively support genome integrity, communication, and long-term survival [ 101 ]. Two experimentally validated unique genes, flgC (chemotaxis) and yutI (nitrogen fixation), define the distinct ecological strategies of Ba. YN. J3. The flgC gene mediates directed motility toward C. cereale spores [ 12 , 102 ], whereas yutI confers the ability to fix nitrogen and grow independently in nutrient-limited conditions [ 103 , 104 ]. These complementary traits equip Ba. YN. J3 with the dual capacity to pursue its pathogenic targets while maintaining nutritional self-sufficiency, underscoring its potential as a robust and environmentally adaptable biocontrol agent. Conclusions This study identifies the oat endophyte B. amyloliquefaciens Ba. YN. J3 as a potent dual-function strain exhibiting both biocontrol and plant growth-promoting (PGP) activities. Phenotypic assays confirmed its broad-spectrum antifungal effects, secretion of multiple cell wall-degrading enzymes (CWDEs), and its ability to enhance the growth of various crop species while inducing systemic resistance. Whole-genome sequencing provided a clear molecular foundation for these traits, revealing a 4.06 Mb circular chromosome enriched with biosynthetic gene clusters for key antifungal secondary metabolites (such as surfactin and fengycin), PGP-related pathways (including IAA biosynthesis and nutrient cycling), and genes conferring tolerance to abiotic stresses. Comparative genomic analysis identified 830 strain-specific genes, including two unique gene families encoding the flagellar rod protein FlgC (involved in chemotaxis) and the nitrogen fixation regulatory protein YutI. These unique genetic determinants provide a mechanistic explanation for the strain’s dual functionality, linking targeted motility to its antagonistic activity and nitrogen fixation to its nutritional self-sufficiency. Moreover, previous greenhouse and field experiments demonstrated that B. amyloliquefaciens Ba. YN. J3 exhibited strong biocontrol efficacy against Colletotrichum cereale , further validating its potential as an effective and reliable biocontrol agent. Collectively, these findings highlight Ba. YN. J3 as a promising and environmentally resilient candidate for the development of sustainable biopesticides and biofertilizers in modern agriculture. Declarations Ethics approval and consent to participate Not applicable. Consent for publication Not applicable. Availability of data and materials The datasets presented in this study can be found in online repositories. The name of the repository and accession number can be found below: NCBI GenBank, accession number: PRJNA1171741 (https://www.ncbi.nlm.nih.gov/bioproject/1171741). Further inquiries can be directed to the corresponding author. All experimental data and materials related to this study are also available from the corresponding author, Bao-zhu Dong, upon reasonable request. Competing interests The authors declare that they have no competing interests. Funding This work was supported by the Science and Technology Program of Inner Mongolia Autonomous Region, “Research and Application of Oat Intertillage Weeding and Chemical Herbicide Reduction and Synergistic Efficiency Enhancement Technology” (Grant No. 2025YFHH0165); the Basic Research Fund for Universities Directly Affiliated with Inner Mongolia Autonomous Region (Grant No. BR251033); the Central Government-Guided Local Science and Technology Development Fund, “Research on Green Production of Oat Grains and High-Quality Compound Feed Processing Technology” (Grant No. 2022ZY0060) and “Application and Promotion of Green Cultivation Technologies for Coarse Cereals in Qingshuihe County” (Grant No. 2022ZY0065); the National Key R&D Program of China, “Research and Demonstration of Green Control Technologies for Diseases, Insect Pests, and Weeds in Minor Grains” (Grant No. 2023YFD1600701-5); and the China Agriculture Research System for Oat and Buckwheat (Grant No. CARS-07-C-3). Authors' contributions Wei Quan: Conceptualization, Methodology, Investigation, Formal Analysis, Writing – Original Draft, Data Curation, Validation. Cheng-Zhong Zheng: Investigation, Formal Analysis. Muhammad Ayaz: Investigation, Software, Visualization, Formal Analysis. Chen Hui: Investigation, Formal Analysis. Chun-Yang Wang: Investigation, Software, Visualization. Chen-Lu Liu: Investigation, Software, Visualization, Formal Analysis, Data Curation, Validation. Zhi-Gang Liu: Software, Visualization, Formal Analysis. Bao-Zhu Dong: Supervision, Project Administration, Funding Acquisition, Conceptualization, Writing – Review & Editing. Hong-You Zhou: Supervision, Project Administration, Funding Acquisition, Conceptualization, Writing – Review & Editing. Acknowledgments We gratefully acknowledge financial support from the Ministry of Agriculture and Rural Affairs, the Ministry of Science and Technology, the Ministry of Education of the People's Republic of China through the Science and Technology Program of the Inner Mongolia Autonomous Region, the Basic Research Fund for Universities Directly Affiliated with Inner Mongolia, the Central Government-Guided Local Science and Technology Development Fund, the National Key R&D Program of China, and the China Agriculture Research System for Oat and Buckwheat. References Wilson D. Endophyte: the evolution of a term, and clarification of its use and definition. Oikos. 1995;104(2):274-276. Doornbos RF, van Loon LC, Bakker PAHM. Impact of root exudates and plant defense signaling on bacterial communities in the rhizosphere: a review. Agron Sustain Dev. 2012;32:227-243. Faeth SH, Fagan WF. Fungal endophytes: common host plant symbionts but uncommon mutualists. 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13:15:19","extension":"png","order_by":21,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":231703,"visible":true,"origin":"","legend":"","description":"","filename":"OnlineFigure2.png","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/d5bd1bb2293f9838fa6e9cb8.png"},{"id":97530389,"identity":"c355c8c2-3552-4af8-90e1-17d7fe692f5a","added_by":"auto","created_at":"2025-12-05 13:15:19","extension":"png","order_by":22,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":570912,"visible":true,"origin":"","legend":"","description":"","filename":"OnlineFigure3.png","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/f4f7030d7f4e8326f180111a.png"},{"id":97530453,"identity":"ecfaf95a-2898-49ff-adcb-42560b417343","added_by":"auto","created_at":"2025-12-05 13:15:21","extension":"png","order_by":23,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":108497,"visible":true,"origin":"","legend":"","description":"","filename":"OnlineFigure4.png","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/ac6cec49ae9a89833e46139c.png"},{"id":97671927,"identity":"ab2683ac-2dae-4174-a071-5694c62e0b74","added_by":"auto","created_at":"2025-12-08 09:33:23","extension":"png","order_by":24,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":1012797,"visible":true,"origin":"","legend":"","description":"","filename":"OnlineFigure5.png","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/5ba4e91c26b06a84a26bb0c9.png"},{"id":97530381,"identity":"b9d20443-bdc5-476c-b6aa-7b0809b794b3","added_by":"auto","created_at":"2025-12-05 13:15:19","extension":"png","order_by":25,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":455955,"visible":true,"origin":"","legend":"","description":"","filename":"OnlineFigure6.png","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/d6d7dc957187ca4a8d9108e5.png"},{"id":97530370,"identity":"1e4e90c2-5ec3-424b-87ed-1d68a4d5ea01","added_by":"auto","created_at":"2025-12-05 13:15:18","extension":"png","order_by":26,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":809844,"visible":true,"origin":"","legend":"","description":"","filename":"OnlineFigure7.png","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/4121ffc8974baf34544676d7.png"},{"id":97530420,"identity":"ce25feb7-b85e-4e19-8f48-32ef22c1a43f","added_by":"auto","created_at":"2025-12-05 13:15:20","extension":"png","order_by":27,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":420822,"visible":true,"origin":"","legend":"","description":"","filename":"OnlineFigure8.png","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/8bb402b61ffd80de480089e3.png"},{"id":97530444,"identity":"7a2d873d-7833-4407-841a-05fc39f169ef","added_by":"auto","created_at":"2025-12-05 13:15:21","extension":"xml","order_by":28,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":227359,"visible":true,"origin":"","legend":"","description":"","filename":"b640308437474b34976796635ba930a61structuring.xml","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/8df532f922cae41d26563ecb.xml"},{"id":97530432,"identity":"c9f8c336-8243-462e-9786-dd579655f95a","added_by":"auto","created_at":"2025-12-05 13:15:21","extension":"html","order_by":29,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":248096,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/78c2be9a22d42981c5dc1b44.html"},{"id":97530363,"identity":"c077c90c-17b0-4e93-bdc2-bf3e35dd2183","added_by":"auto","created_at":"2025-12-05 13:15:18","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":1603261,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eInhibition of six pathogens by dual-culture, volatile and nonvolatile substances.\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Figure1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/64647b6a3163322d8013dc13.jpg"},{"id":97530379,"identity":"d85887d2-14a1-46a6-a05d-67680d37af44","added_by":"auto","created_at":"2025-12-05 13:15:19","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":453016,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eMorphological abnormalities and cytoplasmic leakage of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eC. cereale\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e conidia after cocultivation with Ba. YN. J3.\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Figure2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/0e5ef29c6833f5d7f2a054b6.jpg"},{"id":97530380,"identity":"abe099fe-3226-4751-b685-077289f5e466","added_by":"auto","created_at":"2025-12-05 13:15:19","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":1209302,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003e\u003cstrong\u003eIn vitro\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e screening for plant growth-promoting (PGP) traits and cell wall-degrading enzyme (CWDE) activities of Ba. YN. J3.\u003c/strong\u003e PGP activities are as follows: (A) nitrogen fixation, (B) organic phosphate solubilization, (C) inorganic phosphate solubilization, (D) potassium solubilization, and (H) siderophoreproduction. The following assays for determining CWDE activity were used: (E) cellulase, (F) protease, (G) amylase, (I) pectinase, (J) β-1,3-glucanase, and (K) chitinase.\u003c/p\u003e","description":"","filename":"Figure3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/fcdd9553492cab122350b1d1.jpg"},{"id":97530383,"identity":"5e7058b1-6578-48c2-8bd6-c7e30b0530a0","added_by":"auto","created_at":"2025-12-05 13:15:19","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":223016,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGrowth of strain Ba. YN. J3 is under abiotic stresses such as different pH values (4–11), temperatures (10–45 °C), salt concentrations (1–15%) and PEG6000 concentrations (1–15%).\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Figure4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/04696abb9fbc8ae7a13f80d5.jpg"},{"id":97530361,"identity":"61381d09-e391-472a-91de-d9f88dca798b","added_by":"auto","created_at":"2025-12-05 13:15:17","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":3589858,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003ePlant growth-promoting effects of Ba.YN. Effects of J3 on various crop seedlings.\u003c/strong\u003e For each panel, the seedling on the left is the uninoculated control, and the seedling on the right was inoculated with strain Ba. YN. J3. The crops are (A) maize, (B) sunflower, (C) wheat, and (D) tomato.\u003c/p\u003e","description":"","filename":"Figure5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/9f2d0c311363d6673e00b8d2.jpg"},{"id":97530373,"identity":"ac92bb60-a0cd-4ceb-982e-e158a2bd8063","added_by":"auto","created_at":"2025-12-05 13:15:18","extension":"jpg","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":2045656,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eEffects of Ba.YN. Effects of J3 treatment on the activities of antioxidant enzymes (SOD, POD, CAT, and PPO) in oat seedlings.