A multitissue transcriptomic analysis reveals a potential mechanism whereby Brevibacillus laterosporus S62-9 promotes broiler growth.

Brevibacillus laterosporus S62-9 has been shown to improve broiler growth performance and immunity. In the present study, we aimed to evaluate the effects of B. laterosporus S62-9 on the immunity and lipid metabolism of broilers by means of transcriptomic analysis. A total of 160 1-day-old broilers were randomly allocated to a S62-9 group, the diet of which was supplemented with 106 CFU/g B. laterosporus S62-9 daily, and a control group, which was not. After 42 d of feeding, the broilers in the S62-9 group had higher body mass (7.2%) and feed conversion ratio (5.19%) than the control group. Supplementation with B. laterosporus S62-9 resulted in lower serum total cholesterol and low-density lipoprotein-cholesterol concentrations and higher high-density lipoprotein-cholesterol concentrations. An analysis of the fatty acid composition of the broiler's thigh muscles revealed that the proportions of the unsaturated fatty acids myristoleic acid (C14:1) and arachidonic acid (C20:1) were higher for birds in the S62-9 group. Transcriptomic analysis also showed an upregulation of immunity-related genes in the S62-9 group. Gene Ontology functional enrichment analysis showed that the mitogen-activated protein kinase pathway was enriched in the liver, the defense response was enriched in the duodenum, and immunoglobulin-related entries were enriched in the jejunum of the S62-9 group. Furthermore, the expression of key genes involved in unsaturated fatty acid synthesis (SCD, encoding stearoyl-CoA desaturase) and fatty acid metabolism (HACD2, encoding 3-hydroxyacyl-CoA dehydratase 2) was upregulated in the liver, and the expression of genes associated with fat biosynthesis and accumulation, such as PLIN1, encoding perilipin 1, and FABP4, encoding fatty acid binding protein 4, was upregulated in the ileum of the birds in the S62-9 group. In summary, supplementation with B. laterosporus S62-9 could improve immune defense and the fatty acid metabolism of broiler chickens, thereby enhancing their disease resistance and promoting growth and development.

Biochemical characterisation and production kinetics of high molecular-weight (HMW) putative antibacterial proteins of insect pathogenic Brevibacillus laterosporus isolates.

BACKGROUND: Bacterial genomes often encode structures similar to phage capsids (encapsulins) and phage tails which can be induced spontaneously or using genotoxic compounds such as mitomycin C. These high molecular-weight (HMW) putative antibacterial proteins (ABPs) are used against the competitive strains under natural environment. Previously, it was unknown whether these HMW putative ABPs originating from the insect pathogenic Gram-positive, spore-forming bacterium Brevibacillus laterosporus (Bl) isolates (1821L, 1951) are spontaneously induced during the growth and pose a detrimental effect on their own survival. Furthermore, no prior work has been undertaken to determine their biochemical characteristics. RESULTS: Using a soft agar overlay method with polyethylene glycol precipitation, a narrow spectrum of bioactivity was found from the precipitated lysate of Bl 1951. Electron micrographs of mitomycin C- induced filtrates showed structures similar to phage capsids and contractile tails. Bioactivity assays of cell free supernatants (CFS) extracted during the growth of Bl 1821L and Bl 1951 suggested spontaneous induction of these HMW putative ABPs with an autocidal activity. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis of spontaneously induced putative ABPs showed appearance of ~ 30 kDa and ~ 48 kDa bands of varying intensity across all the time intervals during the bacterial growth except in the initial hours. Statistically, spontaneously induced HMW putative ABPs of Bl 1951 exhibited a significant decrease in the number of viable cells of its producer strain after 18 h of growth in liquid. In addition, a significant change in pH and prominent bioactivity of the CFS of this particular time period was noted. Biochemically, the filtered supernatant derived from either Bl 1821L or Bl 1951 maintained bioactivity over a wide range of pH and temperature. CONCLUSION: This study reports the spontaneous induction of HMW putative ABPs (bacteriocins) of Bl 1821L and Bl 1951 isolates during the course of growth with potential autocidal activity which is critically important during production as a potential biopesticide. A narrow spectrum of putative antibacterial activity of Bl 1951 precipitate was found. The stability of HMW putative ABPs of Bl 1821L and Bl 1951 over a wide range of pH and temperature can be useful in expanding the potential of this useful bacterium beyond the insecticidal value.

