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1.
Molecules ; 26(15)2021 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-34361631

RESUMO

The cyclic lipopeptide produced from Bacillus pumilus strain HY1 was isolated from Korean soybean sauce cheonggukjang. The chemical structures of the surfactin isomers were analyzed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and electrospray ionization tandem mass spectrometry (ESI-MS/MS). The five potential surfactin isoforms were detected with protonated masses of m/z 994.7, 1008.7, 1022.7, 1036.7, and 1050.7 and different structures in combination with Na+, K+, and Ca2+ ions. ESI-MS/MS analysis revealed that the isolated surfactin possessed the precise amino acid sequence LLVDLL and hydroxyl fatty acids with 12 to 16 carbons. The surfactin content during cheonggukjang fermentation increased from 0.3 to 51.2 mg/kg over 60 h of fermentation. The mixture of five surfactin isoforms of cheonggukjang inhibited the growth of two cancer cell lines. The growth of both MCF-7 and Caco-2 cells was strongly inhibited with 100 µg/µL of surfactin. This study is the first-time report of five surfactin isomers of Bacillus pumilus strain HY1 during Korean soybean sauce cheonggukjang fermentation, which has cytotoxic properties.


Assuntos
Bacillus pumilus/metabolismo , Proliferação de Células/efeitos dos fármacos , Alimentos e Bebidas Fermentados/microbiologia , Lipopeptídeos , Soja/microbiologia , Células CACO-2 , Humanos , Lipopeptídeos/química , Lipopeptídeos/isolamento & purificação , Lipopeptídeos/farmacologia , Células MCF-7
2.
Braz J Biol ; 83: e243874, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34378658

RESUMO

In recent days, cheapest alternative carbon source for fermentation purpose is desirable to minimize production cost. Xylanases have become attractive enzymes as their potential in bio-bleaching of pulp and paper industry. The objective of the present study was to identify the potential ability on the xylanase production by locally isolated Bacillus pumilus BS131 by using waste fiber sludge and wheat bran media under submerged fermentation. Culture growth conditions were optimized to obtain significant amount of xylanase. Maximum xylanase production was recorded after 72 hours of incubation at 30 °C and 7 pH with 4.0% substrate concentration. In the nutshell, the production of xylanase using inexpensive waste fiber sludge and wheat-bran as an alternative in place of expensive xylan substrate was more cost effective and environment friendly.


Assuntos
Bacillus pumilus , Bacillus , Bacillus/metabolismo , Bacillus pumilus/metabolismo , Fibras na Dieta , Endo-1,4-beta-Xilanases/metabolismo , Fermentação , Concentração de Íons de Hidrogênio , Esgotos , Temperatura
3.
Sci Rep ; 11(1): 17196, 2021 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-34433897

RESUMO

Heavy metals contaminate the soil that alters the properties of soil and negatively affect plants growth. Using microorganism and plant can remove these pollutants from soil. The present investigation was designed to evaluate the induced effect of Bacillus pumilus on maize plant in Cadmium (Cd) contaminated soil. Three different concentrations of Cd (i.e. 0.25, 0.50 and 0.75 mg kg-1) were applied in soil under which maize plants were grown. The germination percentage, shoot length, leaf length, number of leaves, root length, fresh weight and nutrient uptake by maize plant were determined. The experiment was conducted by using complete randomized design (CRD) with three replicates. The result indicated that germination percentage, Shoot length, leaf length, root length, number of leaves, and plant fresh weight were reduced by 37, 39, 39, 32 and 59% respectively at 0.75 mg kg-1 of CdSO4 concentration but when maize seeds inoculated with Bacillus pumilus significantly increased the germination percentage, shoot length, leaf length, number of leaves, plant fresh weight at different concentrations of CdSO4. Moreover, the plant protein were significantly increased by 60% in T6 (0.25 mg kg-1 of CdSO4 + inoculated seed) and Peroxidase dismutase (POD) was also significantly higher by 346% in T6 (0.25 mg kg-1 of CdSO4 + inoculated seed), however, the Superoxide dismutase (SOD) was significantly higher in T5 (0.75 mg kg-1 of CdSO4 + uninoculated seed) and was 769% higher as compared to control. The Cd contents in Bacillus pumilus inoculated maize roots and shoots were decreased. The present investigations indicated that the inoculation of maize plant with Bacillus pumilus can help maize plants to withstand Cd stress but higher concentration of Cd can harm the plant. The Bacillus pumilus has good potential to remediate Cd from soil, and also have potential to reduce the phyto availability and toxicity of Cd.