\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Figure6.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/85f62b9961cb383d4de6ed62.jpg"},{"id":97530371,"identity":"e8ad962a-5ca8-437a-b5a4-e6a96e2daf6e","added_by":"auto","created_at":"2025-12-05 13:15:18","extension":"jpg","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":4178281,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGenomic features and functional gene classification of Ba. YN. J3. \u003c/strong\u003eCircular maps depict the chromosome (A) and a plasmid (B), with gene functions summarized in a histogram (C). Moving from the periphery to the center of the circular maps (A and B), the concentric rings display the following genomic features. The outermost circle (1) marks the genomic coordinates in kilobases (kb). The subsequent two circles (2 and 3) show the annotated protein-coding genes on the forward (+) and reverse (-) strands, which are color-coded on the basis of their COG functional categories. Circle 4 indicates the positions of noncoding RNAs (ncRNAs). The next circle (5) visualizes the GC content as a deviation from the genome-wide average, where red peaks and blue troughs represent regions of higher and lower GC content, respectively. The innermost circle (6) represents the GC skew [(G-C)/(G+C)], with orange and green peaks indicating the local abundances of G over C and C over G, respectively. Panel (C) presents a histogram quantifying the distribution of annotated genes across the various COG functional categories.\u003c/p\u003e","description":"","filename":"Figure7.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/114cbd981e5bd13b1b5b447c.jpg"},{"id":97530365,"identity":"0bd30361-2990-43ec-8609-e8d91c5cad53","added_by":"auto","created_at":"2025-12-05 13:15:18","extension":"jpg","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":1792781,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eComparative genomics reveals the phylogenetic position and unique features of Ba. YN. J3. \u003c/strong\u003eThis figure compares the genomes of strains Ba. YN. J3 against nine other \u003cem\u003eBacillus\u003c/em\u003ereference strains to elucidate their evolutionary and functional differences. A phylogenetic tree was constructed on the basis of whole-genome single nucleotide polymorphisms (SNPs) to determine the evolutionary position of Ba. YN. J3 (B), with pairwise SNP counts indicating the genomic distance between strains (A). Small-scale genomic variations were further assessed through an analysis of insertions and deletions (indels) (C). At the gene content level, the numbers of shared and unique gene families were compared (D), and pangenome analysis, visualized as a flower plot, highlighted the core, accessory, and strain-specific genes (E).\u003c/p\u003e","description":"","filename":"Figure8.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/f78fb62115e999bfc724d875.jpg"},{"id":97893275,"identity":"a779111a-2bcc-44ae-8b0e-39280c5efc97","added_by":"auto","created_at":"2025-12-10 15:29:37","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":17036062,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/e95963d4-cdce-4266-98d8-e934871f2402.pdf"},{"id":97670949,"identity":"f12a61da-9a48-407d-9f5d-b22bc397e4be","added_by":"auto","created_at":"2025-12-08 09:31:34","extension":"docx","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":9385984,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryMaterial.docx","url":"https://assets-eu.researchsquare.com/files/rs-8097513/v1/236c33bfe13c9e8397b16270.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Whole-Genome Analysis Reveals the Growth-Promoting and Biocontrol Potential of Bacillus amyloliquefaciens Ba. YN. J3 isolated from Avena sativa","fulltext":[{"header":"1. Background","content":"\u003cp\u003eMicroorganisms residing within plant tissues and the rhizosphere play crucial roles in plant development by suppressing pathogens and improving nutrient availability [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. The plant endophytes are a unique group of microbes that colonize the internal tissues of plants without causing apparent harm to their host [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Previous studies have shown that endophytes can promote plant growth by producing phytohormones, such as auxin and gibberellin, or by acting as biofertilizers through mechanisms such as nitrogen fixation and solubilization of phosphate and potassium [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Additionally, certain endophytes can significantly suppress the infection and spread of plant pathogens, thereby protecting plants from disease. Given their dual role in growth promotion and disease control, endophytes have garnered considerable attention as promising tools in sustainable agricultural production.\u003c/p\u003e\u003cp\u003eBeneficial agricultural bacteria promote plant growth through two primary mechanisms: the production of phytohormones and the increase in nutrient availability [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. Phytohormones such as auxin and gibberellin, which are produced by many bacterial strains, have been shown to facilitate plant growth and improve tolerance to abiotic stresses such as high salinity [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. The synthesis of indole-3-acetic acid (IAA), a key auxin, is often governed by critical gene clusters such as the \u003cem\u003etrp\u003c/em\u003e operon [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. For example, the well-studied rhizobacteria \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e SQR9 and \u003cem\u003eB. thuringiensis\u003c/em\u003e RZ2MS9 produce IAA, a trait attributed to biosynthetic genes, including \u003cem\u003etrp\u003c/em\u003e and \u003cem\u003eipdC\u003c/em\u003e [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. In addition to phytohormone production, the nutrient supply provided by endophytic and rhizosphere bacteria is increased through processes such as biological nitrogen fixation and solubilization of phosphorus and potassium. These functions are linked to specific genetic determinants; for example, the \u003cem\u003enif\u003c/em\u003e, \u003cem\u003erpo\u003c/em\u003e, and \u003cem\u003entr\u003c/em\u003e genes are involved in nitrogen fixation, whereas genes such as \u003cem\u003epho\u003c/em\u003e, \u003cem\u003eglt\u003c/em\u003e, and \u003cem\u003ekdp\u003c/em\u003e are associated with phosphorus and potassium solubilization, respectively [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eAntagonism, the direct inhibition of plant pathogens, is a major biocontrol mechanism employed by beneficial bacteria [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. This inhibition is often mediated by a diverse array of bioactive secondary metabolites, including lipopeptides, siderophores, and bacteriocins [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. Lipopeptides, a prominent class of these metabolites, include compounds such as surfactin, iturin, and fengycin, which are synthesized by large enzymatic complexes called nonribosomal peptide synthetases (NRPSs) [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. The biosynthesis of these molecules is governed by extensive gene clusters; for example, the production of fengycin is directed by the \u003cem\u003efen\u003c/em\u003e gene cluster, which encodes five large peptide synthetases [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Siderophores represent another critical class of antagonistic compounds. These low-molecular-weight molecules are secreted to chelate ferric iron from the environment, thereby limiting its availability to competing pathogens [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. The pseudobactin siderophore, for example, is a key factor in the biocontrol of \u003cem\u003eBotrytis cinerea\u003c/em\u003e by \u003cem\u003ePseudomonas\u003c/em\u003e spp., and its production is regulated by gene clusters such as \u003cem\u003eiucABCD\u003c/em\u003e [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eSuccessful colonization of host tissues is a prerequisite for the beneficial functions of endophytes. This process often begins with chemotaxis, where microorganisms recognize compounds secreted by the plant and move toward its surface before entering through natural openings, wounds, or direct penetration [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. Chemotactic activity, alongside the secretion of cell wall-degrading enzymes (CWDEs), is therefore key mechanisms facilitating endophytic colonization. Numerous studies have documented this phenomenon in various endophytic bacteria, such as \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e SQR9, \u003cem\u003eP. putida\u003c/em\u003e KT2440, and \u003cem\u003eRhizobium leguminosarum\u003c/em\u003e N5 [\u003cspan additionalcitationids=\"CR22 CR23\" citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]. The specific chemoattractants identified include a range of sugars and organic acids that guide the targeted movement of \u003cem\u003eB. cereus\u003c/em\u003e YL6 [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. Similarly, fatty acids enhance the motility of \u003cem\u003eB. flexus\u003c/em\u003e KLBMP 4941 [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e], whereas malic acid, glucose, and fructose promote the colonization of \u003cem\u003eB. velezensis\u003c/em\u003e S3-1 [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. This chemical attraction is a conserved ecological strategy, as genera, including \u003cem\u003eAzospirillum\u003c/em\u003e, \u003cem\u003ePseudomonas\u003c/em\u003e, \u003cem\u003eSerratia\u003c/em\u003e, and \u003cem\u003eEnterobacter\u003c/em\u003e, also display chemotactic responses to host-derived compounds, highlighting the importance of rhizosphere colonization across diverse plant-associated bacteria [\u003cspan additionalcitationids=\"CR29\" citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eWhole-genome sequencing and comparative genomics are powerful tools for uncovering the genetic basis of the diverse traits observed among biocontrol bacteria. This approach can reveal the key genetic determinants underlying their biocontrol and plant growth-promoting functions. For example, a whole-genome analysis of the sugarcane endophyte \u003cem\u003eP. aeruginosa\u003c/em\u003e B18 revealed its genetic potential for nitrogen fixation, phosphorus solubilization, and the production of IAA, siderophores, and antibacterial compounds [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e]. Similarly, the genome of the endophyte \u003cem\u003eB. velezensis\u003c/em\u003e K1 contains genes associated with the induction of plant resistance, the production of phytohormones, nitrogen fixation, phosphate solubilization, and colonization [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e]. Furthermore, comparative genomic analysis employs a suite of methods to elucidate the genetic basis of phenotypic diversity among bacteria. At a fine scale, analyses of single nucleotide polymorphisms (SNPs), insertions/deletions (indels), and structural variations (SVs) are used to resolve genomic differences, providing insights into genetic diversity and linking specific variations to functional traits [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e, \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e]. At the gene content level, pangenome analysis identifies the core genome shared by a group of strains, as well as the accessory and strain-specific genes that may confer unique adaptations. The comparison of gene family evolution can further illuminate the distinct biocontrol functions of different strains [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e]. These identified genetic variations can ultimately be correlated with phenotypic traits through association analyses to pinpoint the molecular basis of specific activities.