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1.
Extremophiles ; 24(6): 923-935, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-33030592

RESUMO

The aerobic thermoalkaliphile Caldalkalibacillus thermarum strain TA2.A1 is a member of a separate order of alkaliphilic bacteria closely related to the Bacillales order. Efforts to relate the genomic information of this evolutionary ancient organism to environmental adaptation have been thwarted by the inability to construct a complete genome. The existing draft genome is highly fragmented due to repetitive regions, and gaps between and over repetitive regions were unbridgeable. To address this, Oxford Nanopore Technology's MinION allowed us to span these repeats through long reads, with over 6000-fold coverage. This resulted in a single 3.34 Mb circular chromosome. The profile of transporters and central metabolism gives insight into why the organism prefers glutamate over sucrose as carbon source. We propose that the deamination of glutamate allows alkalization of the immediate environment, an excellent example of how an extremophile modulates environmental conditions to suit its own requirements. Curiously, plant-like hallmark electron transfer enzymes and transporters are found throughout the genome, such as a cytochrome b6c1 complex and a CO2-concentrating transporter. In addition, multiple self-splicing group II intron-encoded proteins closely aligning to those of a telomerase reverse transcriptase in Arabidopsis thaliana were revealed. Collectively, these features suggest an evolutionary relationship to plant life.


Assuntos
Bacillaceae/genética , Bacillaceae/metabolismo , Genômica , Aerobiose , Evolução Biológica , Sequências Repetitivas de Ácido Nucleico
2.
Nat Commun ; 11(1): 2837, 2020 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-32503992

RESUMO

Group II introns are ubiquitous self-splicing ribozymes and retrotransposable elements evolutionarily and chemically related to the eukaryotic spliceosome, with potential applications as gene-editing tools. Recent biochemical and structural data have captured the intron in multiple conformations at different stages of catalysis. Here, we employ enzymatic assays, X-ray crystallography, and molecular simulations to resolve the spatiotemporal location and function of conformational changes occurring between the first and the second step of splicing. We show that the first residue of the highly-conserved catalytic triad is protonated upon 5'-splice-site scission, promoting a reversible structural rearrangement of the active site (toggling). Protonation and active site dynamics induced by the first step of splicing facilitate the progression to the second step. Our insights into the mechanism of group II intron splicing parallels functional data on the spliceosome, thus reinforcing the notion that these evolutionarily-related molecular machines share the same enzymatic strategy.


Assuntos
Íntrons/genética , Precursores de RNA/metabolismo , Processamento de RNA , RNA Bacteriano/metabolismo , Spliceossomos/metabolismo , Bacillaceae/genética , Domínio Catalítico/genética , Cristalografia por Raios X , Simulação de Dinâmica Molecular , Mutagênese , Conformação de Ácido Nucleico , Precursores de RNA/genética , RNA Bacteriano/genética , Análise Espaço-Temporal
3.
Arch Microbiol ; 202(9): 2373-2378, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32583126

RESUMO

A Gram-positive, endospore-forming, rod-shaped bacterium with a single flagellum, and a motile strain, designated CX253, was isolated from bioaerosols. The isolate is facultatively anaerobic, is able to grow at 25-45 â„ƒ (optimum 37 â„ƒ) and pH 6.5-10.0 (optimum 7.5), and can tolerate up to 5.0% NaCl (w/v) under aerobic conditions. The diagnostic diamino acid in the cell wall of strain CX253T is meso-diaminopimelic acid, while major isoprenoid quinone is menaquinone 6 (MK-6) along with a smaller amount of MK-7 (20%). The polar lipid profile is composed of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phospholipids and glycolipids. The major cellular fatty acid is iso-C15:0 and anteiso-C15:0. Phylogenetic analysis based on 16S rRNA gene and genome sequence grouped strain CX253T into the genus Bacillus. The strain was most closely related to Bacillus thermotolerans CCTCC AB 2012108 T by comparison of 16S rRNA gene sequence (97.2% similarity) and to Bacillus wudalianchiensis CCTCC AB 2015266 T by comparison of gyrB gene sequence (80.1% similarity). The draft genome of strain CX253T comprised 3,929,195 bp with a G + C content of 43.3 mol%. The average nucleotide identity and digital DNA-DNA hybridization values between strain CX253T and phylogenetically related Bacillus species were lower than 95% and 70%, respectively. Thus, the polyphasic evidence generated through phenotypic, chemotaxonomic and genomic methods confirmed that strain CX253T (= GDMCC 1.1608 T = KACC 21318 T) was a novel species of the genus Bacillus, for which the name Bacillus aerolatus sp. nov. is proposed.


