Search Results (39)

The endophytic bacterium Bacillus subtilis 10-4 is a potent bioinoculant, previously shown to enhance growth and resilience to abiotic/biotic stresses across various crops. However, the genetic basis underlying these beneficial traits remains unexplored. In this study, a whole-genome analysis of B. subtilis 10-4 was performed to gain the molecular determinants of its plant-beneficial effects. The Illumina MiSeq-based assembly revealed a genome of 4,278,582 bp (43.5% GC content) distributed across 19 contigs, encoding 4314 predicted protein-coding sequences, 42 tRNAs, and 6 rRNAs. This genomic architecture is comparable to other sequenced B. subtilis strains. The genomic annotation identified 331 metabolic subsystems with a total number of 1668 functions, predominantly associated with amino acid (281) (16.9%) and carbohydrate (247) (14.9%) metabolism. In silico genomic analysis uncovered a diverse repertoire of genes significant for plant growth and stress resilience. These included genes for colonization (i.e., exopolysaccharide production, biofilm formation, adhesion, motility, and chemotaxis), nutrient acquisition (i.e., nitrogen, phosphorus, iron, potassium, and sulfur metabolisms), and synthesis of bioactive compounds (auxins, salicylic acid, siderophores, gamma-aminobutyric acid, vitamins, and volatiles) and antimicrobials. The latter was supported by identified biosynthetic gene clusters (BGCs) for known antimicrobials (100% similarity) bacilysin, bacillaene, subtilosin A, and bacillibactin, as well as clusters for surfactin (82%), fengycin (80%), and plipastatin (46%), alongside a unique terpene cluster with no known similarity. Additionally, genes conferring abiotic stress tolerance via glutathione metabolism, osmoprotectants (e.g., proline, glycine betaine), detoxification, and general stress response were identified. The genomic evidence was consistent with observed plant growth improvements in laboratory assays (radish, oat) and a field trial (wheat) upon 10-4 inoculation. Thus, the findings elucidate the genomic background of B. subtilis 10-4’s beneficial effects, solidifying its potential for utilization as a bioinoculant in sustainable crop production under changing climate accompanied by multiple environmental stresses. Full article

Longya Lily (Lilium brownii var. viridulum) bulb rot, a devastating soil-borne disease caused by Fusarium oxysporum f. sp. lilii (Fol L1-1), severely compromises yield and quality of this economically significant crop. In this study, strain R12 was isolated from the rhizosphere soil of asymptomatic Longya lily plants and identified as Bacillus velezensis. The strain significantly disrupted the spore germination and hyphal morphology of Fol L1-1. In pot experiments, R12 not only effectively suppressed disease development but also promoted plant growth, a trait potentially linked to its indole-3-acetic acid (IAA) production capacity. Genomic analysis revealed a 4,015,523 bp circular chromosome (46.42% GC content) harboring gene clusters for the synthesis of diverse secondary metabolites, including surfactin, fengycin, difficidin, and bacillibactin. These findings highlight the potential of B. velezensis R12 as a biocontrol agent and provide insights into its mechanisms for suppressing phytopathogens and promoting plant growth. Full article

The Altai Republic remains a geographic region with an uncovered microbial diversity hiding yet undescribed potential species. Here, we describe the strain al37.1^T from the Altai soil. It showed genomic similarity with the Bacillus mycoides strain DSM 2048^T. However, the in silico DNA–DNA hybridization (DDH) was 61.6%, which satisfies the accepted threshold for delineating species. The isolate formed circular, smooth colonies, in contrast to the rhizoidal morphology typical of B. mycoides. The strain showed optimal growth under the following conditions: pH 6.5, NaCl concentration 0.5% w/v, and +30 °C. The major fraction of fatty acids was composed of C_16:0 (34.77%), C_18:1 (15.20%), C_14:0 (9.06%), and C_18:0 (7.88%), which were sufficiently lower in DSM 2048^T (C_16:0–15.6%, C_14:0–3.7%). In contrast to DSM 2048^T, al37.1^T utilized glycerol, D-mannose, and D-galactose, while being unable to assimilate D-sorbitol, D-melibiose, and D-raffinose. The strain contains biosynthetic gene clusters (BGCs) associated with the production of fengycin, bacillibactin, petrobactin, and paeninodin, as well as loci coding for insecticidal factors, such as Spp1Aa, chitinases, Bmp1, and InhA1/InhA2. The comparative analysis with the 300 closest genomes demonstrated that these BGCs and Spp1Aa could be considered core for the whole group. Most of the strains, coupled with al37.1^T, contained full nheABC and hblABC operons orchestrating the synthesis of enteric toxins. We observed a cytotoxic effect (≈19 and 22% reduction in viability) of the strain on the PANC-1 cell line. Given the unique morphological features and genome-derived data, we propose a new species, B. altaicus, represented by the type strain al37.1^T. Full article