\u003c/p\u003e\u003cp\u003eThe oat endophyte \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e YN-J3 (hereafter Ba. YN. J3) has previously been shown to exert a significant biocontrol effect against oat anthracnose in both greenhouse and field assays [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e]. Further studies have demonstrated its capacity for targeted chemotaxis toward fungal mycelia and its ability to inhibit spore germination and appressorium formation in \u003cem\u003eColletotrichum cereale\u003c/em\u003e [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. Building on these findings, the present study was designed to comprehensively characterize the multifaceted potential of Ba. YN. J3. We evaluated its broad-spectrum antifungal activity, production of cell wall-degrading enzymes (CWDEs), plant growth-promoting (PGP) traits, and ability to induce host plant resistance. To elucidate the molecular mechanisms underlying these beneficial properties, we subsequently performed whole-genome sequencing and comparative genomic analyses. The primary objective was to identify the key genes and gene clusters responsible for its biocontrol and PGP functions, with a specific focus on identifying unique genetic determinants related to chemotaxis and the suppression of appressorium formation.\u003c/p\u003e"},{"header":"2. Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003e2.1 Microbial strains and culture conditions\u003c/h2\u003e\u003cp\u003eThe endophytic bacterium Ba. YN. J3 was previously isolated from the stems of oat (\u003cem\u003eAvena sativa\u003c/em\u003e cv. Baiyan No. 2). For long-term storage, stock cultures were maintained at -80\u0026deg;C in Luria\u0026ndash;Bertani (LB) broth containing 30% (v/v) glycerol. For routine experiments, Ba. YN. J3 was cultured in LB medium at 28\u0026deg;C with shaking [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThe fungal pathogens used in this study included \u003cem\u003eC. cereale\u003c/em\u003e, \u003cem\u003eSclerotinia sclerotiorum\u003c/em\u003e, \u003cem\u003eDrechslera glomerata\u003c/em\u003e, \u003cem\u003eFusarium oxysporum\u003c/em\u003e, \u003cem\u003eRhizoctonia solani\u003c/em\u003e, and \u003cem\u003eAlternaria alternata\u003c/em\u003e. The fungal stock cultures were stored under the same conditions as the bacterial cultures. For experimental use, mycelial plugs from the frozen stocks were inoculated onto the center of potato dextrose agar (PDA) plates and incubated at 25\u0026deg;C for 7\u0026ndash;14 days to obtain fresh cultures for subsequent assays.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec4\" class=\"Section2\"\u003e\u003ch2\u003e2.2 \u003cem\u003eIn\u003c/em\u003e vitro antifungal activity assays\u003c/h2\u003e\u003cp\u003eDual-culture assay: Antagonistic activity was evaluated via a dual-culture plate assay. A 5-mm mycelial plug from an actively growing fungal culture was placed at the center of a potato dextrose agar (PDA) plate. A 24-h-old culture of Ba. YN. J3 was then streaked in a line 2.5 cm from the plug. The plates inoculated with only the fungal plug served as the control. After incubation at 25\u0026deg;C for 7\u0026ndash;14 days, the fungal colony radius was measured for both the control (Rc) and treatment (Rt) plates. The percentage of growth inhibition was calculated as follows: Inhibition (%) = [(Rc\u0026thinsp;\u0026minus;\u0026thinsp;Rt)/Rc] \u0026times; 100 [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eIndirect Inhibition Assay: The effects of volatile organic compounds (VOCs) were assessed via the sealed double-dish method [\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e]. A PDA plate was centrally inoculated with a 5-mm fungal plug. This plate was then inverted over a second plate, which was subsequently streaked with Ba. YN. J3. The pairs of plates were sealed together with Parafilm and incubated at 25\u0026deg;C for 7\u0026ndash;14 days. The control consisted of a fungal plate paired with an uninoculated LB agar plate. Fungal growth inhibition was calculated as described above.\u003c/p\u003e\u003cp\u003eNon-Volatile Metabolite Assay: The effects of nonvolatile metabolites were evaluated via the use of cell-free supernatants [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e]. The Ba. YN. J3 was cultured in LB broth at 37\u0026deg;C with shaking (180 rpm) for 48 h. The culture was centrifuged (12,000 \u0026times; \u003cem\u003eg\u003c/em\u003e, 15 min), and the supernatant was sterilized by filtration through a 0.22-\u0026micro;m membrane. This cell-free supernatant was incorporated into molten PDA (previously cooled to ~\u0026thinsp;50\u0026deg;C) to a final concentration of 10% (v/v). The amended PDA was poured into plates, which were then centrally inoculated with 5-mm fungal plugs and incubated at 25\u0026deg;C for 7\u0026ndash;14 days. PDA plates supplemented with sterile LB broth served as the control. Fungal growth inhibition was calculated as described above.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec5\" class=\"Section2\"\u003e\u003ch2\u003e2.3 Abiotic stress tolerance assays\u003c/h2\u003e\u003cp\u003eThe tolerance of Ba. YN. J3 to various abiotic stresses was determined by measuring its growth (OD600) in LB broth under different conditions [\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e, \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e]. For all the assays, 200 \u0026micro;L of an overnight Ba. YN. J3 culture was inoculated into 20 mL of the respective test medium.\u003c/p\u003e\u003cp\u003eTemperature tolerance: Cultures were incubated at 10, 15, 20, 25, 30, 35, 40, or 45\u0026deg;C with shaking (180 rpm). After 24 h, growth was quantified by measuring the OD₆₀₀.\u003c/p\u003e\u003cp\u003epH tolerance: Cultures were grown at 28\u0026deg;C in LB broth adjusted to pH values of 4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10.0, or 11.0. After 24 h, growth was quantified by measuring the OD₆₀₀.\u003c/p\u003e\u003cp\u003eSalinity tolerance: Cultures were grown at 28\u0026deg;C in LB broth supplemented with NaCl to final concentrations of 1, 3, 5, 7, 9, or 11% (w/v). After 24 h, growth was quantified by measuring the OD₆₀₀.\u003c/p\u003e\u003cp\u003eDrought tolerance: Drought stress was simulated via LB broth supplemented with PEG6000 at concentrations of 1, 3, 5, 7, 9, 11, 13, or 15% (w/v). Cultures were incubated at 28\u0026deg;C, and growth was quantified by measuring the OD₆₀₀ after 48 h.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec6\" class=\"Section2\"\u003e\u003ch2\u003e2.4 Screening for Plant Growth Promotion (PGP) and Enzymatic Activities\u003c/h2\u003e\u003cp\u003eThe PGP and enzymatic activities of Ba. YN. J3 were assessed via qualitative plate assays.\u003c/p\u003e\u003cp\u003eNitrogen Fixation: Nitrogen fixation ability was assayed by observing the growth of Ba. YN. J3 on Ashby's nitrogen-free medium [\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eNutrient solubilization: Phosphate and potassium solubilization activities were evaluated by the formation of clearing zones (halos) around colonies grown on NBRIP medium and Alexandrov medium, respectively [\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e, \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eSiderophore Production: Siderophore production was detected by the formation of a pale-yellow halo on Chrome Azurol S (CAS) agar plates [\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eHydrolytic Enzyme Production: The production of various hydrolytic enzymes was screened by observing hydrolysis halos on agar plates supplemented with specific substrates. A positive result was indicated by a clear zone around the colony after 2\u0026ndash;5 days of incubation at 28\u0026deg;C [\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e, \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e]. The substrates used were as follows: 1% (w/v) sodium carboxymethyl cellulose (for cellulase), 1% (w/v) skim milk powder (for protease), 1% (w/v) soluble starch (for amylase), 1% (w/v) apple pectin (for pectinase), 0.5% (w/v) \u003cem\u003ePoria cocos\u003c/em\u003e powder (for β-1,3-glucanase), and 0.5% (w/v) colloidal chitin (for chitinase).\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec7\" class=\"Section2\"\u003e\u003ch2\u003e2.5 \u003cem\u003eIn Planta\u003c/em\u003e Growth Promotion Assays\u003c/h2\u003e\u003cp\u003eThe Ba. YN. J3 was cultured in LB broth (28\u0026deg;C, 180 rpm, 24 h). The bacterial cells were harvested by centrifugation (5,000 \u0026times; g, 10 min), washed once with sterile water, and finally resuspended in sterile water to an optical density at OD₆₀₀=1.0. This stock suspension was then diluted 10-fold with sterile water for inoculation.\u003c/p\u003e\u003cp\u003eFor the plant growth conditions and treatment, seeds of corn (\u003cem\u003eZea mays\u003c/em\u003e cv. Xianyu 335), wheat (\u003cem\u003eTriticum aestivum\u003c/em\u003e cv. Longmai 21), sunflower (\u003cem\u003eHelianthus annuus\u003c/em\u003e cv. SH361), and tomato (\u003cem\u003eSolanum lycopersicum\u003c/em\u003e cv. Antles) were surface sterilized and germinated. Uniform seedlings at the two-leaf stage were transplanted into pots containing a 2:1 (v/v) mixture of nutrient-rich soil and field soil. Each seedling was then inoculated via root drenching with 50 mL of the diluted \u003cem\u003eBa. YN. J3\u003c/em\u003e suspension. The control plants received an equal volume (50 mL) of sterile water. The plants were maintained in a greenhouse under a 16 h/8 h light/dark cycle at 26\u0026deg;C [\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e]. After 21 days, various growth parameters, including plant height, root length, root fresh weight, root dry weight (biomass), and leaf chlorophyll content, were measured.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e\u003ch2\u003e2.6 Defense-Related Enzyme Assays in Oats\u003c/h2\u003e\u003cp\u003eFor detect Bacterial suspensions of Ba. YN. J3 were prepared as described in Section \u003cspan refid=\"Sec9\" class=\"InternalRef\"\u003e2.7\u003c/span\u003e (final OD₆₀₀ diluted to 0.1).\u003c/p\u003e\u003cp\u003eUniform oat seedlings (\u003cem\u003eAvena sativa\u003c/em\u003e cv. Baiyan No. 2) at the two-leaf stage were used for the assay. Plants in the treatment group were inoculated by foliar spraying with the Ba. YN. J3 suspension until the leaves were thoroughly wet. Control plants were sprayed with an equal volume of sterile water. The plants were maintained in the greenhouse under conditions described in Section \u003cspan refid=\"Sec9\" class=\"InternalRef\"\u003e2.7\u003c/span\u003e.\u003c/p\u003e\u003cp\u003eLeaf samples were collected at various time points post-spraying (hps), immediately frozen in liquid nitrogen, and stored at -80\u0026deg;C for analysis.\u003c/p\u003e\u003cp\u003eThe activities of four defense-related enzymes\u0026mdash;superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and polyphenol oxidase (PPO)\u0026mdash;were measured using commercial assay kits (Boxbio, Beijing, China) according to the manufacturer\u0026rsquo;s instructions. Enzyme activities were quantified using a spectrophotometer.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec9\" class=\"Section2\"\u003e\u003ch2\u003e2.7 Genome Sequencing, Assembly, and Annotation\u003c/h2\u003e\u003cp\u003eThe genomic DNA was extracted from an overnight culture of Ba. YN. J3 (grown in LB broth at 28\u0026deg;C with shaking at 180 rpm) using the STE buffer method. The quality and concentration of the extracted DNA were verified via 1% agarose gel electrophoresis and a Qubit fluorometer (Thermo Fisher Scientific, USA), respectively [\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eWhole-genome sequencing was performed by Novogene Co., Ltd. (Beijing, China) on the Pacific Biosciences (PacBio) Sequel platform via single-molecule real-time (SMRT) technology [\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e]. After filtering to remove low-quality data, the resulting high-quality reads were assembled de novo via Canu (v2.0). This process generates a single, complete, and gapless contig for the chromosome [\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eGenome annotation began with the prediction of various components. Coding sequences (CDSs) were predicted via GeneMarkS (v4.17) [\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e]. Repetitive elements were identified via RepeatMasker (v4.0.5) for interspersed repeats and Tandem Repeats Finder (v4.07b) for tandem repeats [\u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e]. Noncoding RNAs (ncRNAs), including tRNAs and rRNAs, were predicted via tRNAscan-SE (v1.3.1) and rRNAmmer (v1.2), respectively [\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e], whereas other ncRNAs were identified via searches of the Rfam database [\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e, \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e]. Finally, mobile genetic elements such as genomic islands, prophages, and CRISPR arrays were identified via IslandPath-DIOMB (v0.2), phiSpy (v2.3), and CRISPRdigger (v1.0), respectively [\u003cspan additionalcitationids=\"CR60\" citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThe functional annotation of the predicted genes was performed by aligning their sequences against the KEGG [\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e, \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e], SwissProt, and Pfam databases. Specific attention was given to pathways related to nitrogen fixation, phosphate/potassium solubilization, siderophore biosynthesis, and hydrolytic enzymes. Gene clusters for secondary metabolite biosynthesis were identified via antiSMASH (v4.0.2) [\u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e64\u003c/span\u003e], and carbohydrate-active enZymes (CAZymes) were annotated via the dbCAN database [\u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e].