Assuntos
Aerossóis , Bacillus/classificação , Microbiologia Ambiental , Bacillaceae/genética , Bacillus/genética , Bacillus/isolamento & purificação , Técnicas de Tipagem Bacteriana , Composição de Bases , Parede Celular/química , Ácido Diaminopimélico/análise , Ácidos Graxos/análise , Glicolipídeos/análise , Hibridização de Ácido Nucleico , Fosfolipídeos/análise , Filogenia , RNA Ribossômico 16S/genética , Especificidade da Espécie
4.
PLoS One ; 15(5): e0232742, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32374788

RESUMO

Caves formed by sulfuric acid dissolution have been identified worldwide. These caves can host diverse microbial communities that are responsible for speleogenesis and speleothem formation. It is not well understood how microbial communities change in response to surface water entering caves. Illumina 16S rRNA sequencing and bioinformatic tools were used to determine the impact of surface water on the microbial community diversity and function within a spring pool found deep in the Monte Conca Cave system in Sicily, Italy. Sulfur oxidizers comprised more than 90% of the microbial community during the dry season and were replaced by potential anthropogenic contaminants such as Escherichia and Lysinibacillus species after heavy rains. One sampling date appeared to show a transition between the wet and dry seasons when potential anthropogenic contaminants (67.3%), sulfur-oxidizing bacteria (13.6%), and nitrogen-fixing bacteria (6.5%) were all present within the spring pool.


Assuntos
Cavernas/microbiologia , Microbiota/genética , Microbiologia da Água , Molhabilidade , Bacillaceae/genética , Sequência de Bases , DNA Bacteriano/genética , Secas , Escherichia/genética , Sulfeto de Hidrogênio/análise , RNA Ribossômico 16S/genética , Chuva , Estações do Ano , Sicília , Microbiologia do Solo , Sulfatos/análise
5.
PLoS One ; 15(4): e0231426, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32271848

RESUMO

Demand for agricultural crop continues to escalate in response to increasing population and damage of prime cropland for cultivation. Research interest is diverted to utilize soils with marginal plant production. Moisture stress has negative impact on crop growth and productivity. The plant growth promoting rhizobacteria (PGPR) and plant growth regulators (PGR) are vital for plant developmental process under moisture stress. The current study was carried out to investigate the effect of PGPR and PGRs (Salicylic acid and Putrescine) on the physiological activities of chickpea grown in sandy soil. The bacterial isolates were characterized based on biochemical characters including Gram-staining, P-solubilisation, antibacterial and antifungal activities and catalases and oxidases activities and were also screened for the production of indole-3-acetic acid (IAA), hydrogen cyanide (HCN) and ammonia (NH3). The bacterial strains were identified as Bacillus subtilis, Bacillus thuringiensis and Bacillus megaterium based on the results of 16S-rRNA gene sequencing. Chickpea seeds of two varieties (Punjab Noor-2009 and 93127) differing in sensitivity to drought were soaked for 3 h before sowing in fresh grown cultures of isolates. Both the PGRs were applied (150 mg/L), as foliar spray on 20 days old seedlings of chickpea. Moisture stress significantly reduced the physiological parameters but the inoculation of PGPR and PGR treatment effectively ameliorated the adverse effects of moisture stress. The result showed that chickpea plants treated with PGPR and PGR significantly enhanced the chlorophyll, protein and sugar contents. Shoot and root fresh (81%) and dry weights (77%) were also enhanced significantly in the treated plants. Leaf proline content, lipid peroxidation and antioxidant enzymes (CAT, APOX, POD and SOD) were increased in reaction to drought stress but decreased due to PGPR. The plant height (61%), grain weight (41%), number of nodules (78%) and pod (88%), plant yield (76%), pod weight (53%) and total biomass (54%) were higher in PGPR and PGR treated chickpea plants grown in sandy soil. It is concluded from the present study that the integrative use of PGPR and PGRs is a promising method and eco-friendly strategy for increasing drought tolerance in crop plants.