Brevibacillus Laterosporus Osteomyelitis and Hardware Infection in a Young, Immunocompetent Patient Successfully Treated by Oral Minocycline and Ceftriaxone via a Peripherally Inserted Central Catheter for Six Weeks.

Brevibacillus laterosporus, an endospore-forming, aerobic, Gram-positive bacillus, is not only a potential biocontrol agent against plant pathogens but also a rare cause of human infection. Its pathogenicity in humans, especially in immunocompetent individuals, is still not fully understood. Skin infections caused by B. laterosporus are typically opportunistic, entering the skin through cuts, wounds, or other breaches in the skin's protective barrier, as in the case presented here. LEARNING POINTS: Skin infections caused by Brevibacillus laterosporus are generally considered opportunistic and more likely to occur in individuals with compromised immune systems or pre-existing skin conditions.Brevibacillus laterosporus infections in immunocompetent patients are exceedingly rare, and information about optimal treatment strategies is limited.

Brevibacillus laterosporus: A Probiotic with Important Applications in Crop and Animal Production.

Brevibacillus laterosporus (B. laterosporus) is widely distributed in nature and demonstrates significant potential for applications in biological control, environmental protection, agricultural production, and clinical medicine. This review provides a comprehensive overview of the applications of B. laterosporus in crop cultivation and animal feeding, as well as an examination of the antimicrobial peptides produced by B. laterosporus and their antibacterial mechanisms. B. laterosporus enhances crop cultivation by secreting hydrolases to improve nutrient absorption capabilities, synthesizing hormones to promote crop growth, and producing proteins to inhibit the reproduction of harmful organisms. B. laterosporus has been used to improve animal production by regulating the structure of the intestinal microbiota and inhibiting the growth of pathogenic bacteria through the secretion of various antimicrobial peptides. The bactericidal activity of Brevilaterins secreted by B. laterosporus is attributed to their ability to bind to lipopolysaccharide/lipid II molecules on the cell membrane, thereby altering permeability. Brevilaterins also inhibit bacterial reproduction by affecting relevant gene pathways in the cell membranes of pathogenic bacteria. These pathways include ATP synthesis, peptidoglycan biosynthesis, membrane transport, and cellular metabolism. In conclusion, B. laterosporus exhibits substantial potential as a probiotic activity in crop and animal production. However, applications of B. laterosporus in animal production could be improved, necessitating further research to elucidate the underlying probiotic mechanisms.

Evaluation of probiotic properties of a Brevibacillus laterosporus strain.

Brevibacillus laterosporus is a strain of probiotic bacteria that has been widely used in pest control, cash crop, and other production areas. However, few studies have been conducted on its use as a feed additive in animals. Therefore, the probiotic potential of B. laterosporus PBC01 was evaluated by characterizing hydrophobicity, auto-aggregation activity, bile salt and simulated gastrointestinal fluid tolerance, bienzymatic, and antibacterial activity. Antibiotic susceptibility, hemolysis assays, and supplemental feeding of mice were also performed to evaluate safety features. Our results showed that B. laterosporus PBC01 had moderate hydrophobicity, high auto-agglutination ability. Meanwhile, B. laterosporus PBC01 had good tolerance to bile salt and simulated gastrointestinal fluid. It had the ability to secrete protease, cellulase, and to inhibit various pathogens. In addition, B. laterosporus PBC01 was sensitive to many antibiotics, and did not produce hemolysin. In the safety assessment of mice, it did not cause any deaths, nor did it affect the cell components of blood, antioxidant capacity, and reproductive health. The study indicated the great probiotic characteristics and safety of B. laterosporus PBC01. This may provide a theoretical basis for the clinical application and development of probiotic-based feed additives.