\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec10\" class=\"Section2\"\u003e\u003ch2\u003e2.8 Comparative Genomic Analysis\u003c/h2\u003e\u003cp\u003eFor the comparative genomic analysis, the genome of Ba. YN. J3 was compared against nine other \u003cem\u003eBacillus\u003c/em\u003e and \u003cem\u003ePriestia\u003c/em\u003e strains selected for its known or potential biocontrol properties (detailed in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e5\u003c/span\u003e). The whole-genome alignments and synteny analyses were performed via MUMmer and LASTZ. These alignments were then used to identify single nucleotide polymorphisms (SNPs), insertions/deletions (indels), and structural variations (SVs) [\u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e66\u003c/span\u003e, \u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e67\u003c/span\u003e]. To assess evolutionary relationships, a phylogenetic tree was constructed via PhyML with 1,000 bootstrap replicates on the basis of the concatenated alignment of single-copy core genes. Pangenome analysis was conducted via CD-HIT (50% identity, 70% coverage thresholds) to identify core and strain-specific gene sets. Finally, key genomic features of Ba. YN. J3 were visualized on a circular map generated with Circos (v0.66).\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 5\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eStrain information required for comparative genome analysis\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"3\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eStrain name\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eStrain name in report\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eGenebank accession number\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eB. amyloliquefaciens\u003c/em\u003e YN.J3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBa. YN. J3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003ePRJNA1171741\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eB. amyloliquefaciens\u003c/em\u003e GKT04\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBa.GKT04\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCP072120.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eB. amyloliquefaciens\u003c/em\u003e DSM 7\u0026thinsp;=\u0026thinsp;ATCC 23350\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBa.DSM7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eFN597644.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eB. amyloliquefaciens\u003c/em\u003e ZKY01\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBa.ZKY01\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCP044132.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eB.velezensis\u003c/em\u003e FZB42\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBv.FZB42\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCP000560.2\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eB. subtilis subsp. subtilis str.\u003c/em\u003e 168\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBs.168\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eAL009126.3\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003ePriestia megaterium NBRC\u003c/em\u003e 15308\u0026thinsp;\u003cem\u003e=\u003c/em\u003e\u0026thinsp;ATCC 14581\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003ePm.14581\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCP035094.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eB. cereus\u003c/em\u003e FORC_047\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBc.FORC.047\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCP017060.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eB. halotolerans\u003c/em\u003e ZB201702\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBh.ZB201702\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCP029364.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eB. pumilus\u003c/em\u003e SAFR-032\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eBp.SAFR.032\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCP000813.4\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e\u003ch2\u003e2.9 Statistical analysis\u003c/h2\u003e\u003cp\u003eAll experimental data were presented as the mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation (SD) of at least five biological replicates. One-way analysis of variance (ANOVA) was used to determine statistical significance. Mean separation was performed via Duncan's multiple range test at a significance level of \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05. All the statistical analyses were conducted via SPSS Statistics v26.0 (IBM Corp., Armonk, NY, USA).\u003c/p\u003e\u003c/div\u003e"},{"header":"3. Results","content":"\u003cdiv id=\"Sec13\" class=\"Section2\"\u003e\u003ch2\u003e3.1 Ba. YN. J3 Exhibits Broad-Spectrum Antifungal Activity\u003c/h2\u003e\u003cp\u003eThe biocontrol potential of Ba. YN. J3 against six major phytopathogenic fungi were evaluated. In dual-culture assays, Ba. YN. J3 significantly inhibited the mycelial growth of \u003cem\u003eC. cereale\u003c/em\u003e, \u003cem\u003eS. sclerotiorum\u003c/em\u003e, \u003cem\u003eD. glomerata\u003c/em\u003e, \u003cem\u003eF. oxysporum\u003c/em\u003e, \u003cem\u003eR. solani\u003c/em\u003e, and \u003cem\u003eA. alternata\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e1\u003c/span\u003e). To determine the mechanisms of this inhibition, the effects of volatile organic compounds (VOCs) and nonvolatile metabolites were assessed separately. Both VOCs and nonvolatile metabolites significantly suppressed the mycelial expansion of five of the tested pathogens, with inhibition rates ranging from 44.33% to 80.02% (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Notably, for \u003cem\u003eC. cereale\u003c/em\u003e, \u003cem\u003eD. glomerata\u003c/em\u003e, \u003cem\u003eF. oxysporum\u003c/em\u003e, and \u003cem\u003eA. alternata\u003c/em\u003e, the inhibitory effects of the nonvolatile metabolites were significantly stronger than those of the VOCs. Although the colony diameter of \u003cem\u003eR. solani\u003c/em\u003e was not significantly reduced by either treatment, its aerial mycelia were visibly thinner than those of the control (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). These results indicate that Ba. YN. J3 possesses broad-spectrum antifungal activity, which is mediated by both volatile and nonvolatile compounds that inhibit the polar growth and branching of fungal hyphae.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eThe inhibition rates of Ba. YN. J3 against several pathogens\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"4\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eStrains\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colspan=\"3\" nameend=\"c4\" namest=\"c2\"\u003e\u003cp\u003eInhibition rate/%\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eDual culture\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eVolatile substance\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eNon-Volatile substance\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eC. cereale\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e71.65\u0026thinsp;\u0026plusmn;\u0026thinsp;0.64 b\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e65.86\u0026thinsp;\u0026plusmn;\u0026thinsp;1.22 a\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e78.29\u0026thinsp;\u0026plusmn;\u0026thinsp;1.02 c\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eS. sclerotiorum\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e48.13\u0026thinsp;\u0026plusmn;\u0026thinsp;0.74 c\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e46.01\u0026thinsp;\u0026plusmn;\u0026thinsp;0.39 b\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e44.33\u0026thinsp;\u0026plusmn;\u0026thinsp;0.51 a\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eD. glomerata\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e55.13\u0026thinsp;\u0026plusmn;\u0026thinsp;0.28 b\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e51.72\u0026thinsp;\u0026plusmn;\u0026thinsp;0.88 a\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e66.08\u0026thinsp;\u0026plusmn;\u0026thinsp;0.77 c\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eF. oxysporum\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e62.17\u0026thinsp;\u0026plusmn;\u0026thinsp;0.35 b\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e58.57\u0026thinsp;\u0026plusmn;\u0026thinsp;0.78 a\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e62.79\u0026thinsp;\u0026plusmn;\u0026thinsp;0.78 b\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eR. solani\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e48.09\u0026thinsp;\u0026plusmn;\u0026thinsp;0.74 b\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e1.84\u0026thinsp;\u0026plusmn;\u0026thinsp;0.53 a\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e2.37\u0026thinsp;\u0026plusmn;\u0026thinsp;0.70 a\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e\u003cem\u003eA. alternata\u003c/em\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e61.01\u0026thinsp;\u0026plusmn;\u0026thinsp;0.95 b\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e51.73\u0026thinsp;\u0026plusmn;\u0026thinsp;1.54 a\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e80.02\u0026thinsp;\u0026plusmn;\u0026thinsp;0.70 c\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec14\" class=\"Section2\"\u003e\u003ch2\u003e3.2 Ba. YN. J3 Secretes Different Cell-wall-Degrading Enzymes\u003c/h2\u003e\u003cp\u003eTo investigate the biocontrol mechanism of Ba. YN. J3 against \u003cem\u003eC. cereale\u003c/em\u003e, the interaction between the bacterium and fungal conidia was observed. Following cocultivation, the conidia presented severe morphological abnormalities, including cytoplasmic leakage, suggesting that Ba. YN. J3 causes defects in the fungal cell wall (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). On the basis of this observation, we hypothesized that Ba. YN. J3 secretes extracellular cell wall-degrading enzymes (CWDEs). Subsequent plate assays confirmed this, as translucent hydrolysis zones were observed on media containing substrates for cellulase, protease, amylase, pectinase, and chitinase (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eE-G, I, K). In addition, Ba. YN. J3 also tested positive for siderophore production (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eH). Further experiments revealed that Ba. YN. J3 could utilize colloidal chitin as the sole carbon source, confirming chitinase activity, but failed to grow on a medium with colloidal β-1,3-glucan, indicating that it did not produce detectable β-1,3-glucanase under these conditions (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eJ and \u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eK). Collectively, these results demonstrate that Ba. YN. J3 secretes a diverse array of lytic enzymes capable of disrupting the fungal cell wall.