Assuntos
Agricultura , Bacillaceae/fisiologia , Cicer/crescimento & desenvolvimento , Reguladores de Crescimento de Planta/farmacologia , Amônia/metabolismo , Bacillaceae/genética , Bacillaceae/isolamento & purificação , Bacillus megaterium/genética , Bacillus megaterium/isolamento & purificação , Bacillus subtilis/genética , Bacillus subtilis/isolamento & purificação , Bacillus subtilis/fisiologia , Biomassa , Clorofila/análise , Cicer/efeitos dos fármacos , Cicer/metabolismo , Ácidos Indolacéticos/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Reguladores de Crescimento de Planta/metabolismo , Folhas de Planta/metabolismo , Proteínas de Plantas/metabolismo , Raízes de Plantas/metabolismo , Raízes de Plantas/microbiologia , Putrescina/metabolismo , Putrescina/farmacologia , RNA Ribossômico 16S/química , RNA Ribossômico 16S/metabolismo , Chuva , Ácido Salicílico/metabolismo , Ácido Salicílico/farmacologia , Plântula/efeitos dos fármacos , Plântula/crescimento & desenvolvimento , Microbiologia do Solo
6.
Arch Microbiol ; 202(6): 1545-1549, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32246163

RESUMO

A bacterial strain, designated YIM 98839T, was isolated from the hypersaline sediment of Aiding Lake in Xinjiang province, North-West China. The strain was Gram-stain-positive, motile, aerobic, produced oval subterminal or central endospores in swollen sporangia. The whole-cell hydrolysates contain meso-diaminopimelic acid as the diagnostic cell-wall diamino acid. Galactose, fucose and ribose are the major whole-cell sugars. The phospholipids are diphosphatidylglycerol, phosphatidylglycerol and one unknown phospholipid. The predominant menaquinone is MK-7. The major fatty acids are anteiso-C15:0, anteiso-C17:0 and iso-C15:0. The DNA G + C content of the type strain is 37.0 mol%. Phylogenetic analysis indicated that the isolate belongs to the genus Oceanobacillus. However, it differed from its closest relative, Oceanobacillus limi H9BT in many physiological characteristics. Moreover, the DNA-DNA relatedness values between the novel isolate and the relative type strain was 20.2%. Based on comparative analysis of polyphasic taxonomic data, strain YIM 98839T represents a novel species of the genus Oceanobacillus, for which the name Oceanobacillus halotolerans sp. nov. is proposed. The type strain is YIM 98839T (= CGMCC 1.17002T = KCTC 43140T).


Assuntos
Bacillaceae/classificação , Bacillaceae/genética , Bacillaceae/isolamento & purificação , Técnicas de Tipagem Bacteriana , Composição de Bases/genética , Parede Celular/química , China , DNA Bacteriano/genética , Ácido Diaminopimélico/análise , Ácidos Graxos/análise , Lagos/microbiologia , Fosfolipídeos/análise , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
7.
Arch Microbiol ; 202(6): 1469-1476, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32193578

RESUMO

A novel Gram-stain-positive, motile, moderate halophile, strain L9T, was isolated from hides of white goat in China. The isolate grew optimally at 30 °C, at pH 7 and with 5-10% (w/v) NaCl. The predominant menaquinone was MK-7, and the major cellular fatty acids were identified as iso-C15:0 and anteiso-C15:0. The peptidoglycan amino acid type was determined to be A4ß, containing L-ornithine and D-aspartic as diagnostic amino acids. The phospholipids were dominated by diphosphatidylglycerol, phosphatidylglycerol, one unidentified aminophospholipid and two unidentified phospholipids. Genome sequencing resulted in a genome size of 4.0 Mbp and a DNA G + C content of 35.9 mol%. Phylogenetic trees based on the 16S rRNA gene sequences showed the isolate to be closely related to Oceanobacillus limi H9BT (98.2% similarity) and Ornithinibacillus halophilus G8BT (97.5% similarity). The ANI and dDDH values between strain L9T and the closely related species were 69.8-76.1% and 13.0-20.5%, respectively. On the basis of the data presented, strain L9T represents a novel species of the genus Ornithinibacillus, for which the name Ornithinibacillus caprae sp. nov. is proposed. The type strain is L9T (= KCTC 43176T = CGMCC 1.17659T).