Action mechanism of the potential biocontrol agent Brevibacillus laterosporus SN19-1 against Xanthomonas oryzae pv. oryzae causing rice bacterial leaf blight.

The causal agent of rice bacterial leaf blight (BLB) is Xanthomonas oryzae pv. oryzae (Xoo), which causes serious damage to rice, leading to yield reduction or even crop failure. Brevibacillus laterosporus SN19-1 is a biocontrol strain obtained by long-term screening in our laboratory, which has a good antagonistic effect on a variety of plant pathogenic bacteria. In this study, we investigated the efficacy and bacterial inhibition of B. laterosporus SN19-1 against BLB to lay the theoretical foundation and research technology for the development of SN19-1 as a biopesticide of BLB. It was found that SN19-1 has the ability to fix nitrogen, detoxify organic phosphorus, and produce cellulase, protease, and siderophores, as well as IAA. In a greenhouse pot experiment, the control efficiency of SN19-1 against BLB was as high as 90.92%. Further investigation of the inhibitory mechanism of SN19-1 on Xoo found that the biofilm formation ability of Xoo was inhibited and the pathogenicity was weakened after the action of SN19-1 sterile supernatant on Xoo. The activities of enzymes related to respiration and the energy metabolism of Xoo were significantly inhibited, while the level of intracellular reactive oxygen species was greatly increased. Scanning electron microscopy observations showed folds on the surface of Xoo. A significant increase in cell membrane permeability and outer membrane permeability and a decrease in cell membrane fluidity resulted in the extravasation of intracellular substances and cell death. The results of this study highlight the role of B. laterosporus SN19-1 against the pathogen of BLB and help elucidate the underlying molecular mechanisms.

Inoculating exogenous bacterium Brevibacillus laterosporus ZR-11 at maturity stage accelerates composting maturation by regulating physicochemical parameters and indigenous bacterial community succession.

Brevibacillus laterosporus ZR-11, a bio-control strain, was innovatively inoculated at maturity stage of composting to clarify its effect on physicochemical parameters and indigenous bacterial community structure in compost pile. Results revealed that ZR-11 inoculum rapidly increased pile temperature to 52 ºC and raised germination index (GI) value to beyond 85% on day 3, thereby achieving higher pile temperature and GI in the inoculated group than the non-inoculated group almost along maturity stage, and also decreased C/N ratio of the inoculated group to below 20 by composting end (day 8). Also, ZR-11 succeeded in colonizing compost pile along maturity stage. These suggested that ZR-11 as inoculum at maturity stage could accelerate compost maturation and have a potential to participate in bio-fertilizer production. High-throughput sequencing indicated that bacterial community structure experienced substantial succession in the inoculated and non-inoculated groups, and Firmicutes, Proteobacteria, and Actinobacteria were the dominant phyla in the two groups during maturity stage, with their abundances higher in the inoculated group. Saccharomonospora and Ammoniibacillus abundance increased on day 3 while Actinomadura abundance increased on day 6 in the inoculated group. As verified statistically, pile temperature and pH were key factors closely linked to dominant genera abundance, where Saccharomonospora and Ammoniibacillus abundance were positively correlated to pile temperature, while Actinomadura abundance was positively correlated to pile pH. Thus, it was inferred that ZR-11 inoculum could improve parameters such as temperature and pH to modify dominant genera abundance, thus regulating indigenous bacterial community succession, which might in turn promote compost maturation.

Laterosporulin25: A probiotically produced, novel defensin-like bacteriocin and its immunogenic properties.