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec15\" class=\"Section2\"\u003e\u003ch2\u003e3.3 Environmental adaptability and abiotic stress tolerance of Ba. YN. J3\u003c/h2\u003e\u003cp\u003eTo evaluate its potential for field application, the environmental adaptability of Ba. YN. J3 to various abiotic stresses was assessed (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e). The strain demonstrated broad temperature tolerance, growing effectively between 10\u0026deg;C and 45\u0026deg;C, with optimal growth observed at 30\u0026deg;C. It also tolerates a wide pH range from 5.0\u0026ndash;9.0, although growth is significantly inhibited under more strongly acidic (pH\u0026thinsp;\u0026lt;\u0026thinsp;5.0) or alkaline (pH\u0026thinsp;\u0026gt;\u0026thinsp;9.0) conditions. Furthermore, Ba. YN. J3 exhibited tolerance to salinity and osmotic stress. Its growth gradually decreased with increasing NaCl concentration, showing a sharp decline only when the concentration exceeded 7% (w/v). Similarly, in the presence of PEG6000 to simulate drought, the strain growth decreased only slightly up to a concentration of 11% (w/v), beyond which a sharp decline occurred. Collectively, these results indicate that Ba. YN. J3 is well adapted to a wide range of temperatures and pH values and is tolerant to moderate levels of salinity and osmotic stress.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec16\" class=\"Section2\"\u003e\u003ch2\u003e3.4 Ba. YN. J3 promotes the growth of four crop species\u003c/h2\u003e\u003cp\u003ePrevious studies have shown that Ba. YN. J3 promotes oat seedling growth through mechanisms such as IAA synthesis and nutrient provision [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e]. To determine whether this effect extends to other crops, its growth-promoting activity was further evaluated in sunflower, corn, wheat, and tomato. Compared with the uninoculated controls, Ba. YN. J3 treatment significantly increased shoot length, root length, root fresh weight, and root dry weight in all tested species. A significant increase in chlorophyll content was observed only in corn and sunflower, indicating host-specific effects on photosynthetic capacity (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e). The most pronounced growth enhancement was found in corn, followed by sunflower, tomato, and wheat, suggesting that Ba. YN. J3 possesses broad-spectrum plant growth-promoting potential.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec17\" class=\"Section2\"\u003e\u003ch2\u003e3.5 Ba. YN. J3 induces systemic disease resistance in oats\u003c/h2\u003e\u003cp\u003eWe next investigated whether Ba. YN. J3 could induce systemic resistance in oat plants. The activities of four key defense-related antioxidant enzymes\u0026mdash;superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and polyphenol oxidase (PPO)\u0026mdash;were measured at multiple time points after spraying (hps) and compared with those of a sterile water-treated control (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e). Ba. YN. J3 treatment significantly enhanced the activities of all four enzymes (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). In treated seedlings, enzyme activities progressively increased, reached their maximum at 36 hps, and then slightly declined, yet remained significantly higher than those in the control group throughout the experiment. In contrast, enzyme activities in control plants showed negligible variation over time. These findings demonstrate that Ba. YN. J3 effectively activates the antioxidant defense system in oats, thereby enhancing their systemic resistance to stress and potential pathogen attack.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec18\" class=\"Section2\"\u003e\u003ch2\u003e3.6 Genomic features of Ba. YN. J3\u003c/h2\u003e\u003cdiv id=\"Sec19\" class=\"Section3\"\u003e\u003ch2\u003e3.6.1 General Genome and Plasmid Features\u003c/h2\u003e\u003cp\u003eTo elucidate the molecular mechanisms underlying its beneficial activities, the complete genome of Ba. YN. J3 was sequenced. The genome consists of a single circular chromosome of 4,063,196 bp with a GC content of 46.27% and one circular plasmid of 215,473 bp with a GC content of 37.24% (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003e; Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e2\u003c/span\u003e). A total of 4,541 protein-coding genes were predicted, accounting for 90.04% of the genome. In addition, the genome contains 124 noncoding RNA genes (including 27 rRNAs, 87 tRNAs and 10 sRNAs), 15 CRISPR arrays, 5 genomic islands, and 23 pseudogenes (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e2\u003c/span\u003e). The complete genome sequence has been deposited in the GenBank database under accession number PRJNA1171741.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eThe general genome features of Ba. YN. J3\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"2\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eFeature\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eVaule\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eGenome size (bp)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e4,063,196\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eG\u0026thinsp;+\u0026thinsp;C content\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e46.27%\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eTopology\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eCircular\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003ePlasmid\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eTotal number of genes\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e4665\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eTotal size of protein-coding genes\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e3,852,390 bp\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eProtein-coding genes\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e4541\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eAverage CDs size (bp)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e848\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003erRNA number (total)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e27\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003etRNA number\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e87\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003esRNA number\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e10\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eRepetitive sequence(bp)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e35,897(0.839%)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eCRISPR\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e15\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eProphage\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e10\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003ePseudogenes\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e23\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eGls\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e5\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eGene cluster\u003c/p\u003e\u003cp\u003eGenes assigned to NR\u003c/p\u003e\u003cp\u003eGenes assigned to GO\u003c/p\u003e\u003cp\u003eGenes assigned to KEGG\u003c/p\u003e\u003cp\u003eGenes assigned to Pfam\u003c/p\u003e\u003cp\u003eGenes assigned to Swiss-Prot\u003c/p\u003e\u003cp\u003eGenes assigned to CAZy\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e13\u003c/p\u003e\u003cp\u003e4413\u003c/p\u003e\u003cp\u003e2865\u003c/p\u003e\u003cp\u003e4130\u003c/p\u003e\u003cp\u003e2865\u003c/p\u003e\u003cp\u003e3396\u003c/p\u003e\u003cp\u003e161\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec20\" class=\"Section3\"\u003e\u003ch2\u003e3.6.2 Functional Annotation and COG Classification\u003c/h2\u003e\u003cp\u003eThe functional annotation against multiple databases (including NR, GO, KEGG, and SwissProt) assigned functions to a majority of the predicted genes. Analysis of Clusters of Orthologous Groups (COGs) revealed that the most abundant categories were amino acid transport and metabolism (298 genes), transcription (289 genes), and carbohydrate transport and metabolism (248 genes). Significant numbers of genes were also assigned to categories related to cell wall/membrane biogenesis and signal transduction, reflecting the strain's active interaction with its environment (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003eC).\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec21\" class=\"Section3\"\u003e\u003ch2\u003e3.6.3 Genetic basis for biocontrol activities\u003c/h2\u003e\u003cp\u003eConsistent with its observed antifungal activity, the genome of Ba. YN. J3 harbors a rich repertoire of genes associated with biocontrol. Analysis with antiSMASH identified thirteen secondary metabolite biosynthesis-related gene clusters (BGCs), including those for the synthesis of surfactin, fengycin, bacillibactin, and difficidin (Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e3\u003c/span\u003e). Furthermore, the genome contains 286 genes encoding putative hydrolytic enzymes, such as chitinases and proteases, which are implicated in fungal cell wall degradation (Table \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e). The genome also harbors 6 genes for siderophore production and 40 genes related to chemotaxis, providing a genetic basis for its iron competition and targeted motility capabilities (Table\u0026nbsp;\u003cspan refid=\"Tab5\" class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003ePredictive gene clusters involved in the synthesis of secondary metabolites in Ba. YN. J3\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"6\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colspan=\"2\" nameend=\"c2\" namest=\"c1\"\u003e\u003cp\u003eRegion of genome\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colspan=\"4\" nameend=\"c6\" namest=\"c3\"\u003e\u003cp\u003eMost similar known cluster\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eFrom\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eTo\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eType\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eProductions\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003eSimilarity\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003eResources\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e195,469\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e273,197\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eNPRS, transAT-PKS\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003elocillomycin/locillomycin B/locillomycin C\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e35%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e\u003cem\u003eB.subtilis\u003c/em\u003e 916(Luo, 2015)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e342,784\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e408,191\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eNRPS\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003esurfactin\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e82%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e\u003cem\u003eB.subtilis\u003c/em\u003e ATCC21332(Wei, 2004)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e977,079\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e1,108,323\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003ePKS-like\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003ebutirosin A/butirosin B\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e7%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e\u003cem\u003eB.circulans\u003c/em\u003e SANK72073(Kudo, 2005)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1,100,346\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e1,121,086\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eterpene\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1,239,789\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e1,268,678\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eLanthipeptide-class-ii\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1,432,632\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e1,520,843\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003etransAT-PKS\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003emacrolactin H\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e100%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e\u003cem\u003eB.amyloliquefaciens\u003c/em\u003e NJN-6(Yuan, 