Assuntos
Bacillaceae/classificação , Bacillaceae/isolamento & purificação , Cabras/microbiologia , Aminoácidos/análise , Animais , Bacillaceae/genética , Técnicas de Tipagem Bacteriana , Composição de Bases/genética , DNA Bacteriano/genética , Ácidos Graxos/análise , Peptidoglicano/química , Fosfolipídeos/análise , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Cloreto de Sódio/metabolismo
8.
Ecotoxicol Environ Saf ; 192: 110250, 2020 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-32028154

RESUMO

A bacterial strain designated Lysinibacillus fusiformis 15-4 was isolated from oil-free soil on the Qinghai-Tibet Plateau, which can grow well utilizing petroleum hydrocarbons as a carbon source at a lower temperature. To deeply characterize the molecular adaptations and metabolic processes of this strain when grown in a petroleum-containing environment, transcriptome analysis was performed. A total of 4664 genes and the expression of 3969 genes were observed in strain 15-4. When the strain was grown in petroleum-containing medium, 2192 genes were significantly regulated, of which 1312 (60%) were upregulated and 880 (40%) were downregulated. This strain degraded and adapted to petroleum via modulation of diverse molecular processes, including improvements in transporter activity, oxidoreductase/dehydrogenase activity, two-component system/signal transduction, transcriptional regulation, fatty acid catabolism, amino acid metabolism, and environmental stress responses. Many strain-specific genes were involved in the oxidation of hydrocarbon compounds, such as several luciferase family alkane monooxygenase genes, flavin-utilizing monooxygenase family genes, and flavoprotein-like family alkanesulfonate monooxygenase genes. Several cold shock protein genes were also induced suggesting adaptation to cold environments and the potential for petroleum degradation at low temperatures. The results obtained in this study may broaden our understanding of molecular adaptation of bacteria to hydrocarbon-containing environments and may provide valuable data for further study of L. fusiformis.


Assuntos
Bacillaceae/genética , Bacillaceae/metabolismo , Petróleo/metabolismo , Transportadores de Cassetes de Ligação de ATP/metabolismo , Adaptação Fisiológica , Bacillaceae/isolamento & purificação , Biodegradação Ambiental , Proteínas e Peptídeos de Choque Frio/biossíntese , Proteínas e Peptídeos de Choque Frio/genética , Temperatura Baixa , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Hidrocarbonetos/metabolismo , Oxigenases de Função Mista/biossíntese , Oxigenases de Função Mista/genética , Microbiologia do Solo , Tibet
9.
PLoS Comput Biol ; 15(12): e1007564, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31860665

RESUMO

In comparison to protein coding sequences, the impact of mutation and natural selection on the sequence and function of non-coding (ncRNA) genes is not well understood. Many ncRNA genes are narrowly distributed to only a few organisms, and appear to be rapidly evolving. Compared to protein coding sequences, there are many challenges associated with assessment of ncRNAs that are not well addressed by conventional phylogenetic approaches, including: short sequence length, lack of primary sequence conservation, and the importance of secondary structure for biological function. Riboswitches are structured ncRNAs that directly interact with small molecules to regulate gene expression in bacteria. They typically consist of a ligand-binding domain (aptamer) whose folding changes drive changes in gene expression. The glycine riboswitch is among the most well-studied due to the widespread occurrence of a tandem aptamer arrangement (tandem), wherein two homologous aptamers interact with glycine and each other to regulate gene expression. However, a significant proportion of glycine riboswitches are comprised of single aptamers (singleton). Here we use graph clustering to circumvent the limitations of traditional phylogenetic analysis when studying the relationship between the tandem and singleton glycine aptamers. Graph clustering enables a broader range of pairwise comparison measures to be used to assess aptamer similarity. Using this approach, we show that one aptamer of the tandem glycine riboswitch pair is typically much more highly conserved, and that which aptamer is conserved depends on the regulated gene. Furthermore, our analysis also reveals that singleton aptamers are more similar to either the first or second tandem aptamer, again based on the regulated gene. Taken together, our findings suggest that tandem glycine riboswitches degrade into functional singletons, with the regulated gene(s) dictating which glycine-binding aptamer is conserved.