Although multiple vaccines have been developed against infectious diseases, the rapid emergence of new pathogens develops an urgent need for novel strategies to combat infectious diseases. Antimicrobial peptides (AMPs) are excellent agents to fight against infectious diseases having unique multiple mechanisms of action against various pathogens. Apart from the direct applications, AMPs can also be developed as subunit vaccines or could be used as a highly immunogenic carrier protein with highly antigenic but non-immunogenic antigens. Here in the present study, we have identified a novel defensin-like bacteriocin, laterosporulin25 (LS25) upon genome mining of Brevibacillus laterosporus DSM25, a probiotic bacterial strain. By using immunoinformatic tools, we have studied the immunogenic and physiochemical properties of LS25. LS25 is characterized as defensin-like bacteriocin, having 51 amino acids and a molecular weight of 5862.7 Da. The modeled tertiary structure of LS25 is docked with TLR3 and TLR4-MD2 complex to confirm the facilitation of induced immune response that is further validated using molecular dynamics simulations and In-silico immune stimulations. Overall, detailed immunoinformatics analysis suggested LS25 as a potential candidate to be used as an adjuvant or carrier protein for subunit vaccine development, however, further in-vitro and in-vivo experiments are essential to validate its potential.

Linocin M18 protein from the insect pathogenic bacterium Brevibacillus laterosporus isolates.

Brevibacillus laterosporus (Bl) is a Gram-positive and spore-forming bacterium. Insect pathogenic strains have been characterised in New Zealand, and two isolates, Bl 1821L and Bl 1951, are under development for use in biopesticides. However, growth in culture is sometimes disrupted, affecting mass production. Based on previous work, it was hypothesised that Tectiviridae phages might be implicated. While investigating the cause of the disrupted growth, electron micrographs of crude lysates showed structural components of putative phages including capsid and tail-like structures. Sucrose density gradient purification yielded a putative self-killing protein of ~30 kDa. N-terminal sequencing of the ~30 kDa protein identified matches to a predicted 25 kDa hypothetical and a 31.4 kDa putative encapsulating protein homologs, with the genes encoding each protein adjacent in the genomes. BLASTp analysis of the homologs of 31.4 kDa amino acid sequences shared 98.6% amino acid identity to the Linocin M18 bacteriocin family protein of Brevibacterium sp. JNUCC-42. Bioinformatic tools including AMPA and CellPPD defined that the bactericidal potential originated from a putative encapsulating protein. Antagonistic activity of the ~30 kDa encapsulating protein of Bl 1821L and Bl 1951during growth in broth exhibited bacterial autolytic activity. LIVE/DEAD staining of Bl 1821L cells after treatment with the ~30 kDa encapsulating protein of Bl 1821L substantiated the findings by showing 58.8% cells with the compromised cell membranes as compared to 37.5% cells in the control. Furthermore, antibacterial activity of the identified proteins of Bl 1821L was validated through gene expression in a Gram-positive bacterium Bacillus subtilis WB800N. KEY POINTS: • Gene encoding the 31.4 kDa antibacterial Linocin M18 protein was identified • It defined the autocidal activity of Linocin M18 (encapsulating) protein • Identified the possible killing mechanism of the encapsulins.

Dietary Supplementation of Brevibacillus laterosporus S62-9 Improves Broiler Growth and Immunity by Regulating Cecal Microbiota and Metabolites.

Brevibacillus laterosporus has been added as a direct-fed microbiota to chicken. Yet, few studies have reported the effects of B. laterosporus on broiler growth and gut microbiota. The aim of this study was to evaluate the effects of B. laterosporus S62-9 on growth performance, immunity, cecal microbiota, and metabolites in broilers. A total of 160 1-day-old broilers were randomly divided into S62-9 and control groups, with or without 106 CFU/g B. laterosporus S62-9 supplementation, respectively. During the 42 days feeding, body weight and feed intake were recorded weekly. Serum was collected for immunoglobulin determination, and cecal contents were taken for 16S rDNA analysis and metabolome at Day 42. Results indicated that the broilers in S62-9 group showed an increase in body weight of 7.2% and 5.19% improvement in feed conversion ratio compared to the control group. The B. laterosporus S62-9 supplementation promoted the maturation of immune organs and increased the concentration of serum immunoglobulins. Furthermore, the α-diversity of cecal microbiota was improved in the S62-9 group. B. laterosporus S62-9 supplementation increased the relative abundance of beneficial bacteria including Akkermansia, Bifidobacterium, and Lactobacillus, while decreased the relative abundance of pathogens including Klebsiella and Pseudomonas. Untargeted metabolomics revealed that 53 differential metabolites between the two groups. The differential metabolites were enriched in 4 amino acid metabolic pathways, including arginine biosynthesis and glutathione metabolism. In summary, B. laterosporus S62-9 supplementation could improve the growth performance and immunity through the regulation of gut microbiota and metabolome in broilers.