Maize stalk rot (MSR) is one of the most devastating fungal diseases affecting maize worldwide. In recent years, biological control agents have emerged as an environmentally friendly and highly attractive strategy for managing MSR. In this study, two Bacillus strains—B. subtilis KP3P9 and B. siamensis K13C—were shown to effectively inhibit the growth of the MSR pathogen Fusarium graminearum in vitro. Pot experiments showed that inoculation with KP3P9 significantly increased plant height, stem width, above-ground part fresh weight, and total plant fresh weight, whereas K13C significantly improved the stem width and under-ground part fresh weight of maize seedlings (p < 0.05), demonstrating their plant-growth-promoting potential. Moreover, both strains markedly reduced the disease severity indices (DSIs) of maize seedlings, indicating that they can enhance maize resistance to the pathogen. Whole-genome sequencing using Oxford Nanopore (ONT) and Illumina technologies showed that the complete genomes of KP3P9 and K13C contained biosynthetic gene clusters involved in the biosynthesis of antimicrobial secondary metabolites, including fengycin, bacillibactin, subtilin, pulcherriminic acid, subtilosin A, bacilysin, and others. Moreover, both strains exhibited strong antagonistic activity against F. solani (the causal pathogen of apple replant disease), as well as F. oxysporum f. sp. cubense race 1 (Foc1) and tropical race 4 (FocTR4) (pathogens responsible for banana wilt disease), with inhibition rates exceeding 70% in vitro. These results indicate that KP3P9 and K13C are promising biocontrol agents for MSR and other devastating Fusarium diseases. Full article

Bacillus cereus AS_3 and Bacillus cereus AS_5 are bacterial endophytes isolated from sterilized leaves of the medical plant Alectra sessiliflora, which were previously identified using 16S rRNA sequencing. Here, we present the whole-genome sequencing and annotation of strains AS_3 and AS_5, the first genome report of Bacillus cereus strains from A. sessiliflora. The genome of strain AS_3 has 59 contigs, 5 503 542 bp draft circular chromosome, an N₅₀ of 211,274 bp, and an average G+C content of 35.2%; whereas strain AS_5 has 38 contigs, 5,510,121 bp draft circular chromosome, an N₅₀ of 536,033 bp, and an average G+C content of 35.2%. A total of 5679 protein-coding genes, 62 genes coding for RNAs, and 122 pseudogenes in the strain AS_3 genome were identified by the National Center for Biotechnology Information Prokaryotic Annotation pipeline, whereas a total of 5688 gene protein-coding genes were identified in AS_5, with 60 genes coding for RNAs and 120 pseudogenes. Phenotypic analysis and whole-genome sequencing analysis showed that AS_3 and AS_5 share similar characteristics, including Gram-positive, motile, rod-shaped, and endospore-forming have shown a high sequence similarity with Bacillus cereus, type strain ATCC 14579^T. Strains AS_3 and AS_5 had genomic digital DNA–DNA hybridization (dDDH) with the type strain Bacillus cereus ATCC 14579^T of 85.8% and 86%, respectively, and average nucleotide identities (ANIs) of 98% and 98.01%, respectively. Phylogenomic analysis confirmed that strains AS_3 and AS_5 share very similar genomic and phenotypic characteristics, and are closely related to the type strain Bacillus cereus type strain ATCC 14579^T, supporting their classification within the Bacillus cereus species. A total of 10 secondary metabolite gene clusters, including siderophore type petrobactin, terpene type molybdenum cofactor, non-ribosomal peptide synthetase (NRPS) type bacillibactin, and β-lactone type fengycin, were predicted using AntiSMASH software (version 5.0). Putative genes potentially involved in bioremediation and endophytic lifestyle were identified in the genome analysis. Genome sequencing of Bacillus cereus AS_3 and Bacillus cereus AS_5 has provided genomic information and demonstrated potential biotechnological applications. Full article