Assuntos
Aptâmeros de Nucleotídeos/química , Aptâmeros de Nucleotídeos/genética , Glicina/química , Riboswitch/genética , Bacillaceae/classificação , Bacillaceae/genética , Biologia Computacional , Evolução Molecular , Genoma Bacteriano , Modelos Genéticos , Modelos Moleculares , Conformação de Ácido Nucleico , Filogenia , Vibrionaceae/classificação , Vibrionaceae/genética
10.
J Microbiol ; 57(11): 997-1002, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31659686

RESUMO

A Gram-stain-positive, rod-shaped, alkalitolerant, and halophilic bacterium-designated as strain NKC3-5T-was isolated from kimchi that was collected from the Geumsan area in the Republic of Korea. Cells of isolated strain NKC3-5T were 0.5-0.7 µm wide and 1.4-2.8 µm long. The strain NKC3-5T could grow at up to 20.0% (w/v) NaCl (optimum 10%), pH 6.5-10.0 (optimum pH 9.0), and 25-40°C (optimum 35°C). The cells were able to reduce nitrate under aerobic conditions, which is the first report in the genus Salicibibacter. The genome size and genomic G + C content of strain NKC3-5T were 3,754,174 bp and 45.9 mol%, respectively; it contained 3,630 coding sequences, 16S rRNA genes (six 16S, five 5S, and five 23S), and 59 tRNA genes. Phylogenetic analysis based on 16S rRNA showed that strain NKC3-5T clustered with bacterium Salicibibacter kimchii NKC1-1T, with a similarity of 96.2-97.6%, but formed a distinct branch with other published species of the family Bacillaceae. In addition, OrthoANI value between strain NKC3-5T and Salicibibacter kimchii NKC1-1T was far lower than the species demarcation threshold. Using functional genome annotation, the result found that carbohydrate, amino acid, and vitamin metabolism related genes were highly distributed in the genome of strain NKC3-5T. Comparative genomic analysis revealed that strain NKC3-5T had 716 pan-genome orthologous groups (POGs), dominated with carbohydrate metabolism. Phylogenomic analysis based on the concatenated core POGs revealed that strain NKC3-5T was closely related to Salicibibacter kimchii. The predominant polar lipids were phosphatidylglycerol and two unidentified lipids. Anteiso-C15:0, iso-C17:0, anteiso-C17:0, and iso-C15:0 were the major cellular fatty acids, and menaquinone-7 was the major isoprenoid quinone present in strain NKC3-5T. Cell wall peptidoglycan analysis of strain NKC3-5T showed that meso-diaminopimelic acid was the diagnostic diamino acid. The phephenotypic, genomic, phylogenetic, and chemotaxonomic properties reveal that the strain represents a novel species of the genus Salicibibacter, for which the name Salicibibacter halophilus sp. nov. is proposed, with the type strain NKC3-5T (= KACC 21230T = JCM 33437T).


Assuntos
Bacillaceae/classificação , Bacillaceae/isolamento & purificação , Alimentos e Bebidas Fermentados/microbiologia , Filogenia , Bacillaceae/genética , Bacillaceae/fisiologia , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácido Diaminopimélico/metabolismo , Ácidos Graxos/química , Genes Bacterianos/genética , Genômica , Halobacteriales , Concentração de Íons de Hidrogênio , Peptidoglicano/química , RNA Ribossômico 16S/genética , República da Coreia , Tolerância ao Sal , Análise de Sequência de DNA , Cloreto de Sódio/metabolismo , Vitamina K 2/análogos & derivados , Vitamina K 2/química , Sequenciamento Completo do Genoma
11.
Int J Food Microbiol ; 305: 108245, 2019 Sep 16.
Artigo em Inglês | MEDLINE | ID: mdl-31295678

RESUMO

This study aimed to evaluate the performance of Hicrome Bacillus™ agar for isolation and rapid identification of the aerobic spore-forming bacteria most frequently found in honey samples. A collection of 197 bacterial isolates of Bacillus, Brevibacillus, Lysinibacillus, Paenibacillus, and Rummeliibacillus belonging to different species that have been reported in honey were screened for their abilities to grow and for their colony colors and medium appearance in HiCrome Bacillus agar. Also, 21 strains from culture collections were used for comparison and quality controls. A flowchart utilizing a combination of colony and media characteristics in the chromogenic medium and a set of simple biochemical and morphological tests were elaborated for quick presumptive identification. A procedure for direct isolation from honey samples was developed. In conclusion, HiCrome Bacillus agar in combination with simple microbiological tests was highly useful for rapid and reliable identification of most Bacillus, Brevibacillus, Lysinibacillus and Paenibacillus species commonly found in honey samples facilitating isolation from polymicrobial honey.