2012)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1,739,543\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e1,849,654\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003etransAT-PKS, T3PKS, NRPS\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003ebacillaene\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e100%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e\u003cem\u003eB. amyloliquefaciens\u003c/em\u003e SQ-2(Li, 2024)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1,905,107\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e2,042,936\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eNRPS, transAT-PKS, betalactone\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003efengycin\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e100%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e\u003cem\u003eB.subtilis\u003c/em\u003e F-29-3(Vanittanakom, 1986)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2,065,505\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e2,087,388\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eterpene\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2,156,032\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e2,197,132\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eT3PKS\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2,447,181\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e2,553,347\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003etransAT-PKS\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003edifficidin\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e100%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e\u003cem\u003eB.subtilis\u003c/em\u003e ATCC39320(Zimmerman, 1987)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e3,164,202\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e3,215,995\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eNRP-metallophore, NRPS, RiPP-like\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003ebacillibactin\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e100%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e\u003cem\u003eB.siamensis\u003c/em\u003e SCSIO05746(Pan, 2019)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e3,728,795\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e3,770,213\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eOthers\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003ebacilysin\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e100%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e\u003cem\u003eB.velezensis\u003c/em\u003e FZB42(Han, 2021)\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab5\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eComparison of genes related to the biocontrol and growth promotion of Ba. YN. J3 and nine reference \u003cem\u003eBacillus\u003c/em\u003e strains\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"11\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c11\" colnum=\"11\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eStrain name\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colspan=\"5\" nameend=\"c6\" namest=\"c2\"\u003e\u003cp\u003eNumber of biocontrol associated genes\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colspan=\"5\" nameend=\"c11\" namest=\"c7\"\u003e\u003cp\u003eNumber of growth-promoting associated genes\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eA\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eB\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eC\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003eD\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003eE\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003eF\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003eG\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003eH\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003eI\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003eJ\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eBa. YN. J3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e69\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e286\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e40\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e105\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e39\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e63\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e60\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eBa.GKT04\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e12\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e203\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e37\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e18\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e61\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e40\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e0\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eBa.ZKY01\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e27\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e274\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e36\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e19\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e85\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e50\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eBv.FZB42\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e26\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e265\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e12\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e12\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e12\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e73\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e49\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eBa.DSM7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e12\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e269\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e36\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e19\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e68\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e49\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eBs.A168\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e37\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e224\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e19\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e51\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e81\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e116\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e58\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eBh.ZB201702\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e17\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e274\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e41\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e25\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e111\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e56\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e4\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eBp.SAFR.032\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e273\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e49\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e19\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e64\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e49\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e0\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eBc.FORC.047\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e454\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e46\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e14\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e115\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e62\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003ePm.A14581\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e399\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e33\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e23\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e102\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e88\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e5\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003ctfoot\u003e\u003ctr\u003e\u003ctd colspan=\"11\"\u003eNote: A: Secondary metabolites, B: Cell wall-degrading enzymes, C: Siderophores, D: Chemotaxis, E: Colonization ability, F: Nitrogen fixation, G: Phosphate solubilization, H: Potassium solubilization, I: Indole-3-acetic acid (IAA) production, J: Amylase production.\u003c/td\u003e\u003c/tr\u003e\u003c/tfoot\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec22\" class=\"Section3\"\u003e\u003ch2\u003e3.6.4 Genetic basis for plant growth-promoting (PGP) traits\u003c/h2\u003e\u003cp\u003eThe PGP potential of Ba. YN. J3 is also well supported by its genome. We identified 60 genes in the tryptophan biosynthesis pathway leading to IAA production, 39 genes involved in nitrogen metabolism (including the key genes \u003cem\u003enasDE\u003c/em\u003e and \u003cem\u003erocG\u003c/em\u003e), 63 genes related to phosphate solubilization (e.g., the \u003cem\u003epst\u003c/em\u003e system), and 5 genes involved in potassium solubilization (\u003cem\u003ektr\u003c/em\u003e, \u003cem\u003ekdp\u003c/em\u003e) (Table\u0026nbsp;\u003cspan refid=\"Tab5\" class=\"InternalRef\"\u003e4\u003c/span\u003e; Table S2). These genetic features underpin the strain's ability to promote crop growth by producing phytohormones and increasing nutrient availability.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec23\" class=\"Section3\"\u003e\u003ch2\u003e3.6.5 Genetic basis for abiotic stress tolerance\u003c/h2\u003e\u003cp\u003eThe high tolerance of this strain to abiotic stresses is corroborated by the presence of numerous stress response genes (Table \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e). For thermotolerance, the genome encodes several heat shock proteins (e.g., \u003cem\u003ednaK\u003c/em\u003e, \u003cem\u003ednaJ\u003c/em\u003e, and \u003cem\u003eclpB\u003c/em\u003e). For salinity and osmotic stress, genes for the synthesis and transport of osmoprotectants such as proline and betaine (e.g., \u003cem\u003eproA\u003c/em\u003e, \u003cem\u003ebetB\u003c/em\u003e, and \u003cem\u003eopuA\u003c/em\u003e) were identified. Tolerance to pH fluctuations is supported by the presence of the \u003cem\u003eF₁F₀\u003c/em\u003e-ATPase (\u003cem\u003eatp\u003c/em\u003e) and Na⁺/H⁺ antiporter (\u003cem\u003enhaC\u003c/em\u003e) gene clusters. Additionally, the genome encodes two-component regulatory systems (\u003cem\u003ecomP/comA\u003c/em\u003e, \u003cem\u003edegS/degU\u003c/em\u003e) and the alternative sigma factor SigB, which enable rapid adaptation to environmental changes. Overall, this genetic arsenal explains the robust environmental adaptability of Ba. YN. J3.\u003c/p\u003e\u003c/div\u003e\u003c/div\u003e\u003cdiv id=\"Sec24\" class=\"Section2\"\u003e\u003ch2\u003e3.7 Comparative Genomic Analysis of Ba. YN. J3 within nine \u003cem\u003eBacillus\u003c/em\u003e strains\u003c/h2\u003e\u003cp\u003eTo investigate the evolutionary context of Ba. YN. J3, a comparative genomic analysis was conducted against nine representative \u003cem\u003eBacillus\u003c/em\u003e and \u003cem\u003ePriestia\u003c/em\u003e strains (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e5\u003c/span\u003e). A phylogenetic tree based on whole-genome single-nucleotide polymorphisms (SNPs) revealed that Ba. YN. J3 clustered within a well-supported clade (bootstrap value\u0026thinsp;=\u0026thinsp;88) together with the well-characterized biocontrol strains \u003cem\u003eB. velezensis\u003c/em\u003e FZB42, \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e ZKY01, and \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e GKT04, indicating a close evolutionary relationship among them (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003eB).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eGenome-wide variation analysis further clarified these relationships. Pairwise comparisons revealed extensive SNP divergence between Ba. YN. J3 and the reference genomes, ranging from approximately 2,000 SNPs (vs. \u003cem\u003eB. cereus\u003c/em\u003e FORC_047) to more than 189,000 SNPs (vs. \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e DSM7) (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003eA; Table S3). Similarly, the number of insertion/deletion (InDel) events increased with phylogenetic distance, from 27 (vs. \u003cem\u003eB. cereus\u003c/em\u003e FORC_047) to 1,194 (vs. \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e DSM7) (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003eC).