Bacillus amyloliquefaciens is a well-recognized biocontrol agent and plant growth promoter. This study characterized the endophytic B. amyloliquefaciens LJ1, isolated from Nanguo pear fruit, through whole-genome sequencing and functional analyses. The B. amyloliquefaciens LJ1 genome (3,947,365 bp, 46.48% GC content) encodes 3757 protein-coding sequences. Genomic analysis revealed diverse carbohydrate-active enzymes (CAZymes) and 12 secondary metabolite biosynthetic gene clusters, including those potentially producing surfactin, fengycin, bacillibactin, and bacilysin. Safety assessments, including hemolysis, indole production, biogenic amine production, and a 21 day mice-feeding trial, indicated no adverse effects, suggesting B. amyloliquefaciens LJ1 is non-pathogenic. In vitro assays demonstrated significant inhibitory activity against Penicillium expansum, a major post-harvest pathogen, by suppressing spore germination and germ-tube elongation. These results suggest that B. amyloliquefaciens LJ1 possesses significant biocontrol potential and could be a promising agent for sustainable disease management in Nanguo pear and potentially other crops. Full article

Dietary protein levels greatly influence gut microbial ecosystems; however, their effects on gut archaea and associated functions in ruminants require further elucidation. This study evaluated the impact of varying dietary protein levels on gut archaeal composition, antimicrobial resistance (AMR) genes, virulence factors, and functional capacities in sheep. Eighteen ewes (Yunnan semi-fine wool breed, uniparous, 2 years old, and averaging 50 ± 2 kg body weight) were randomly assigned to diets containing an 8.5 (low; H_1), 10.3 (medium; H_m), or 13.9% (high; H_h) crude protein level from the 35th day of pregnancy to the 90th day postpartum. The total duration of the experiment was approximately 202 days. A total of nine fecal samples (three from each group) were analyzed via 16S rRNA and metagenomics sequencing. Higher archaeal alpha diversity and richness were observed in the H_m and H_h groups compared to the H_l group (p < 0.05). A Beta diversity analysis revealed the archaeal community’s distinct clustering mode based on protein levels. The methanogenic genera Methanobrevibacter and Methanocorpusculum were dominant across the three groups, and their abundance was influenced by protein intake. A functional prediction analysis indicated moderate changes in amino acid and carbohydrate metabolism, which are particularly associated with methane production, an important source of greenhouse gases. AMR genes (e.g., tetA (60), patA, vat, and Erm methyltransferase) and virulence factors (Bacillibactin, LPS) were significantly enriched when animals were fed high-protein diets. Our results demonstrated that dietary protein levels significantly influence gut archaeal composition, AMR gene enrichment, and related functional pathways. Medium-protein diets promoted greater archaeal diversity, whereas high-protein diets favored resistance gene proliferation and enhanced methanogenic activity. Optimizing dietary protein intake may enhance gut health, mitigate antimicrobial resistance risk, and reduce methane emissions, thereby supporting livestock sustainability and environmental protection. Full article