Physiological properties and genomic insights into the plant growth-promoting rhizobacterium Brevibacillus laterosporus K75 isolated from maize rhizosphere.

BACKGROUND: When looking for a safer alternative to pesticides that are potentially harmful to living organisms, one of the directions worth looking at are plant growth-promoting rhizobacteria. The purpose of the research was a comprehensive characterization of Brevibacillus laterosporus K75, a strain isolated from maize rhizosphere. Many studies have proved B. laterosporus to be a biocontrol agent; however, little is known about B. laterosporus as a plant growth-promoting rhizobacterium. RESULTS: Ninety strains were screened for plant growth-promoting activities. Four strains with the best plant growth-promoting traits (Rhodococcus qingshengii K8, Bacillus subtilis subsp. stercoris K73, Brevibacillus laterosporus K75, and Brevibacillus laterosporus K89) were used to research their effect on maize growth. Under sterile conditions, B. laterosporus K75 showed the best stimulatory effect, significantly improving the weight of roots, shoots and leaves, and considerably increasing content of chlorophyll. In unsterilized soil, B. laterosporus K75 significantly improved length of roots and weight of leaves compared to the K73, K89, and untreated control. Moreover, B. laterosporus K75 significantly increased specific leaf area compared to the untreated control and to other inoculant treatments. The genome of B. laterosporus K75 was compared to the recently published B. laterosporus MG64. Genome-mining displayed differences in identified plant growth-promoting genes and biosynthetic gene clusters of secondary metabolites. The B. laterosporus K75 genome possessed additional genes involved in indole-3-acetic acid production and phosphate solubilization that could be attributed to its ability to enhance maize growth. CONCLUSION: Our study demonstrated that B. laterosporus K75 is a promising candidate for use in inoculant formulation, effectively facilitating maize growth. © 2022 Society of Chemical Industry.

Brevibacillus laterosporus BL1, a promising probiotic, prevents obesity and modulates gut microbiota in mice fed a high-fat diet.

Scope: Probiotics are a potential preventive strategy for obesity. However, with discrete efficacy and limited species of probiotics, there is a demand for novel strains with excellent anti-obesity properties. This study aimed to investigate the effects of Brevibacillus laterosporus BL1 on preventing obesity in high-fat diet (HFD)-fed mice. Methods and results: C57BL/6 male mice were randomly assigned to four groups (n = 10) and fed a control diet, HFD, HFD plus B. laterosporus BL1, and HFD plus supernatant of B. laterosporus BL1, respectively for 8 weeks. The results showed that prophylactic B. laterosporus BL1 treatment reduced body weight gain by 41.26% in comparison to the HFD group, and this difference was accompanied by a reduction in body fat mass and the weight of inguinal white adipose tissues and epididymal white adipose tissue (-33.39%, -39.07%, and -43.75%, respectively). Moreover, the B. laterosporus BL1-mediated improvements in lipid profile, insulin resistance, and chronic inflammation were associated with the regulation of gene expression related to lipid metabolism and enhancement of brown adipose tissue thermogenesis. Particularly, B. laterosporus BL1 intervention significantly improved HFD-induced gut flora dysbiosis, as evidenced by a reverse in the relative abundance of Bacillota and Bacteroidota, as well as an increase in the relative abundance of bacteria that produce short-chain fatty acids (SCFAs), which in turn increased SCFAs levels. Conclusion: Our findings found for the first time that B. laterosporus BL1 may be a promising probiotic for prevention of obesity associated with the regulation of gut microbiota.

Structural Diversity, LC-MS-MS Analysis and Potential Biological Activities of Brevibacillus laterosporus Extract.