\u003c/p\u003e\u003cp\u003eStructural variation (SV) analysis provided a macroscopic view of genome organization and synteny (Figure \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e). Strong collinearity was observed between Ba. YN. J3 and its closest relatives (e.g., \u003cem\u003eB. velezensis\u003c/em\u003e FZB42), although several local rearrangements such as inversions and translocations were evident. In contrast, comparisons with more distantly related species (\u003cem\u003eP. megaterium\u003c/em\u003e A14581 and \u003cem\u003eB. cereus\u003c/em\u003e FORC_047) revealed extensive genomic rearrangements and large-scale segmental losses, consistent with their distinct phylogenetic positions.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec25\" class=\"Section2\"\u003e\u003ch2\u003e3.7 Gene family analysis of Ba. YN. J3 and nine comparative \u003cem\u003eBacillus\u003c/em\u003e strains\u003c/h2\u003e\u003cp\u003eGene family analysis was conducted to compare the functional potential of Ba. YN. J3 with nine reference strains. Among the reference genomes, the number of gene families ranged from 2,271 in \u003cem\u003eB. cereus\u003c/em\u003e FORC_047 to 2,932 in \u003cem\u003eB. subtilis\u003c/em\u003e 168 (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003eD). The genome of Ba. YN. J3 contained 4,541 genes, of which 3,846 (84.7%) were clustered into 2,836 gene families.\u003c/p\u003e\u003cp\u003eA key outcome of this comparative analysis was the identification of strain-specific (unique) gene families. The number of unique gene families among the reference strains ranged from 0 to 184, with \u003cem\u003eB. cereus\u003c/em\u003e FORC_047 possessing the most. In contrast, Ba. YN. J3 encoded ten unique gene families. Although eight of these genes could not be functionally annotated, two were identified as being critically associated with key biocontrol and plant growth-promoting (PGP) traits: the \u003cem\u003eflgC\u003c/em\u003e gene family, encoding a flagellar basal body rod proteins involved in chemotaxis, and the \u003cem\u003eyutI\u003c/em\u003e gene family, which encodes a putative nitrogen fixation regulatory proteins. These unique gene families may therefore contribute to the specific beneficial functions observed in Ba. YN. J3.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec26\" class=\"Section2\"\u003e\u003ch2\u003e3.8 Core‒pan analysis between Ba. YN. J3 and nine comparative \u003cem\u003eBacillus strains\u003c/em\u003e\u003c/h2\u003e\u003cp\u003eTo further explore the genomic basis of the unique traits of Ba. YN. J3, a core-pan analysis was conducted with nine reference \u003cem\u003eBacillus\u003c/em\u003e strains. The analysis identified a core genome of 1,097 genes, which were highly conserved across all ten strains and were primarily involved in essential cellular processes such as primary metabolism, genetic information processing, and environmental adaptation.\u003c/p\u003e\u003cp\u003eIn addition to the core genome, Ba. YN. J3 possessed 830 strain-specific genes (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003eE). Of these, only 40 genes (4.82%) could be functionally annotated. Notably, these annotated unique genes were enriched in functions relevant to the strain\u0026rsquo;s ecological niche, including signal transduction, transport, resistance, and transcriptional regulation, suggesting that these genes may play important roles in the specialized biocontrol and plant-associated lifestyle of Ba. YN. J3.\u003c/p\u003e\u003c/div\u003e"},{"header":"4. Discussion","content":"\u003cp\u003eIn this study, we demonstrated that the oat endophyte \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e Ba. YN. J3 is a potent strain that exhibits both biocontrol and plant growth-promoting (PGP) functions. Its strong efficacy against oat anthracnose is attributed to multiple synergistic mechanisms, including inhibition of fungal mycelial growth through both volatile (VOCs) and non-volatile metabolites, degradation of fungal cell walls, and induction of host defense enzymes. In addition, Ba. YN. J3 significantly enhanced the growth of multiple crop species. Whole-genome and comparative genomic analyses revealed the genetic basis for these traits, identifying two unique gene families related to chemotaxis (\u003cem\u003eflgC\u003c/em\u003e) and nitrogen fixation (\u003cem\u003eyutI\u003c/em\u003e), as well as 40 strain-specific genes associated with plant-associated functions such as signal transduction, transport, resistance, and transcriptional regulation. Together, these findings establish a clear link between the unique genomic repertoire of Ba. YN. J3 and its dual biocontrol and PGP capacities.\u003c/p\u003e\u003cp\u003eMembers of the genus \u003cem\u003eBacillus\u003c/em\u003e are well-known biocontrol agents that suppress plant pathogens through the secretion of diverse antifungal compounds, which inhibit mycelial growth and spore germination [\u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e68\u003c/span\u003e]. The principal molecular mechanisms underlying this antagonism include the production of lipopeptides, siderophores, and cell wall-degrading enzymes (CWDEs) [\u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e69\u003c/span\u003e, \u003cspan citationid=\"CR70\" class=\"CitationRef\"\u003e70\u003c/span\u003e]. Consistent with this paradigm, Ba. YN. J3 exhibited broad-spectrum antifungal activity, with its metabolites causing fungal cell wall disruption and cytoplasmic leakage. Genomic analyses corroborated these observations, revealing a rich set of genes involved in biocontrol, including those encoding CWDEs, siderophore biosynthetic enzymes, and secondary metabolite biosynthesis. Specifically, thirteen biosynthetic gene clusters (BGCs) were identified, five of which encode nonribosomal peptide synthetases (NRPSs), suggesting that the antifungal activity of Ba. YN. J3 results from the synergistic action of CWDEs, siderophores, and NRPS-derived lipopeptides.\u003c/p\u003e\u003cp\u003eConsistent with its strong antagonistic activity, the Ba. YN. J3 genome contains BGCs for the three major families of \u003cem\u003eBacillus\u003c/em\u003e lipopeptides\u0026mdash;surfactin, iturin, and fengycin\u0026mdash;synthesized by NRPS and PKS complexes. These compounds inhibit phytopathogens such as \u003cem\u003eBotrytis dothidea\u003c/em\u003e and \u003cem\u003eFusarium graminearum\u003c/em\u003e by disrupting fungal membranes [\u003cspan additionalcitationids=\"CR72 CR73\" citationid=\"CR71\" class=\"CitationRef\"\u003e71\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR74\" class=\"CitationRef\"\u003e74\u003c/span\u003e]. The surfactin cluster is regulated by quorum-sensing components ComX, ComA/ComP, and RapC [\u003cspan citationid=\"CR75\" class=\"CitationRef\"\u003e75\u003c/span\u003e, \u003cspan citationid=\"CR76\" class=\"CitationRef\"\u003e76\u003c/span\u003e], whereas the iturin A cluster, controlled by DegU and DegQ, directs the synthesis of potent antifungal peptides [\u003cspan citationid=\"CR77\" class=\"CitationRef\"\u003e77\u003c/span\u003e]. The complete fengycin cluster (\u003cem\u003eppsA\u0026ndash;E\u003c/em\u003e) further underscores the antifungal potential of this strain. In addition to these canonical clusters, additional BGCs for locillomycin, macrolactin, bacillaene, and difficidin broaden the antimicrobial spectrum of Ba. YN. J3. This diverse array of secondary metabolites provides a strong molecular basis for its biocontrol efficiency.\u003c/p\u003e\u003cp\u003eIron acquisition via siderophore production represents another key mechanism of pathogen suppression [\u003cspan citationid=\"CR78\" class=\"CitationRef\"\u003e78\u003c/span\u003e]. Ba. YN. J3 produces siderophores, as indicated by clear halos on Chrome Azurol S (CAS) agar, and harbors essential siderophore biosynthetic genes (\u003cem\u003efhuA\u003c/em\u003e, \u003cem\u003efhuB\u003c/em\u003e, \u003cem\u003efhuC\u003c/em\u003e, \u003cem\u003efhuD\u003c/em\u003e, and \u003cem\u003efhuG\u003c/em\u003e). Similar siderophore-mediated biocontrol mechanisms have been reported in \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e TA-1 [\u003cspan citationid=\"CR79\" class=\"CitationRef\"\u003e79\u003c/span\u003e] and \u003cem\u003eB. subtilis\u003c/em\u003e MBI 600 [\u003cspan citationid=\"CR80\" class=\"CitationRef\"\u003e80\u003c/span\u003e], as well as in other \u003cem\u003eBacillus\u003c/em\u003e species [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e, \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e, \u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e]. These findings collectively confirm that siderophore-mediated iron competition constitutes a critical component of the antagonistic repertoire of Ba. YN. J3.\u003c/p\u003e\u003cp\u003eBa. YN. J3 also promotes plant growth through multiple pathways. Indole-3-acetic acid (IAA) biosynthesis genes (\u003cem\u003etrpA\u0026ndash;P\u003c/em\u003e) were identified in its genome, consistent with previous reports of its IAA-producing potential [\u003cspan citationid=\"CR81\" class=\"CitationRef\"\u003e81\u003c/span\u003e, \u003cspan citationid=\"CR82\" class=\"CitationRef\"\u003e82\u003c/span\u003e]. Furthermore, the strain was capable of growth on nitrogen-free, phosphorus-, and potassium-solubilizing media, reflecting its nutrient-cycling capacity. Genome annotation revealed key genes involved in nitrogen fixation (\u003cem\u003eyutI\u003c/em\u003e, \u003cem\u003enifU\u003c/em\u003e, \u003cem\u003enifS\u003c/em\u003e, \u003cem\u003enifF\u003c/em\u003e, \u003cem\u003egltB\u003c/em\u003e, \u003cem\u003enarK\u003c/em\u003e), nitrogen metabolism (\u003cem\u003enasE\u003c/em\u003e, \u003cem\u003enasD\u003c/em\u003e, \u003cem\u003egudB\u003c/em\u003e, \u003cem\u003erocG\u003c/em\u003e), phosphorus solubilization (\u003cem\u003epstA\u003c/em\u003e, \u003cem\u003epstB1\u003c/em\u003e, \u003cem\u003epstB2\u003c/em\u003e, \u003cem\u003epstC\u003c/em\u003e, \u003cem\u003epstS\u003c/em\u003e, \u003cem\u003eugpC\u003c/em\u003e, \u003cem\u003epykF\u003c/em\u003e), and potassium metabolism (\u003cem\u003ekimA\u003c/em\u003e, \u003cem\u003ektrC\u003c/em\u003e, \u003cem\u003etrkA\u003c/em\u003e, \u003cem\u003ekdpD\u003c/em\u003e). Comparative genomics further revealed a unique nitrogen fixation gene family, suggesting that Ba. YN. J3 promotes plant growth through IAA secretion and mobilization of essential nutrients.\u003c/p\u003e\u003cp\u003eBa. YN. J3 also exhibited remarkable tolerance to abiotic stress, thriving under a wide range of conditions, including high temperatures (10\u0026ndash;45\u0026deg;C), pH (5.0\u0026ndash;9.0), salinity (up to 7% NaCl), and osmotic stress (up to 11% PEG6000). Genomic analysis revealed numerous stress-response genes, including those encoding heat shock proteins (GroEL, DnaK, DnaJ, ClpB), osmotic adaptation proteins (OpuA, ProP, TreB), salt tolerance proteins (ProABC, BetAB, CspABC), and pH homeostasis proteins (AtpABCDEFGH, NhaC) [\u003cspan additionalcitationids=\"CR84 CR85\" citationid=\"CR83\" class=\"CitationRef\"\u003e83\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR86\" class=\"CitationRef\"\u003e86\u003c/span\u003e]. Regulatory elements such as the sigma factor SigB and two-component systems (DegS/DegU and ComP/ComA) [\u003cspan citationid=\"CR87\" class=\"CitationRef\"\u003e87\u003c/span\u003e] further enhance environmental adaptability by fine-tuning stress-responsive gene expression.\u003c/p\u003e\u003cp\u003eChemotaxis, the directed movement toward chemical stimuli, is essential for host colonization and environmental sensing [\u003cspan citationid=\"CR88\" class=\"CitationRef\"\u003e88\u003c/span\u003e, \u003cspan citationid=\"CR89\" class=\"CitationRef\"\u003e89\u003c/span\u003e]. Previous studies demonstrated that Ba. YN. J3 actively migrates toward \u003cem\u003eColletotrichum cereale\u003c/em\u003e hyphae [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. Genomic analysis revealed a complete chemotaxis and flagellar assembly system, including \u003cem\u003emcp\u003c/em\u003e, \u003cem\u003echeA\u003c/em\u003e, \u003cem\u003echeB\u003c/em\u003e, \u003cem\u003echeC\u003c/em\u003e, \u003cem\u003echeD\u003c/em\u003e, \u003cem\u003echeR\u003c/em\u003e, \u003cem\u003echeW\u003c/em\u003e, \u003cem\u003efliC\u003c/em\u003e, \u003cem\u003efliD\u003c/em\u003e, \u003cem\u003eflhA\u003c/em\u003e, \u003cem\u003eflhB\u003c/em\u003e, \u003cem\u003eflgB\u0026ndash;D\u003c/em\u003e, and \u003cem\u003emotA/B\u003c/em\u003e, suggesting that its robust motility and environmental sensing capacity may underlie host-targeting behavior.