Assuntos
Bacillus pumilus/metabolismo , Cádmio/toxicidade , Poluentes do Solo/toxicidade , Zea mays/efeitos dos fármacos , Bacillus pumilus/patogenicidade , Biodegradação Ambiental , Cádmio/metabolismo , Sementes/efeitos dos fármacos , Sementes/metabolismo , Sementes/microbiologia , Poluentes do Solo/metabolismo , Estresse Fisiológico , Zea mays/metabolismo , Zea mays/microbiologia
4.
Toxins (Basel) ; 13(5)2021 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-33919181

RESUMO

Zearalenone (ZEA) is a mycotoxin widely occurring in many agricultural commodities. In this study, a purified bacterial isolate, Bacillus sp. S62-W, obtained from one of 104 corn silage samples from various silos located in the United States, exhibited activity to transform the mycotoxin ZEA. A novel microbial transformation product, ZEA-14-phosphate, was detected, purified, and identified by HPLC, LC-MS, and NMR analyses. The isolate has been identified as belonging to the genus Bacillus according to phylogenetic analysis of the 16S rRNA gene and whole genome alignments. The isolate showed high efficacy in transforming ZEA to ZEA-14-phosphate (100% transformation within 24 h) and possessed advantages of acid tolerance (work at pH = 4.0), working under a broad range of temperatures (22-42 °C), and a capability of transforming ZEA at high concentrations (up to 200 µg/mL). In addition, 23 Bacillus strains of various species were tested for their ZEA phosphorylation activity. Thirteen of the Bacillus strains showed phosphorylation functionality at an efficacy of between 20.3% and 99.4% after 24 h incubation, suggesting the metabolism pathway is widely conserved in Bacillus spp. This study established a new transformation system for potential application of controlling ZEA although the metabolism and toxicity of ZEA-14-phosphate requires further investigation.


Assuntos
Bacillus/metabolismo , Zearalenona/metabolismo , Bacillus/genética , Bacillus pumilus/genética , Bacillus pumilus/metabolismo , Cromatografia Líquida de Alta Pressão , Cromatografia Gasosa-Espectrometria de Massas , Fosforilação , Filogenia , RNA Ribossômico 16S/genética , Alinhamento de Sequência
5.
Microb Cell Fact ; 20(1): 40, 2021 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-33557838

RESUMO

BACKGROUND: Microbial surfactants called biosurfactants, thanks to their high biodegradability, low toxicity and stability can be used not only in bioremediation and oil processing, but also in the food and cosmetic industries, and even in medicine. However, the high production costs of microbial surfactants and low efficiency limit their large-scale production. This requires optimization of management conditions, including the possibility of using waste as a carbon source, such as food processing by-products. This papers describes the production and characterization of the biosurfactant obtained from the endophytic bacterial strain Bacillus pumilus 2A grown on various by-products of food processing and its potential applications in supporting plant growth. Four different carbon and nitrogen sources, pH, inoculum concentration and temperature were optimized within Taguchi method. RESULTS: Optimization of bioprocess within Taguchi method and experimental analysis revealed that the optimal conditions for biosurfactant production were brewer's spent grain (5% w/v), ammonium nitrate (1% w/v), pH of 6, 5% of inoculum, and temperature at 30 °C, leading to 6.8 g/L of biosurfactant. Based on gas chromatography-mass spectrometry and Fourier transform infrared spectroscopy analysis produced biosurfactant was determined as glycolipid. Obtained biosurfactant has shown high and long term thermostability, surface tension of 47.7 mN/m, oil displacement of 8 cm and the emulsion index of 69.11%. The examined glycolipid, used in a concentration of 0.2% significantly enhanced growth of Phaseolus vulgaris L. (bean), Raphanus L. (radish), Beta vulgaris L. (beetroot). CONCLUSIONS: The endophytic Bacillus pumilus 2A produce glycolipid biosurfactant with high and long tem thermostability, what makes it useful for many purposes including food processing. The use of brewer's spent grain as the sole carbon source makes the production of biosurfactants profitable, and from an environmental point of view, it is an environmentally friendly way to remove food processing by products. Glycolipid produced by endophytic Bacillus pumilus 2A significantly improve growth of Phaseolus vulgaris L. (bean), Raphanus L. (radish), Beta vulgaris L. (beetroot). Obtained results provide new insight to the possible use of glycolipids as plant growth promoting agents.


Assuntos
Bacillus pumilus , Beta vulgaris/crescimento & desenvolvimento , Endófitos , Phaseolus/crescimento & desenvolvimento , Raphanus/crescimento & desenvolvimento , Tensoativos , Bacillus pumilus/química , Bacillus pumilus/metabolismo , Endófitos/química , Endófitos/metabolismo , Tensoativos/isolamento & purificação , Tensoativos/metabolismo , Tensoativos/farmacologia
6.
World J Microbiol Biotechnol ; 36(12): 181, 2020 Nov 09.
Artigo em Inglês | MEDLINE | ID: mdl-33164140