Lake Mariout is Egypt's degraded coastal marine habitat that encompasses a variety of wastes. The biodiversity and hard environmental conditions allow the co-existence of organisms with high resistance and rich metabolism, making them potential candidates for screening and isolating novel microbial strains. A bacterial isolate (BF202) cultured from the marine sediments of Alexandria's Mariout Lake (Egypt) was tested for its antimicrobial and anticancer potential. The phylogenetic analysis of the isolated strain's 16S rDNA and gyrB revealed that BF202 belongs to Brevibacillus laterosporus (B. laterosporus). Antibiosis of B. laterosporus was confirmed against microbial pathogens including Escherichia coli, Klebsiella pneumoniae, Salmonella typhi, and Staphylococcus aureus. The highest antibacterial activity was detected on glucose peptone medium after 18 h of incubation at 35 °C, and at pH of 7.0 in the presence of mannose and ammonium carbonate as carbon and nitrogen sources, respectively. The cytotoxicity of the methanolic extract against breast cancer (MCF-7) and normal Vero cell lines, using the MTT test, revealed IC50 values of 7.93 and 23.79 µg/mL, respectively. To identify apoptotic and necrotic cells, a flow cytometric analysis using annexin V-FITC/PI dual-labeling was utilized and recorded a higher number of necrotic cells compared to apoptotic ones. Similarly, the cell cycle S-phase arrest was reported. The LC-MS-MS investigation of B. laterosporus extract and the molecular networking database analysis demonstrated five strategic diketopiperazine compounds with antimicrobial and anticancer activities. Taken together, this research shows that the crude extract of B. laterosporus might be an effective agent against drug-resistant bacteria and malignant disorders due to its richness in diketopiperazines.

Antibacterial Regularity Mining Beneath the Systematic Activity Database of Lipopeptides Brevilaterins: An Instructive Activity Handbook for Its Food Application.

Bacterial contamination is a primary threat to food safety. Therefore, the persistent development of natural antibacterial agents has become essential work. The present essay attempts to establish a systematic antibacterial activity database to instruct the food application of brevilaterins, promising antibacterial lipopeptides from Brevibacillus laterosporus S62-9. Minimum inhibitory concentrations (MIC) and minimum bactericidal concentrations (MBC) were systematically collected from 43 species of standard bacteria and 140 strains of isolated bacteria (food spoilage bacteria and antibiotic-resistant bacteria) using a broth dilution method. The results showed that brevilaterins performed a broad-spectrum inhibitory (0.5~128 µg/mL) and bactericidal activity (1~256 µg/mL), especially efficient against Gram-positive bacteria and spoilage bacteria from grain products. Moreover, brevilaterins not only inhibit and kill multiple antibiotic-resistant bacteria but do not readily develop resistance, with a small specific value of MBC/MIC (1~8). Furthermore, brevilaterins would interact with negatively charged sodium dodecyl sulfate and bind amphipathic soybean phospholipid with an affinity constant of KD = 4.70 × 10-4 M. No significant activity difference was found between brevilaterin B and brevilaterin C. Collectively, this work contributed rich antibacterial data of brevilaterins and revealed the antibacterial regularity beneath these data, which can be used as an activity handbook to instruct their application in food safety.

An accurate strategy for pointing the key biocatalytic sites of bre2691A protein for modification of the brevilaterin from Brevibacillus laterosporus.