\u003c/p\u003e\u003cp\u003eComparative genomics integrating SNP, InDel, SV, and core/pan-genome analyses provided further insights into the genetic basis of strain-specific traits [\u003cspan citationid=\"CR90\" class=\"CitationRef\"\u003e90\u003c/span\u003e]. Ba. YN. J3 is closely related to well-characterized biocontrol strains such as \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e GKT04, \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e ZKY01, and \u003cem\u003eB. velezensis\u003c/em\u003e FZB42, and shares functional features of antagonism and rhizosphere colonization [\u003cspan citationid=\"CR91\" class=\"CitationRef\"\u003e91\u003c/span\u003e]. However, its genome also contains 961,698 nonsynonymous mutations affecting key functional genes for lipopeptide synthesis, hormone production (\u003cem\u003eipdC\u003c/em\u003e), nutrient solubilization (\u003cem\u003ephoA\u003c/em\u003e, \u003cem\u003eppk\u003c/em\u003e), nitrogen fixation (\u003cem\u003enif\u003c/em\u003e), and siderophore biosynthesis. Such variations are known to modulate metabolite diversity [\u003cspan citationid=\"CR92\" class=\"CitationRef\"\u003e92\u003c/span\u003e], exopolysaccharide (EPS) formation [\u003cspan citationid=\"CR93\" class=\"CitationRef\"\u003e93\u003c/span\u003e], motility [\u003cspan citationid=\"CR94\" class=\"CitationRef\"\u003e94\u003c/span\u003e], and iron acquisition [\u003cspan citationid=\"CR95\" class=\"CitationRef\"\u003e95\u003c/span\u003e], and are likely to contribute to the unique functional profile and ecological adaptability of Ba. YN. J3 [\u003cspan citationid=\"CR96\" class=\"CitationRef\"\u003e96\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eAlthough only 7.4% of the 447 strain-specific genes of Ba. YN. J3 were functionally annotated, these genes underscore its ecological specialization. Many contribute to nutrient acquisition and stress resistance. For example, \u003cem\u003emalL\u003c/em\u003e [\u003cspan citationid=\"CR97\" class=\"CitationRef\"\u003e97\u003c/span\u003e] correlates with strong hydrolytic enzyme activity, enabling efficient degradation of environmental macromolecules. Other unique genes, including \u003cem\u003eligA\u003c/em\u003e [\u003cspan citationid=\"CR98\" class=\"CitationRef\"\u003e98\u003c/span\u003e], quorum-sensing regulators \u003cem\u003ecomP\u0026ndash;comA\u003c/em\u003e [\u003cspan citationid=\"CR99\" class=\"CitationRef\"\u003e99\u003c/span\u003e], and small acid-soluble spore proteins (SASPs) [\u003cspan citationid=\"CR100\" class=\"CitationRef\"\u003e100\u003c/span\u003e], collectively support genome integrity, communication, and long-term survival [\u003cspan citationid=\"CR101\" class=\"CitationRef\"\u003e101\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eTwo experimentally validated unique genes, \u003cem\u003eflgC\u003c/em\u003e (chemotaxis) and \u003cem\u003eyutI\u003c/em\u003e (nitrogen fixation), define the distinct ecological strategies of Ba. YN. J3. The \u003cem\u003eflgC\u003c/em\u003e gene mediates directed motility toward \u003cem\u003eC. cereale\u003c/em\u003e spores [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR102\" class=\"CitationRef\"\u003e102\u003c/span\u003e], whereas \u003cem\u003eyutI\u003c/em\u003e confers the ability to fix nitrogen and grow independently in nutrient-limited conditions [\u003cspan citationid=\"CR103\" class=\"CitationRef\"\u003e103\u003c/span\u003e, \u003cspan citationid=\"CR104\" class=\"CitationRef\"\u003e104\u003c/span\u003e]. These complementary traits equip Ba. YN. J3 with the dual capacity to pursue its pathogenic targets while maintaining nutritional self-sufficiency, underscoring its potential as a robust and environmentally adaptable biocontrol agent.\u003c/p\u003e"},{"header":"Conclusions","content":"\u003cp\u003eThis study identifies the oat endophyte \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e Ba. YN. J3 as a potent dual-function strain exhibiting both biocontrol and plant growth-promoting (PGP) activities. Phenotypic assays confirmed its broad-spectrum antifungal effects, secretion of multiple cell wall-degrading enzymes (CWDEs), and its ability to enhance the growth of various crop species while inducing systemic resistance.\u003c/p\u003e\u003cp\u003eWhole-genome sequencing provided a clear molecular foundation for these traits, revealing a 4.06 Mb circular chromosome enriched with biosynthetic gene clusters for key antifungal secondary metabolites (such as surfactin and fengycin), PGP-related pathways (including IAA biosynthesis and nutrient cycling), and genes conferring tolerance to abiotic stresses.\u003c/p\u003e\u003cp\u003eComparative genomic analysis identified 830 strain-specific genes, including two unique gene families encoding the flagellar rod protein FlgC (involved in chemotaxis) and the nitrogen fixation regulatory protein YutI. These unique genetic determinants provide a mechanistic explanation for the strain\u0026rsquo;s dual functionality, linking targeted motility to its antagonistic activity and nitrogen fixation to its nutritional self-sufficiency.\u003c/p\u003e\u003cp\u003eMoreover, previous greenhouse and field experiments demonstrated that \u003cem\u003eB. amyloliquefaciens\u003c/em\u003e Ba. YN. J3 exhibited strong biocontrol efficacy against \u003cem\u003eColletotrichum cereale\u003c/em\u003e, further validating its potential as an effective and reliable biocontrol agent. Collectively, these findings highlight Ba. YN. J3 as a promising and environmentally resilient candidate for the development of sustainable biopesticides and biofertilizers in modern agriculture.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets presented in this study can be found in online repositories. The name of the repository and accession number can be found below: NCBI GenBank, accession number: PRJNA1171741 (https://www.ncbi.nlm.nih.gov/bioproject/1171741). Further inquiries can be directed to the corresponding author. All experimental data and materials related to this study are also available from the corresponding author, Bao-zhu Dong, upon reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis work was supported by the Science and Technology Program of Inner Mongolia Autonomous Region, \u0026ldquo;Research and Application of Oat Intertillage Weeding and Chemical Herbicide Reduction and Synergistic Efficiency Enhancement Technology\u0026rdquo; (Grant No. 2025YFHH0165); the Basic Research Fund for Universities Directly Affiliated with Inner Mongolia Autonomous Region (Grant No. BR251033); the Central Government-Guided Local Science and Technology Development Fund, \u0026ldquo;Research on Green Production of Oat Grains and High-Quality Compound Feed Processing Technology\u0026rdquo; (Grant No. 2022ZY0060) and \u0026ldquo;Application and Promotion of Green Cultivation Technologies for Coarse Cereals in Qingshuihe County\u0026rdquo; (Grant No. 2022ZY0065); the National Key R\u0026amp;D Program of China, \u0026ldquo;Research and Demonstration of Green Control Technologies for Diseases, Insect Pests, and Weeds in Minor Grains\u0026rdquo; (Grant No. 2023YFD1600701-5); and the China Agriculture Research System for Oat and Buckwheat (Grant No. CARS-07-C-3).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors\u0026apos; contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWei Quan: Conceptualization, Methodology, Investigation, Formal Analysis, Writing \u0026ndash; Original Draft, Data Curation, Validation.\u003c/p\u003e\n\u003cp\u003eCheng-Zhong Zheng: Investigation, Formal Analysis.\u003c/p\u003e\n\u003cp\u003eMuhammad Ayaz: Investigation, Software, Visualization, Formal Analysis.\u003c/p\u003e\n\u003cp\u003eChen Hui: Investigation, Formal Analysis.\u003c/p\u003e\n\u003cp\u003eChun-Yang Wang: Investigation, Software, Visualization.\u003c/p\u003e\n\u003cp\u003eChen-Lu Liu: Investigation, Software, Visualization, Formal Analysis, Data Curation, Validation.\u003c/p\u003e\n\u003cp\u003eZhi-Gang Liu: Software, Visualization, Formal Analysis.\u003c/p\u003e\n\u003cp\u003eBao-Zhu Dong: Supervision, Project Administration, Funding Acquisition, Conceptualization, Writing \u0026ndash; Review \u0026amp; Editing.\u003c/p\u003e\n\u003cp\u003eHong-You Zhou: Supervision, Project Administration, Funding Acquisition, Conceptualization, Writing \u0026ndash; Review \u0026amp; Editing.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe gratefully acknowledge financial support from the Ministry of Agriculture and Rural Affairs, the Ministry of Science and Technology, the Ministry of Education of the People\u0026apos;s Republic of China through the Science and Technology Program of the Inner Mongolia Autonomous Region, the Basic Research Fund for Universities Directly Affiliated with Inner Mongolia, the Central Government-Guided Local Science and Technology Development Fund, the National Key R\u0026amp;D Program of China, and the China Agriculture Research System for Oat and Buckwheat.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eWilson D. 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Scientifica. 2012;2012(1):963401.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"bmc-genomics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"gics","sideBox":"Learn more about [BMC Genomics](http://bmcgenomics.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/gics","title":"BMC Genomics","twitterHandle":"#BMCGenomics","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Bacillus amyloliquefaciens, Endophyte, Biocontrol, Plant growth promotion, Whole-genome sequencing, Comparative genomics","lastPublishedDoi":"10.21203/rs.3.rs-8097513/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8097513/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e\u003cp\u003eEndophytic bacteria serve as important resources for the development of novel biocontrol agents for sustainable agriculture. The present study provides a detailed characterization of a newly isolated oat endophyte, \u003cem\u003eBacillus amyloliquefaciens\u003c/em\u003e Ba. YN. J3, using an integrated analysis of phenotypic, genomic, and comparative genomic data to uncover its biocontrol and plant growth-promoting (PGP) potential.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e\u003cp\u003eThe current findings indicate that Ba. YN. J3 possessed efficient PGP and biocontrol potential both \u003cem\u003ein vitro\u003c/em\u003e and \u003cem\u003ein planta\u003c/em\u003e. Additionally, Ba. YN. J3 showed broad-spectrum antifungal activity against six major phytopathogens and was found to produce multiple cell wall-degrading enzymes (CWDEs) and siderophores, significantly increasing the growth of several crop species and regulating defense enzymes in oats. The complete 4.06 Mb genome of Ba. YN. J3 contains numerous gene clusters encoding vital secondary metabolites (e.g., surfactin, fengycin), CWDEs, and proteins associated with PGP functions and chemotaxis. The genome also harbors a robust set of genes related to abiotic stress tolerance, suggesting its potential to survive and function effectively in challenging field environments. Furthermore, comparative genomic analysis revealed 830 strain-specific genes, including two unique gene families critically linked to chemotaxis (flagellar rod protein FlgC) and nitrogen fixation (regulatory protein YutI).\u003c/p\u003e\u003ch2\u003eConclusions\u003c/h2\u003e\u003cp\u003eThis integrated study elucidates the potent dual function of Ba. YN. J3 and its unique genetic determinants. The \u003cem\u003eflgC\u003c/em\u003e and \u003cem\u003eyutI\u003c/em\u003e gene families, in particular, offer novel insights into the molecular basis of its targeted antagonism and nutritional self-sufficiency. Hence, the current findings highlight Ba. YN. J3 as a promising candidate for the development of effective biopesticides and biofertilizers.\u003c/p\u003e","manuscriptTitle":"Whole-Genome Analysis Reveals the Growth-Promoting and Biocontrol Potential of Bacillus amyloliquefaciens Ba. YN. 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