Assuntos
Bacillaceae/isolamento & purificação , Bacillus/isolamento & purificação , Contagem de Colônia Microbiana/métodos , Mel/microbiologia , Bacillaceae/classificação , Bacillaceae/genética , Bacillaceae/crescimento & desenvolvimento , Bacillus/classificação , Bacillus/genética , Bacillus/crescimento & desenvolvimento , Contagem de Colônia Microbiana/instrumentação , Microbiologia de Alimentos
12.
Mol Biol Rep ; 46(4): 4385-4395, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31201678

RESUMO

The ferulic acid esterase (FAE) gene from Geobacillus thermoglucosidasius DSM 2542T was cloned into pET28a(+) expression vector and characterized and is being reported in this study for the first time in Geobacillus. The enzyme, designated as GthFAE, was purified by heat shock and ion-exchange column chromatography. In addition, a second clone containing a Histidine tag was expressed and purified by affinity column chromatography demonstrating future potential for scale-up. FAE gene contains an open reading frame (ORF) of 759-bp encoding a hypothetical 252 amino acid protein, a molecular mass of 28.11 kDa and an isoelectric point of 5.53. From this study it was found that GthFAE had optimal activity at 50 °C and pH of 8.5. Furthermore, the enzyme has been found to retain 64% of its activity after two days incubation at 50 °C and exhibited a high level of functionality with p-nitrophenyl caprylate (C8). Km, Vmax, kcat and kcat/Km values for p-nitrophenyl caprylate were determined as 0.035 mM, 11,735 µmol/min/mg protein, 5491 (1/s) and 156,885 s-1 mM-1 respectively. The combination of higher activity and stability compared to previously reported FAEs makes GthFAE a potential candidate for use in the paper manufacturing industry.


Assuntos
Bacillaceae/enzimologia , Bacillaceae/genética , Hidrolases de Éster Carboxílico/química , Sequência de Aminoácidos , Clonagem Molecular , Estabilidade Enzimática , Geobacillus/genética , Concentração de Íons de Hidrogênio , Cinética , Peso Molecular , Especificidade por Substrato
13.
ACS Synth Biol ; 8(7): 1642-1654, 2019 07 19.
Artigo em Inglês | MEDLINE | ID: mdl-31242391

RESUMO

Bacterial microcompartments (BMCs) are organelles that host specific biochemical reactions for both anabolic and catabolic functions. Engineered morphologically diverse BMCs bearing heterologous enzymatic pathways have shown enhanced productivity for commodity chemicals, which makes BMCs an important focus for metabolic engineering. Gaining control of BMC assembly and incorporation of a heterologous enzymatic cargo has yet to be achieved in thermophiles. Herein, we address this by first conducting a detailed bioinformatic analysis of the propanediol utilization (pdu) operon in the thermophile Parageobacillus thermoglucosidasius. We then demonstrated, in vivo, the ability to assemble the native BMCs at an elevated temperature of 60 °C. Heterologous expression of Pdu shell proteins from P. thermoglucosidasius in Bacillus subtilis resulted in the assembly of a single tubular BMC with an average length of 1.4 µm; BMCs assembled after a 20 min induction of expression of the shell operons. Moreover, we show that it is possible to target the monomeric superfolder GFP (msfGFP) to the interior of the compartment by fusion of an N-terminal sequence of the propanediol utilization protein (PduP) of at least 24 amino acids. This study establishes the feasibility of constructing cell factories for small molecules in industrially important Bacillus and Geobacillus spp. by heterologous cargo-carrying BMC production and assembly. Additionally, the study provides experimental confirmation that BMCs are produced in thermophilic bacteria, which opens a path for future research on repurposing the native organelles to provide new functionality at elevated temperatures.