RESUMO

Although the use of degrading-bacteria is one of the most efficient methods for the bioremediation of polluted sites, detection, selection and proliferation of the most efficient and competing bacteria is still a challenge. The objective of this multi-stage research was to investigate the effects of the selected bacterial strains on the degradation of anthracene, florentine, naphthalene, and oil, determined by biochemical tests. In the first stage, using the following tests: (a) biosurfactant production (emulsification, oil spreading, number of drops, drop collapse, and surface tension), (b) biofilm production, (c) activity of laccase enzyme, and (d) exopolysaccaride production, the three bacterial strains with the highest degrading potential including Bacillus pumilus, B. aerophilus, and Marinobacter hydrocarbonoclasticus were chosen. In the second stage using the following tests: (a) bacterial growth, (b) laccase enzyme activity, and (c) biosurfactant production (emulsification, oil spreading, and collapse of droplet) the degrading ability of the three selected bacterial strains plus Escherichia coli were compared. Different bacterial strains were able to degrade anthracene, florentine, naphthalene, and oil by the highest rate, three days after inoculation (DAI). However, M. hydrocarbonoclasticus showed the highest rate of florentine degradation. Although with increasing pollutant concentration the degrading potential of the bacterial strains significantly decreased, M. hydrocarbonoclasticus was determined as the most efficient bacterial strain.


Assuntos
Bactérias/crescimento & desenvolvimento , Bactérias/metabolismo , Poluentes Ambientais/química , Hidrocarbonetos Policíclicos Aromáticos/química , Antracenos/química , Bacillus/crescimento & desenvolvimento , Bacillus/metabolismo , Bacillus pumilus/crescimento & desenvolvimento , Bacillus pumilus/metabolismo , Bactérias/isolamento & purificação , Biodegradação Ambiental , Biofilmes , Biocombustíveis/análise , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/metabolismo , Lacase/metabolismo , Marinobacter/crescimento & desenvolvimento , Marinobacter/metabolismo , Naftalenos/química , Polissacarídeos Bacterianos/metabolismo , Tensoativos/metabolismo
7.
Int J Mol Sci ; 21(21)2020 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-33167434

RESUMO

Unpredictable influenza pandemics, annual epidemics, and sporadic poultry-to-human avian influenza virus infections with high morbidity and mortality rates dictate a need to develop new antiviral approaches. Targeting cellular pathways and processes is a promising antiviral strategy shown to be effective regardless of viral subtypes or viral evolution of drug-resistant variants. Proteomics-based searches provide a tool to reveal the druggable stages of the virus life cycle and to understand the putative antiviral mode of action of the drug(s). Ribonucleases (RNases) of different origins not only demonstrate antiviral effects that are mediated by the direct RNase action on viral and cellular RNAs but can also exert their impact by signal transduction modulation. To our knowledge, studies of the RNase-affected cell proteome have not yet been performed. To reveal cellular targets and explain the mechanisms underlying the antiviral effect employed by the small extra-cellular ribonuclease of Bacillus pumilus (binase) both in vitro and in vivo, qualitative shotgun and quantitative targeted proteomic analyses of the influenza A virus (IAV) H1N1pdm09-infected A549 cells upon binase treatment were performed. We compared proteomes of mock-treated, binase-treated, virus-infected, and virus-infected binase-treated cells to determine the proteins affected by IAV and/or binase. In general, IAV demonstrated a downregulating strategy towards cellular proteins, while binase had an upregulating effect. With the help of bioinformatics approaches, coregulated cellular protein sets were defined and assigned to their biological function; a possible interconnection with the progression of viral infection was conferred. Most of the proteins downregulated by IAV (e.g., AKR1B1, AKR1C1, CCL5, PFN1, RAN, S100A4, etc.) belong to the processes of cellular metabolism, response to stimulus, biological regulation, and cellular localization. Upregulated proteins upon the binase treatment (e.g., AKR1B10, CAP1, HNRNPA2B1, PFN1, PPIA, YWHAB, etc.) are united by the processes of biological regulation, cellular localization, and immune and metabolic processes. The antiviral activity of binase against IAV was expressed by the inversion of virus-induced proteomic changes, resulting in the inhibition of virus-associated processes, including nuclear ribonucleoprotein export (NCL, NPM1, Nup205, and Bax proteins involved) and cytoskeleton remodeling (RDX, PFN1, and TUBB) induced by IAV at the middle stage of single-cycle infection in A549 cells. Modulation of the immune response could be involved as well. Overall, it seems possible that binase exerts its antiviral effects in multiple ways.