Spore-forming Bacilli have been explored due to their potential biotechnological features and applications in human health and functional food research. This study focuses on the genetic and phenotypical characterization of the functional probiotic properties of Bacillus velezensis P45, a strain isolated from fish intestines. B. velezensis P45 exhibited antimicrobial activity against Gram-positive and Gram-negative pathogens and demonstrated strong autoaggregation and biofilm formation properties in vitro. The strain also showed tolerance to gastrointestinal conditions and ability to metabolize and adhere to mucin. In silico analysis confirmed the absence of virulence factors and antibiotic resistance genes, reinforcing its safety as a probiotic candidate. Genome mining revealed the presence of genes related to adhesion, such as fibronectin-binding protein and enolases, and for the synthesis of secondary metabolites, including the antimicrobial lipopeptides fengycin, surfactin, and bacillibactin. In addition, phylogenetic comparison using the yloA (rqcH) gene associated with gut adhesion clustered strain P45 with other probiotic Bacillus and B. velezensis strains, while separating it from pathogenic bacteria. Thus, the strain B. velezensis P45 could be a valuable candidate as a probiotic due to its functional properties and safety. Full article

Aquaculture is one of the fastest-growing sectors in food production. The widespread use of antibiotics in fish farming has been identified as a driver for the development of antibiotic resistance. One of the promising approaches to solving this problem is the use of probiotics. There are many promising aquaculture probiotics in the Bacillus genus, which produces non-ribosomal peptides (NRPs). NRPs are known as antimicrobial agents, although evidence is gradually accumulating that they may have other effects, especially at lower (subinhibitory) concentrations. The mechanisms of action of many NRPs remain unexplored, and molecular docking and molecular dynamics studies are invaluable tools for studying such mechanisms. The purpose of this study was to investigate the in silico inhibition of crucial bacterial targets by NRPs. Molecular docking analyses were conducted to assess the binding affinities of the NRPs of Bacillus for protein targets. Among the complexes evaluated, bacillibactin with glutamine synthetase, dihydrofolate reductase, and proaerolysin exhibited the lowest docking scores. Consequently, these complexes were selected for further investigation through molecular dynamics simulations. As a result, three additional potential mechanisms of action for bacillibactin were identified through in silico analyses, including the inhibition of glutamine synthetase, dihydrofolate reductase, and proaerolysin, which are critical bacterial enzymes and considered as the potential antibacterial targets. These findings were further supported by in vitro antagonism assays using bacillibactin-producing Bacillus velezensis strains MT55 and MT155, which demonstrated strong inhibitory activity against Pseudomonas aeruginosa and Aeromonas veronii. Full article

This study aims to elucidate the genomic characteristics of Bacillus subtilis MGE 2012, a strain isolated from Korean traditional fermented food, meju, which contributes to its high enzyme activity and potential applications. The whole genome sequence of B. subtilis MGE 2012 was assembled using MEGAHIT, annotated using RAST and BLASTKOALA v3.1. Phylogenetic analysis placed MGE 2012 within the Bacillus clade, showing high similarity to B. subtilis NCIB 3610 and B. subtilis ATCC 6051. AntiSMASH analysis identified 14 biosynthetic gene clusters (BGCs) capable of producing various secondary metabolites, including subtilosin, bacillibactin, fengycin, bacilysin, plipastatin, and surfactin. This study provides an overview of the whole genome and secondary metabolite profile of B. subtilis MGE 2012, emphasizing its potential applications in biotechnology. While the primary focus of this study was to explore the genomic characteristics and secondary metabolite profile, future research could delve deeper into genome mining for enzyme activities and their applications. Full article