Assuntos
Bacillaceae/genética , Bacillaceae/metabolismo , Aminoácidos/genética , Aminoácidos/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biologia Computacional/métodos , Geobacillus/genética , Geobacillus/metabolismo , Engenharia Metabólica/métodos , Óperon/genética , Organelas/genética , Organelas/metabolismo , Propilenoglicóis/metabolismo , Temperatura
14.
J Microbiol ; 57(7): 562-568, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31124045

RESUMO

A Gram-positive, aerobic, rod-shaped, spore-forming bacterium, designated YLB-03T, with peritrichous flagella was isolated from deep-sea sediment of the Yap Trench at a depth of 4435 m. The bacterium was found to be catalase-positive but oxidase-negative. Growth of this bacterium was observed at 15-50°C (optimum 37°C), pH 5-10.5 (optimum 7), 0-5% NaCl (optimum 1%, w/v) and 0.1-50 MPa (optimum 0.1 MPa). Phylogenetic analysis based on 16S rRNA gene sequences showed that strain YLB-03T was a member of the genus Lysinibacillus. Strain YLB-03T was closely related to Lysinibacillus sinduriensis BLB-1T and Lysinibacillus chungkukjangi 2RL3-2T (98.4%), Lysinibacillus halotolerans LAM-612T (98.0%), Lysinibacillus telephonicus KT735049T (97.5%), Lysinibacillus endophyticus C9T (97.5%), Lysinibacillus composti NCCP-36T and Lysinibacillus massiliensis 4400831T (97.3%). The ANI and the GGDC DNA-DNA hybridization estimate values between strain YLB-03T and closely related type strains were 73.7-76.3% and 34.7-38.7%, respectively. The principal fatty acids were anteiso-C15:0 and iso-C15:0. The G+C content of the chromosomal DNA was 39.6 mol%. The respiratory quinone was determined to be MK-7. The diagnostic amino acids in the cell wall peptidoglycan contained Lys-Asp (type A4α) and the cell-wall sugars were glucose and xylose. The polar lipids included diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, and an unidentified phospholipid. The combined genotypic and phenotypic data showed that strain YLB-03T represents a novel species within the genus Lysinibacillus, for which the name Lysinibacillus yapensis sp. nov. is proposed, with the type strain YLB-03T (= MCCC 1A12698T = JCM 32871T).


Assuntos
Bacillaceae/classificação , Sedimentos Geológicos/microbiologia , Bacillaceae/genética , Bacillaceae/isolamento & purificação , Técnicas de Tipagem Bacteriana/métodos , Oceano Pacífico , Filogenia , RNA Ribossômico 16S/genética
15.
Syst Appl Microbiol ; 42(3): 360-372, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30982666

RESUMO

The species Bacillus badius is one of the oldest members of the genus Bacillus isolated from faeces of children and was classified based on its ability to form endospores [8]. In 16S rRNA gene sequence and phylogenetic analysis, Bacillus badius DSM 23T shared low similarity (93.0%) and distant relationship with B. subtilis, the type species of the genus Bacillus indicating that it does not belong to this genus. Additional strains of the species, B. badius DSM 5610, DSM 30822 and B. encimensis SGD-V-25 (which has been recently reclassified as a member of this species) were included in the study to consider intraspecies diversity. Detailed molecular phylogenetic and comparative genome analysis clearly showed that the strains of B. badius were consistently retrieved outside the cluster of Bacillus sensu stricto and also distantly related to the genera Domibacillus and Quasibacillus. Further, the data from biochemical reactions (inability to ferment most carbohydrates), polar lipids profile (presence of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and an aminophosphoglycolipid) and fatty acids supported the molecular analysis. Thus the four B. badius strains; DSM 23T, DSM 5610, DSM 30822 and SGD-V-25 displayed sufficient demarcating phenotypic characteristics that warrant their classification as members of a novel genus and single species, for which the name Pseudobacillus badius gen. nov. comb. nov. is proposed with Pseudobacillus badius DSM 23T (= ATCC 14574T) as the type strain. Additionally, based on our findings from phenotypic, chemotaxonomic and genotypic parameters, Bacillus wudalianchiensis DSM 100757T was reclassified as Pseudobacillus wudalianchiensis comb. nov.


Assuntos
Bacillaceae/classificação , Filogenia , Bacillaceae/química , Bacillaceae/genética , DNA Bacteriano/genética , Ácidos Graxos/análise , Genoma Bacteriano/genética , Lipídeos/análise , Hibridização de Ácido Nucleico , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Especificidade da Espécie , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
16.
Nat Commun ; 10(1): 1115, 2019 03 07.
Artigo em Inglês | MEDLINE | ID: mdl-30846700