Assuntos
Endorribonucleases/farmacologia , Vírus da Influenza A/efeitos dos fármacos , Células A549 , Animais , Antivirais/metabolismo , Antivirais/farmacologia , Bacillus pumilus/enzimologia , Bacillus pumilus/metabolismo , Linhagem Celular , Endorribonucleases/metabolismo , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Humanos , Vírus da Influenza A/genética , Vírus da Influenza A/patogenicidade , Influenza Humana/virologia , Proteoma , Proteômica/métodos , Ribonucleases/metabolismo , Replicação Viral/efeitos dos fármacos
8.
Int J Biol Macromol ; 164: 3340-3348, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-32871119

RESUMO

The bioactive form of thermostable and alkali stable pectinase of Bacillus pumilus dcsr1 is a homodimer of the molecular mass of 60 kDa with a pI of 4.6. The enzyme is optimally active at 50 °C and pH 10.5, and its Michaelis constant (Km), maximum rate of reaction (Vmax), activation energy (Ea), and temperature quotient (Q10) values (for citrus pectin) are 0.29 mg mL-1, 116 µmole mg-1 min-1, 74.73 KJmol-1 and 1.57, respectively. The enzyme has a shelf life of one and a half years at room temperature as well as 4 °C. The activity of the enzyme is stimulated by Mn2+ and Ca2+ and inhibited by Hg+, Cd2+, Co2+, Zn2+, Fe2+, Pb2+, EDTA and urea to a varied extent. The conformational studies of the enzyme revealed a high ß-sheet content in the bioactive dimer, and high α-helix in the inactive monomer. The Circular Dichroism (CD) spectra of the dimer in the presence of inhibitors suggested a marked decrease in ß-sheet, and a significant increase in α-helix, suggesting a key role of ß-sheets in the enzyme catalysis. Based on the end product analysis, the enzyme is an exopolygalacturonase with a unique ability of transglycosylation. When ramie fibers were treated with the enzyme, removal of gummy material (pectin) was visible, confirming its applicability in the degumming process.


Assuntos
Bacillus pumilus/enzimologia , Glicosídeo Hidrolases/química , Glicosídeo Hidrolases/isolamento & purificação , Bacillus/enzimologia , Bacillus pumilus/metabolismo , Proteínas de Bactérias/química , Boehmeria/química , Boehmeria/metabolismo , Cisteína Endopeptidases/química , Cisteína Endopeptidases/isolamento & purificação , Estabilidade Enzimática , Concentração de Íons de Hidrogênio , Cinética , Peso Molecular , Pectinas/química , Poligalacturonase/química , Polissacarídeo-Liases/química , Especificidade por Substrato , Temperatura
9.
Chemosphere ; 260: 127559, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32673872

RESUMO

Bacillus pumilus SWU7-1 was isolated from strontium ion (Sr(II))-uncontaminated soil, its biosorption potential was evaluated, and the effect of γ-ray radiation treatment on its biosorption was discussed. Domesticated under Sr(II) stress promoted the biosorption ability of B. pumilus to Sr(II), and the biosorption efficiency increased from 46.09% to 94.69%. At a lower initial concentration, the living bacteria had the ability to resist the biosorption of Sr(II). The optimal initial concentration range was 54-130 mg/L. The biosorption profile was better matched by Langmuir than Freundlich model, showing that the biosorption process of Sr(II) by the experimental strain was closer to the surface adsorption. According to Langmuir model, the maximum biosorption capacity of B. pumilus on Sr (II) was 299.4 mg/g. During the bacterial growth in the biosorption process, the changes in biosorption capacity and efficiency can be divided into two phases, and a pseudo-second-order model is followed in each phase. There was no significant difference in the biosorption efficiency of bacteria with different culture time after γ-ray radiation, and all of them were above 90%, which showed that B. pumilus had significant radiation resistance under experimental conditions. This study emphasized the potential application of B. pumilus in the treatment of radioactive Sr(II) pollution by biosorption.


Assuntos
Adsorção , Bacillus pumilus/metabolismo , Biodegradação Ambiental , Isótopos de Estrôncio/metabolismo , Cinética , Microbiologia do Solo , Poluentes Radioativos do Solo , Poluentes Químicos da Água/metabolismo
10.
Sci Rep ; 10(1): 10033, 2020 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-32572051

RESUMO

Huge quantities of keratinaceous waste are a substantial and almost totally unexploited protein resource which could be upgraded for use as high value-added products by efficient keratinolytic enzymes. In this study, we found that Bacillus sp. 8A6 can efficiently degrade chicken feather after 24 h growth. According to phylogenetic analysis, the strain (formerly identified as Bacillus pumilus 8A6) belongs to the B. pumilus species clade but it is more closely related to B. safensis. Hotpep predicted 233 putative proteases from Bacillus sp. 8A6 genome. Proteomic analysis of culture broths from Bacillus sp. 8A6 cultured on chicken feathers or on a mixture of bristles and hooves showed high abundance of proteins with functions related to peptidase activity. Five proteases (one from family M12, one from family S01A, two from family S08A and one from family T3) and four oligopeptide and dipeptide binding proteins were highly expressed when Bacillus sp. 8A6 was grown in keratin media compared to LB medium. This study is the first to report that bacterial proteases in families M12, S01A and T3 are involved in keratin degradation together with proteases from family S08.