BACKGROUND: Brevilaterin A-E, a novel class of multi-component cationic antimicrobial lipopeptides, were biosynthesized by a non-ribosomal peptides synthetase (NRPS) in Brevibacillus laterosporus. However, the antimicrobial abilities of different brevilaterin components varied greatly, and this multi-component form was impeding the scale production of the excellent component, and a little information about the brevilaterin biosynthesis mechanism was available to apply in brevilaterin design modification. In this study, we used an accurate strategy that revealed the reason for producing multi-component was the substrate selectivity of bre2691A protein being not enough specific and pinpointed the key design sites to make the specificity of bre2691A enhanced. RESULTS: Bioinformatic analysis revealed that the biocatalytic site of bre2691A, which was an adenylation domain catalyzed and recognized methionine, leucine, valine and isoleucine and thus introduced them into brevilaterins and caused different components (brevilaterin A-E), was consisted of A1 ~ A10 residues named specificity-conferring code. Coupling molecular docking simulations with mutation studies identified A2 and A7 as critical residues, where determined substrate-specificity and impacted activity. The in virto activity assay showed that the A2 mutant (G193A) would lose activity against methionine and have no effect on the other three amino acids, the A7 mutant (G285C) would enhance the catalytic activity against four substrates, especially against leucine at almost a double activity. When the A2 and A7 residues were synchronously mutated, this mutant would be more focused on recognizing leucine. CONCLUSIONS: An accurate strategy that combined with bioinformatics and site-directed mutation techniques revealed the pivotal site A2 and A7 positions of bre2691A protein that could be used to design and modify brevilaterins, thus further providing a reasonable direction of genetic engineering for Brevibacillus laterosporus. A deeper understanding of the function of crucial residues in the adenylation domain would make it get more accurate and highly efficient design and more fully utilized. Furthermore, it would contribute to biotechnological applications, namely for the large centralized synthesis of antimicrobial peptides, or for the optimization of their production.

Brevilaterin B from Brevibacillus laterosporus has selective antitumor activity and induces apoptosis in epidermal cancer.

Brevilaterins as antimicrobial peptides (AMPs) secreted by a newly discovered species Brevibacillus laterosporus, had been demonstrated to display excellent antibacterial and antifungal activities; however, very limited information about their new bioactivity was ever developed. Herein, we discovered Brevilaterin B, an AMP produced by Br. laterosporus S62-9, exhibited a new anticancer activity and investigated its anticancer details. Proliferation, membrane permeability and apoptotic rate of cell lines were studied by methods of CCK-8 Assay, LDH Assay and Annexin V-FITC/PI Kits, respectively. ROS levels and mitochondrial membrane potential of tested cells were further detected through the fluorescent probes DCFH-DA and JC-1. Brevilaterin B exhibited broad-spectrum anticancer activity in a dose-dependent manner. It selectively inhibited the proliferation of epidermal cancer cell A431 but had no effect on its control normal cells in a dose of 2.0 µg/mL. In comparision, typical morphological characteristics of apoptosis and an apoptotic ratio of 71.0% in A431 were observed after treatment by 2.0-3.0 µg/mL of Brevilaterin B. The ROS levels increased by 21.3% and mitochondrial membrane potential reduced by 48.8% from A431 were further occurred, indicating Brevilaterin B's anticancer action was mainly focus on the mitochondrion of cancer cells. In total, Brevilaterin B we reported above maybe believed to be a potential application as an anticancer medicament, increasing its commercial value.

Transcriptome Analysis Reveals the Algicidal Mechanism of Brevibacillus laterosporus against Microcystis aeruginosa through Multiple Metabolic Pathways.

It is widely accepted that eutrophication has played an important role in the formation of harmful cyanobacterial blooms in recent decades, which impacts water quality and ecological environment and causes huge economic losses. Algicidal bacteria have a promising application prospect in controlling cyanobacterial blooms in aquaculture water. Here, the process of the algicidal bacterium Brevibacillus laterosporus strain Bl-zj acting on Microcystis aeruginosa was explored using transcriptome analysis to elucidate the algicidal mechanism. The results of the co-culture of bacterium and alga showed a strong alga-lysing effect of B. laterosporus against M. aeruginosa with an extreme morphology deformation of the algal cells. A total of 2744 differentially expressed genes of B. laterosporus were identified, which were mainly involved in the metabolism of amino acid, carbohydrate, and lipid. In the co-cultured group, the expression of genes mainly enriched in valine, leucine and isoleucine degradation, and fatty acid degradation were significantly increased. However, the expression of the genes related to ribosome were mainly inhibited. Transcriptome analysis showed that B. laterosporus obtained ATP and energy by the degradation of valine, leucine, isoleucine, and fatty acids, and destroyed algal cells by efflux pump transporters, secretion of hydrolytic enzymes, antibiotics, proteases, and other secondary metabolites, resulting in algal death and achieving the algicidal effect.