Bacterial strain TSO55 was isolated from a commercial field of wheat (Triticum turgidum L. subsp. durum), under organic amendments, located in the Yaqui Valley, Mexico. Morphological and microscopical characterization showed off-white irregular colonies and Gram-positive bacillus, respectively. The draft genome sequence of this strain revealed a genomic size of 5,489,151 bp, with a G + C content of 35.21%, N50 value of 245,934 bp, L50 value of 8, and 85 contigs. Taxonomic affiliation showed that strain TSO55 belongs to Bacillus paranthracis, reported as an emergent human pathogen. Genome annotation identified 5743 and 5587 coding DNA sequences (CDSs), respectively, highlighting genes associated with indole production, phosphate and potassium solubilization, and iron acquisition. Further in silico analysis indicated the presence of three CDSs related to pathogenicity islands and a high pathogenic potential (77%), as well as the presence of multiple gene clusters related to antibiotic resistance. The in vitro evaluation of plant growth promotion traits was negative for indole production and phosphate and potassium solubilization, and it was positive but low (18%) for siderophore production. The biosynthetic gene cluster for bacillibactin (siderophore) biosynthesis was confirmed. Antifungal bioactivity of strain TSO55 evaluated against wheat pathogenic fungi (Alternaria alternata TF17, Bipolaris sorokiniana TPQ3, and Fusarium incarnatum TF14) showed minimal fungal inhibition. An antibiotic susceptibility assay indicated resistance to three of the six antibiotics evaluated, up to a concentration of 20 µg/mL. The beta hemolysis result on blood agar reinforced TSO55’s pathogenic potential. Inoculation of B. paranthracis TSO55 on wheat seedlings resulted in a significant decrease in root length (−8.4%), total plant height (−4.2%), root dry weight (−18.6%), stem dry weight (−11.1%), and total plant dry weight (−15.2%) compared to the control (uninoculated) treatment. This work highlights the importance of analyzing the microbiological safety of organic amendments before application. Comprehensive genome-based taxonomic affiliation and bioprospecting of microbial species introduced to the soil by organic agricultural practices and any microbial inoculant will prevent the introduction of dangerous species with non-beneficial traits for crops, which affect sustainability and generate potential health risks for plants and humans. Full article

Strain TE5 was isolated from a wheat (Triticum turgidum L. subsp. durum) rhizosphere grown in a commercial field of wheat in the Yaqui Valley in Mexico. In this work, we present strain TE5 as a promising biological control agent against Bipolaris sorokiniana. First, after its genome sequencing through Illumina NovaSeq, this strain showed a genome size of 4,262,927 bp, with a 43.74% G + C content, an N50 value of 397,059 bp, an L50 value of 4 bp, and 41 contigs (>500 bp). Taxonomical affiliation was carried out by using overall genome relatedness indexes (OGRIs) and the construction of a phylogenomic tree based on the whole genome. The results indicated that strain TE5 identifies with Bacillus cabrialesii subsp. cabrialesii. Genomic annotation using Rapid Annotation Using Subsystems Technology (RAST) and Rapid Prokaryotic Genome Annotation (Prokka) indicated the presence of 4615 coding DNA sequences (CDSs) distributed across 330 subsystems, which included gene families associated with biocontrol, stress response, and iron competition. Furthermore, when the antiSMASH 7.1 platform was used for genome mining, the results indicated the presence of seven putative biosynthetic gene clusters related to the production of biocontrol metabolites, namely subtilosin A, bacillibactin, fengycin, bacillaene, bacilysin, surfactin, and rhizocticin A. Moreover, the antifungal activity of strain TE5 and its cell-free extract (CFE) was evaluated against Bipolaris sorokiniana, an emergent wheat pathogen. The results of in vitro dual confrontation showed fungal growth inhibition of 67% by strain TE5. Additionally, its CFE almost completely inhibited (93%) the growth of the studied phytopathogenic fungus on liquid media. Further observations of the impact of these bacterial metabolites on fungal spore germination exhibited inhibition of fungal spores through degrading the germinative hypha, avoiding mycelium development. Finally, the protective effect of strain TE5 against Bipolaris sorokiniana was evaluated for wheat seedlings. The results showed a significant decrease (83%) in disease severity in comparison with the plant infection without inoculation of the biological control agent. Thus, this work proposes Bacillus cabrialesii subsp. cabrialesii strain TE5 as a promising biological control agent against the wheat pathogen Bipolaris sorokiniana while suggesting lipopeptides as the potential mode of action, together with plant growth and defense stimulation. Full article