Assuntos
Bacillus/enzimologia , Queratinas/metabolismo , Peptídeo Hidrolases/metabolismo , Animais , Bacillus/genética , Bacillus/metabolismo , Bacillus pumilus/enzimologia , Bacillus pumilus/genética , Bacillus pumilus/metabolismo , Galinhas , Cisteína Proteases/genética , Cisteína Proteases/metabolismo , Plumas/metabolismo , Metaloproteases/genética , Metaloproteases/metabolismo , Peptídeo Hidrolases/genética , Filogenia , Proteômica , Serina Proteases/genética , Serina Proteases/metabolismo
11.
Int J Biol Macromol ; 161: 525-530, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32531366

RESUMO

A novel feruloyl esterase (BpFae12) with rosmarinic acid (RA) hydrolysis activity was isolated from Bacillus pumilus W3 and expressed in Escherichia coli BL21 (DE3). With RA as a substrate, the optimal pH and temperature of BpFae12 were pH 8.0 and 50 °C, respectively. The specific enzyme activity was 12.8 U·mg-1. BpFae12 showed the highest activity and substrate affinity toward RA (Vmax of 13.13 U·mg-1, Km of 0.41 mM). Moreover, it also presented strong hydrolysis performance against chlorogenic acid (190.17 U·mg-1). RA was effectively Hydrolyzed into more bioactive caffeic acid and 3,4-dihydroxyphenyllactic acid by BpFae12, which have potential applications in the food industry.


Assuntos
Bacillus pumilus/química , Hidrolases de Éster Carboxílico/química , Cinamatos/química , Depsídeos/química , Hidrólise/efeitos dos fármacos , Bacillus pumilus/metabolismo , Ácidos Cafeicos/química , Ácido Clorogênico/química , Escherichia coli/metabolismo , Concentração de Íons de Hidrogênio , Peso Molecular , Especificidade por Substrato , Temperatura
12.
Mol Biol Rep ; 47(5): 3347-3359, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32248385

RESUMO

Cyclic dipeptides are increasingly gaining importance as considering its significant biological and pharmacological activities. This study was aimed to investigate the anticancer activity of a dipeptide Cyclo(-Pro-Tyr) (DP) identified from marine sponge Callyspongia fistularis symbiont Bacillus pumilus AMK1 and the underlying apoptotic mechanisms in the liver cancer HepG2 cell lines. MTT assay was done to demonstrate the cytotoxic effect of DP in HepG2 cells and mouse Fibroblast McCoy cells. Initially, apoptosis inducing activity of DP was identified using propidium iodide (PI) and acridine orange/ethidium bromide (AO/EB) dual staining, then it was confirmed by DNA fragmentation assay and western blotting analysis of apoptosis related markers Bax, Bcl-2, cytochrome c, caspase-3 and cleaved poly (ADP-ribose) polymerase (PARP). Rhodamine 123 staining was performed to observe DP effects on the mitochondrial membrane potential (MMP) and DCFH-DA (Dichloro-dihydro-fluorescein diacetate) staining was done to measure the intracellular reactive oxygen species (ROS) levels. The MTT results revealed that DP initiated dose-dependent cytotoxicity in HepG2 cells, but no significant toxicity in mouse Fibroblast McCoy cells treated with DP at the specified concentrations. DP induced apoptosis, which is confirmed by the appearance of apoptotic bodies with PI and AO/EB dual staining, and DNA fragmentation. DP significantly elevated the Bax/Bcl-2 ratio, disrupted the mitochondrial membrane potential (MMP), enhanced cytochrome c release from mitochondria, increased caspase-3 activation, the cleavage of PARP and increased intracellular reactive oxygen species (ROS) levels. Besides this, DP successfully inhibited the phosphorylation of PI3K, AKT and increased PTEN expression. These results suggested DP might have anti-cancer effect by initiating apoptosis through mitochondrial dysfunction and downregulating PI3K/Akt signaling pathway in HepG2 cells with no toxicity effect on normal fibroblast cells. Therefore, DP may be developed as a potential alternative therapeutic agent for treating hepatocellular carcinoma.


Assuntos
Apoptose/fisiologia , Carcinoma Hepatocelular/metabolismo , Dipeptídeos/farmacologia , Animais , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Bacillus pumilus/enzimologia , Bacillus pumilus/metabolismo , Callyspongia/microbiologia , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/metabolismo , Dipeptídeos/metabolismo , Células Hep G2/metabolismo , Humanos , Fígado/patologia , Neoplasias Hepáticas/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos
13.
Appl Microbiol Biotechnol ; 104(8): 3445-3457, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32088759

RESUMO

Single-molecule real-time (SMRT) sequencing can be used to identify a wide variety of chemical modifications of the genome, such as methylation. Here, we applied this approach to identify N6-methyl-adenine (m6A) and N4-methyl-cytosine (m4C) modification in the genome of Bacillus pumilus BA06. A typical methylation recognition motif of the type I restriction-modification system (R-M), 5'-TCm6AN8TTGG-3'/3'-AGTN8m6AACC-5', was identified. We confirmed that this motif was a new type I methylation site using REBASE analysis and that it was recognized by a type I R-M system, Bpu6ORFCP, according to methylation sensitivity assays in vivo and vitro. Furthermore, we found that deletion of the R-M system Bpu6ORFCP induced transcriptional changes in many genes and led to increased gene expression in pathways related to ABC transporters, sulfur metabolism, ribosomes, cysteine and methionine metabolism and starch and sucrose metabolism, suggesting that the R-M system in B. pumilus BA06 has other significant biological functions beyond protecting the B. pumilus BA06 genome from foreign DNA.