Broad-spectrum cytotoxicity to cancer cells of Brevilaterin C from Brevibacillus laterosporus and its specific mechanism on human epidermal cancer cells.

Antimicrobial peptides (AMP) from Brevibacillus laterosporus have good prospects as clinical treatments for cancer. Nevertheless, details about their anticancer spectrum and mode of cytotoxicity remain poorly understood. A newly found AMP (named Brevilaterin C) secreted by B. laterosporus S62-9 exhibited strong inhibition on almost cancer cell lines examined at a concentration of 8 µg/ml but was relatively safe for normal cells. We further systematically examined its cytotoxicity and mechanism toward human epidermal cancer cell A431. A dosage of 3 µg/ml of Brevilaterin C could significantly increase lactate dehydrogenase release of tumor cells. Moreover, it could remarkably increase the ratio of apoptosis and reactive oxygen species generation of A431, indicating effective induction of apoptosis. Moreover, the formation of JC-1 aggregates was effectively prevented by a low concentration of Brevilaterin C, indicating its effective induction of A431's apoptosis. Brevilaterin C exhibited broad-spectrum cytotoxicity to cancer cells, indicating a good potential prospect in the medical field.

Structural Organization of Brevilaterin Biosynthesis in Brevibacillus laterosporus S62-9: A Novel MbtH-Independent Cationic Antimicrobial Peptide Synthetase System.

Cationic antimicrobial peptides, produced by nonribosomal peptide synthetases (NRPSs), have received great attention in different applications, including as biocontrol and antimicrobial agents against foodborne pathogenic bacteria. Also, Brevibacillus spp. is a competent microorganism to produce cationic antimicrobial peptides yet has received little attention. Herein, Brevibacillus laterosporus S62-9 genome mining revealed an integrated cationic antimicrobial peptide synthetase system that synthesized brevilaterin. Combining biochemical analysis with bioinformatics elucidated that the A domain from this system was the MbtH-independent enzyme and showed activity against the same amino acid in the structure of brevilaterin. Moreover, the creations of the first three and position 12 residues in the sequence were targeted to bre261, bre270, bre2691A, and bre2662, respectively. Further analysis of the specificity-conferring code of the A domain suggested that a tiny difference would make the activity of the A domain very diverse and the range of substrate selection would be enlarged or narrowed by changing some residues in the code. The dissection of this biosynthesis mechanism would contribute to the successful realization of reasonable artificial design and the modification of bioactive peptides, and this capable organism also would be more fully utilized.

Ultrastructural and pathogenicity of Brevibacillus laterosporus against sinantropic muscoid dipterans.

Brevibacillus laterosporus has entomopathogenic potential against several orders of insects and its wide bioactivity is associated with a variety of strain-specific molecules. In order to avoid the use of synthetic insecticides, along with the need to control insect pests, microbial control has been widely used. Muscoid dipterans are known for their medical-veterinary and sanitary importance, and synanthropy. The enormous biotechnological potential of B. laterosporus has been demonstrated, but there are still few studies with muscoid dipterans. The aim of the study was to verify the mortality of B. laterosporus NRS590 on synanthropic flies and to characterize its different cell stages ultrastructurally. The flies were collected from garbage bins and the colonies were adapted to the laboratory conditions. Bioassays with neo larvae were carried out from the bacterial growth in the phases: vegetative (6 hr), sporangium (20 hr), and free spores (44 hr). An aliquot of each phase was collected for Transmission (TEM) and Scanning Electron Microscopy (SEM). The effectiveness of NRS590 was observed in the sporulation phase, where the corrected mortality was 83.3, 85.1, and 99% for Chrysomya megacephala, Chrysomya putoria, and Musca domestica, respectively. The parasporal body was observed in detail on the entire spore surface. Although our knowledge of this bacterium is growing, it remains to be determined the real virulence factors responsible for the wide entomopathogenic activity observed on muscoid dipterans. Therefore, this study can provide subsidies for the improvement of efficient and safe microbial control techniques for the environment and living beings.