The dispersal of plant pathogens is a threat to the global economy and food industry which necessitates the need to discover efficient biocontrol agents such as bacteria, fungi, etc., inhibiting them. Here, we describe the Bacillus mycoides strain b12.3 isolated from the soil of Olkhon Island in Lake Baikal, Russia. By applying the co-cultivation technique, we found that the strain inhibits the growth of plant pathogens, such as the bacteria Xanthomonas campestris, Clavibacter michiganensis, and Pectobacterium atrospecticum, as well as the fungus Alternaria solani. To elucidate the genomic fundament explaining these activities, we leveraged next-generation whole-genome sequencing and obtained a high-quality assembly based on short reads. The isolate bore seven known BGCs (biosynthetic gene clusters), including those responsible for producing bacillibactin, fengycin, and petrobactin. Moreover, the genome contained insecticidal genes encoding for App4Aa1, Tpp78Ba1, and Spp1Aa1 toxins, thus implicating possible pesticidal potential. We compared the genome with the 50 closest assemblies and found that b12.3 is enriched with BGCs. The genomic analysis also revealed that genomic architecture corresponds to the experimentally observed activity spectrum implying that the combination of produced secondary metabolites delineates the range of inhibited phytopathogens Therefore, this study deepens our knowledge of the biology and ecology of B. mycoides residing in the Lake Baikal region. Full article

The emergence of multidrug-resistant fungi Candida auris is a worldwide health crisis connected with high rates of mortality. There is a critical need to find novel and unique antifungal compounds for treating infections of multidrug-resistant fungi such as C. auris. This study aimed to illustrate that biosynthetic gene clusters in native bacterial isolates are able to produce antifungal compounds against the multidrug-resistant fungus C. auris. It was successfully achieved using large-scale antifungal activity screening, cytotoxicity analysis, and whole genome sequencing integrated with genome mining-guided analysis and liquid chromatography–mass spectrometry (LC/MS). A list of possible gene candidates was initially identified with genome mining methods to predict secondary metabolite gene clusters of antifungal-compound-producing bacteria. Then, gene clusters present in the antifungal-compound-producing bacteria were identified and aligned with the reference genome using comparative genomic approaches. Bacillus halotolerans AQ11M9 was identified through large-scale antifungal activity screening as a natural compound-producer against multidrug-resistant C. auris, while it was nontoxic to normal human skin fibroblast cells (confirmed using a cell viability assay). The genome (4,197,347 bp) of B. halotolerans AQ11M9 with 2931 predicted genes was first mined for detecting and characterizing biosynthetic gene clusters, which revealed 10 candidate regions with antifungal activity. Clusters of AQ11M9 encoded non-ribosomal peptide synthase (NRPS) (bacilysin, bacillibactin, paenibactin, surfactin, plipastin, and fengycin) and polyketide (macrobrevin). The presence of gene clusters with anti-C. auris activity, and surfactin identified through LC/MS, from AQ11M9 suggests the potential of utilizing it as a source for a novel and powerful anti-C. auris compound. Full article

Members of the Bacillus cereus group are spore-forming Gram-positive bacilli that are commonly associated with diarrheal or emetic food poisoning. They are widespread in nature and frequently present in both raw and processed food products. Here, we genetically characterized 24 B. cereus group isolates from foodstuffs. Whole-genome sequencing (WGS) revealed that most of the isolates were closely related to B. cereus sensu stricto (12 isolates), followed by B. pacificus (5 isolates), B. paranthracis (5 isolates), B. tropicus (1 isolate), and “B. bingmayongensis” (1 isolate). The most detected virulence genes were BAS_RS06430, followed by bacillibactin biosynthesis genes (dhbA, dhbB, dhbC, dhbE, and dhbF), genes encoding the three-component non-hemolytic enterotoxin (nheA, nheB, and nheC), a gene encoding an iron-regulated leucine-rich surface protein (ilsA), and a gene encoding a metalloprotease (inhA). Various biofilm-associated genes were found, with high prevalences of tasA and sipW genes (matrix protein-encoding genes); purA, purC, and purL genes (eDNA synthesis genes); lytR and ugd genes (matrix polysaccharide synthesis genes); and abrB, codY, nprR, plcR, sinR, and spo0A genes (biofilm transcription regulator genes). Genes related to fosfomycin and beta-lactam resistance were identified in most of the isolates. We therefore demonstrated that WGS analysis represents a useful tool for rapidly identifying and characterizing B. cereus group strains. Determining the genetic epidemiology, the presence of virulence and antimicrobial resistance genes, and the pathogenic potential of each strain is crucial for improving the risk assessment of foodborne B. cereus group strains. Full article