Assuntos
Bacillus pumilus/genética , Metilação de DNA , Regulação Bacteriana da Expressão Gênica , Genoma Bacteriano , Motivos de Nucleotídeos , Bacillus pumilus/metabolismo , DNA Bacteriano/genética , Redes e Vias Metabólicas/genética
14.
Arch Microbiol ; 202(2): 399-409, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31686120

RESUMO

In addition to rhizobia, other non-symbiotic endophytic bacteria also have been simultaneously isolated from the same root nodules. The existence of non-symbiotic endophytic bacteria in leguminous root nodules is a universal phenomenon. The vast majority of studies have detected endophytic bacteria in other plant tissues. In contrast, little systemic observation has been made on the non-symbiotic endophytic bacteria within leguminous root nodules. The present investigation was carried out to isolate plant growth-promoting endophytic non-symbiotic bacteria from indigenous leguminous Sphaerophysa salsula and their influence on plant growth. A total of 65 endophytic root nodule-associated bacteria were isolated from indigenous legume S. salsula growing in the northwestern arid regions of China. When combining our previous work with the current study, sequence analysis of the nifH gene revealed that the strain belonging to non-nodulating Bacillus pumilus Qtx-10 had genes similar to those of Rhizobium leguminosarum Qtx-10-1. The results indicated that horizontal gene transfer could have occurred between rhizobia and non-symbiotic endophyties. Under pot culture conditions, out of the 20 representative endophytic isolates, 15 with plant growth-promoting traits, such as IAA production, ACC deaminase, phosphate solubilization, chitinase, siderophore, and fungal inhibition activity showed plant growth-promoting activity with respect to various plant parameters such as chlorophyll content, fresh weight of plant, shoot length, nodule number per plant and average nodule weight per plant when co-inoculated with rhizobial bioinoculant Mesorhizobium sp. Zw-19 under N-free culture conditions. Among them, Bacillus pumilus Qtx-10 and Streptomyces bottropensis Gt-10 were excellent plant growth-promoting bacteria, which enhanced the seeding fresh weight by 87.5% and the shoot length by 89.4%, respectively. The number of nodules grew more than 31.89% under field conditions. Our findings indicate the frequent presence of these non-symbiotic endophytic bacteria within root nodules, and that they help to improve nodulation and nitrogen fixation in legume plants through synergistic interactions with rhizobia.


Assuntos
Bacillus pumilus/metabolismo , Fabaceae/crescimento & desenvolvimento , Fabaceae/microbiologia , Mesorhizobium/metabolismo , Nódulos Radiculares de Plantas/microbiologia , Streptomyces/metabolismo , Carbono-Carbono Liases , China , Endófitos/isolamento & purificação , Transferência Genética Horizontal , Mesorhizobium/genética , Fixação de Nitrogênio , Filogenia , Desenvolvimento Vegetal/fisiologia , Sideróforos
15.
J Microbiol ; 58(2): 131-141, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31872373

RESUMO

Response regulator (RR) is known a protein that mediates cell's response to environmental changes. The effect of RR from extremophiles was still under investigation. In this study, response regulator homologs were mined from NGS data of Antarctic bacteria and overexpressed in Escherichia coli. Sixteen amino acid sequences were annotated corresponding to response regulators related to the two-component regulatory systems; of these, 3 amino acid sequences (DRH632, DRH1601 and DRH577) with high homology were selected. These genes were cloned in pRadGro and expressed in E. coli. The transformant strains were subjected to various abiotic stresses including oxidative, osmotic, thermal stress, and acidic stress. There was found that the robustness of E. coli to abiotic stress was increased in the presence of these response regulator homologs. Especially, recombinant E. coli overexpressing drh632 had the highest survival rate in oxidative, hypothermic, osmotic, and acidic conditions. Recombinant E. coli overexpressing drh1601 showed the highest tolerance level to osmotic stress. These results will be applicable for development of recombinant strains with high tolerance to abiotic stress.


Assuntos
Extremófilos/genética , Regulação Bacteriana da Expressão Gênica , Estresse Oxidativo/genética , Estresse Fisiológico/genética , Regiões Antárticas , Bacillus/genética , Bacillus/metabolismo , Bacillus pumilus/genética , Bacillus pumilus/metabolismo , Proteínas de Bactérias/genética , Clonagem Molecular , Biologia Computacional , Escherichia coli/genética , Escherichia coli/fisiologia , Extremófilos/metabolismo , Genoma Bacteriano , Estresse Oxidativo/fisiologia , Estresse Fisiológico/fisiologia , Transformação Bacteriana
16.
Appl Microbiol Biotechnol ; 104(2): 833-852, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31848654

RESUMO

Bacillus pumilus, an endospore-forming soil bacterium, produces a wide array of extracellular proteins, such as proteases, which are already applied in the chemical, detergent and leather industries. Small noncoding regulatory RNAs (sRNAs) in bacteria are important RNA regulators that act in response to various environmental signals. Here, an RNA-seq-based transcriptome analysis was applied to B. pumilus SCU11, a strain that produces extracellular alkaline protease, across various growth phases of the protease fermentation process. Through bioinformatics screening of the sequencing data and visual inspection, 84 putative regulatory sRNAs were identified in B. pumilus, including 21 antisense sRNAs and 63 sRNAs in intergenic regions. We experimentally validated the expression of 48 intergenic sRNAs by quantitative RT-PCR (qRT-PCR). Meanwhile, the expression of 6 novel sRNAs was confirmed by northern blotting, and the expression profiles of 5 sRNAs showed close correlation with the growth phase. We revealed that the sRNA Bpsr137 was involved in flagellum and biofilm formation in B. pumilus. The identification of a global set of sRNAs increases the inventory of regulatory sRNAs in Bacillus and implies the important regulatory roles of sRNA in B. pumilus. These findings will contribute another dimension to the optimization of crucial metabolic activities of B. pumilus during a productive fermentation process.


Assuntos
Bacillus pumilus/crescimento & desenvolvimento , Bacillus pumilus/genética , Peptídeo Hidrolases/metabolismo , Pequeno RNA não Traduzido/biossíntese , Bacillus pumilus/metabolismo , Northern Blotting , Biologia Computacional , Fermentação , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Pequeno RNA não Traduzido/genética , Reação em Cadeia da Polimerase em Tempo Real , Análise de Sequência de RNA
17.
Pol J Microbiol ; 69(3): 357-365, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33574865

RESUMO

The capacity of four bacterial strains isolated from productive soil potato fields to solubilize tricalcium phosphate on Pikovskaya agar or in a liquid medium was evaluated. A bacterial strain was selected to evaluate in vitro capacity of plant-growth promotion on Solanum tuberosum L. culture. Bacterial strain A3 showed the highest value of phosphate solubilization, reaching a 20 mm-diameter halo and a concentration of 350 mg/l on agar and in a liquid medium, respectively. Bacterial strain A3 was identified by 16S rDNA analysis as Bacillus pumilus with 98% identity; therefore, it is the first report for Bacillus pumilus as phosphate solubilizer. Plant-growth promotion assayed by in vitro culture of potato microplants showed that the addition of bacterial strain A3 increased root and stems length after 28 days. It significantly increased stem length by 79.3%, and duplicated the fresh weight of control microplants. In this paper, results reported regarding phosphorus solubilization and growth promotion under in vitro conditions represent a step forward in the use of innocuous bacterial strain biofertilizer on potato field cultures.


Assuntos
Bactérias/metabolismo , Fosfatos/metabolismo , Microbiologia do Solo , Solanum tuberosum/crescimento & desenvolvimento , Bacillus pumilus/classificação , Bacillus pumilus/genética , Bacillus pumilus/isolamento & purificação , Bacillus pumilus/metabolismo , Bactérias/classificação , Bactérias/genética , Bactérias/isolamento & purificação , Cinética , Filogenia , Raízes de Plantas/crescimento & desenvolvimento , Caules de Planta/crescimento & desenvolvimento , RNA Ribossômico 16S/genética , Rizosfera , Solanum tuberosum/metabolismo , Sacarose/metabolismo
18.
Appl Microbiol Biotechnol ; 103(20): 8375-8381, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31444521

RESUMO

Because of the emergence of antibiotic resistance, we must investigate new antibiotical agents. The present study was designed to find new compounds with antibacterial activity from metabolites of Bacillus pumilus. We found that the concentrated fermentation broth of Bacillus pumilus has antibacterial property. By high-performance liquid chromatography (HPLC), three compounds with antibacterial activity were first isolated from the ethyl acetate layer of fermentation broth of Bacillus pumilus. And then their structures were identified by nuclear magnetic resonance (NMR) and mass spectrometry (MS) analysis. According to the data, the compound 1, compound 2, and compound 3 were determined to be 3,4-dipentylhexane-2,5-diol, 1,1'-(4,5-dibutylcyclohexane-1,2-diyl)bis(ethan-1-ol), and 1,1'-(4,5-dibutyl-3,6-dimethylcyclohexane-1,2-diyl)bis(ethan-1-one). And all of them exhibited potent inhibitory effects against a panel of pathogenic bacteria including Staphylococcus aureus ATCC6538, Micrococcus luteus CMCC28001, Variant Salmonella gallinarum CVCC79207, Pasteurella multocida CVCC474, Swine Salmonella, Salmonella enterica ATCC13076, Swine Escherichia coli K88, Chicken Escherichia coli O78. Given its antibacterial activity, 3,4-dipentylhexane-2,5-diol, 1,1'-(4,5-dibutylcyclohexane-1,2-diyl)bis(ethan-1-ol), 1,1'-(4,5-dibutyl-3,6-dimethylcyclohexane-1,2-diyl)bis(ethan-1-one) are assumed to be promising agents for further development as antibacterial agents.


Assuntos
Antibacterianos/isolamento & purificação , Bacillus pumilus/metabolismo , Meios de Cultura/química , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Antibacterianos/química , Bacillus pumilus/crescimento & desenvolvimento , Cromatografia Líquida de Alta Pressão , Espectroscopia de Ressonância Magnética , Testes de Sensibilidade Microbiana
19.
BMC Genomics ; 20(1): 327, 2019 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-31039790

RESUMO

BACKGROUND: Bacillus pumilus is a Gram-positive and endospore-forming bacterium broadly existing in a variety of environmental niches. Because it produces and secrets many industrially useful enzymes, a lot of studies have been done to understand the underlying mechanisms. Among them, scoC was originally identified as a pleiotropic transcription factor negatively regulating protease production and sporulation in B. subtilis. Nevertheless, its role in B. pumilus largely remains unknown. RESULTS: In this study we successfully disrupted scoC gene in B. pumilus BA06 and found increased total extracellular protease activity in scoC mutant strain. Surprisingly, we also found that scoC disruption reduced cell motility possibly by affecting flagella formation. To better understand the underlying mechanism, we performed transcriptome analysis with RNA sequencing. The result showed that more than one thousand genes were alternated at transcriptional level across multiple growth phases, and among them the largest number of differentially expressed genes (DEGs) were identified at the transition time point (12 h) between the exponential growth and the stationary growth phases. In accordance with the altered phenotype, many protease genes especially the aprE gene encoding alkaline protease were transcriptionally regulated. In contrast to the finding in B. subtilis, the aprN gene encoding neutral protease was transcriptionally downregulated in B. pumilus, implicating that scoC plays strain-specific roles. CONCLUSIONS: The pleiotropic transcription factor ScoC plays multiple roles in various cellular processes in B. pumilus, some of which were previously reported in B. subtilis. The supervising finding is the identification of ScoC as a positive regulator for flagella formation and bacterial motility. Our transcriptome data may provide hints to understand the underlying mechanism.


Assuntos
Bacillus pumilus/genética , Proteínas de Bactérias/antagonistas & inibidores , Regulação Bacteriana da Expressão Gênica , Pleiotropia Genética , Transcriptoma , Bacillus pumilus/citologia , Bacillus pumilus/crescimento & desenvolvimento , Bacillus pumilus/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Movimento Celular , Endopeptidases/metabolismo , Flagelos/fisiologia , Fenótipo
20.
Chemosphere ; 223: 448-454, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30784751

RESUMO

Arsenic contaminated in rice plants can cause many physiological, biochemical and productivity in rice. This also had a negative impact on human health. To decrease arsenic in grains, a combination of leonardite as amendment and arsenic-resistance endophytic bacteria was investigated. The results showed that 1% (w/v) leonardite (91.86 ±â€¯2.04%) had the highest efficiency in adsorbing initial arsenic concentration of 2 mg L-1, which was higher than bagasse fly ash (16.25 ±â€¯3.97%), rice husk ash (10.36 ±â€¯1.28%), and sawdust fly ash (63.00 ±â€¯5.67%) under the same condition. This was due to the higher aluminium and iron contents of leonardite strongly binding to arsenic anions. Meanwhile, Bacillus pumilus had an ability to decrease arsenic accumulation in rice grains to levels below those achieved by Pseudomonas sp. and Bacillus thuringiensis. This was possibly due to B. pumilus producing higher siderophore. Interestingly, a combination of microbe and leonardite addition could decrease arsenic accumulation in grains to below the permissible limit (0.2 mg As kg-1 for inorganic arsenic). It could also reduce oxidative stress and showed down-regulation of Lsi1, Lsi2 and OsPT4 at the heading stage, which coincided with low arsenic and high silicon accumulation in roots. Therefore, this result could be used to decrease arsenic accumulation in grains in arsenic-contaminated paddy fields, improved rice plants defense and endured of arsenic stress, and increased rice productivity.


Assuntos
Arsênio/metabolismo , Bactérias/metabolismo , Endófitos/metabolismo , Minerais/farmacologia , Oryza/metabolismo , Bacillus pumilus/metabolismo , Grão Comestível/química , Humanos , Raízes de Plantas/metabolismo , Poluentes do Solo
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