RESUMO

The genome of the thermophilic bacterium, Aeribacillus pallidus 8, encodes the bacteriocin pallidocin. It belongs to the small class of glycocins and is posttranslationally modified, containing an S-linked glucose on a specific Cys residue. In this study, the pallidocin biosynthetic machinery is cloned and expressed in Escherichia coli to achieve its full biosynthesis and modification. It targets other thermophilic bacteria with potent activity, demonstrated by a low minimum inhibitory concentration (MIC) value. Moreover, the characterized biosynthetic machinery is employed to produce two other glycopeptides Hyp1 and Hyp2. Pallidocin and Hyp1 exhibit antibacterial activity against closely related thermophilic bacteria and some Bacillus sp. strains. Thus, heterologous expression of a glycocin biosynthetic gene cluster including an S-glycosyltransferase provides a good tool for production of hypothetical glycocins encoded by various bacterial genomes and allows rapid in vivo screening.


Assuntos
Bacillaceae/metabolismo , Bacteriocinas/biossíntese , Sequência de Aminoácidos , Antibacterianos/biossíntese , Antibacterianos/química , Antibacterianos/farmacologia , Bacillaceae/genética , Bacteriocinas/genética , Bacteriocinas/farmacologia , Clonagem Molecular , Dissulfetos/química , Escherichia coli/genética , Escherichia coli/metabolismo , Genes Bacterianos , Glicopeptídeos/biossíntese , Glicopeptídeos/genética , Glicopeptídeos/farmacologia , Testes de Sensibilidade Microbiana , Família Multigênica , Estrutura Terciária de Proteína , Homologia de Sequência de Aminoácidos
17.
J Appl Microbiol ; 126(6): 1742-1750, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30817048

RESUMO

AIMS: Hot springs have always drawn attention due to their unique chemical richness and the presence of different microbial communities. The use of hot spring bacteria in concrete technology is our primary focus; isolation of an alkaliphilic bacterium from the Bakreshwar hot springs having longer survival and better efficacy towards cementitious environment was the basis of our study's design. METHODS AND RESULTS: A novel facultative anaerobic and highly alkaliphilic bacterial strain (BKH4; GenBank accession no. KX622782) belonging to the family 'Bacillaceae' and homologous (99%) with Lysinibacillus fusiformis was isolated from Bakreshwar hot springs. The isolated coccoid-type Gram-positive bacterium grows well in a defined semi-synthetic medium (pH 12·0 and 65°C). This bacterium survives for more than a month and shows better efficacy in enhancing compressive strengths (>50%), ultrasonic pulse velocity (>25%) and durability of the cementitious mortar when incorporated at a concentration of 104  cells per ml of water used. CONCLUSION: The novel bacterium BKH4 is more effective for the enhancement of the bioconcrete properties. SIGNIFICANCE AND IMPACT OF THE STUDY: BKH4 bacterium will add a new dimension to future concrete technology for its usefulness in strength enhancement and durability due to its alkaliphilic nature and longer survival within a cementitious environment.


Assuntos
Álcalis/metabolismo , Bacillaceae/fisiologia , Biotecnologia , Materiais de Construção/microbiologia , Fontes Termais/microbiologia , Bacillaceae/genética , Bacillaceae/metabolismo , Força Compressiva , DNA Bacteriano/genética , Temperatura Alta , Concentração de Íons de Hidrogênio , Hibridização de Ácido Nucleico , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
18.
Antonie Van Leeuwenhoek ; 112(8): 1161-1167, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30820713

RESUMO

A strain of a Gram-positive, strictly aerobic, motile, rod-shaped, endospore forming bacterium was originally isolated from rhizospheric soil of a pepper plant when screening and bioprospecting for plant beneficial microorganisms. Phylogenetic analysis of the 16S rRNA gene sequences indicated that this strain, PB300T, is closely related to Lysinibacillus macroides DMS 54T (99.6%) and Lysinibacillus xylanilyticus DSM 23493T (99.4%). In phenotypic characterisation, the novel strain was found to grow between 15 and 40 °C and tolerate up to 10% (w/v) NaCl. Furthermore, the strain was found to grow in media with pH 5 to 10 (optimal growth at pH 7.0). The predominant cellular fatty acids were observed to be iso-C15 : 0 (56.6 %), anteiso-C15 : 0 (14.6%), C16 :1ω7C alcohol (9.3%) and C16 : 0 (7.1%). The cell wall peptidoglycan contains lysine-aspartic acid, as in its close relatives. A draft genome was completed and the DNA G + C content was determined to be 37.5% (mol content). A phylogenomic analysis of the core genome of the new strain and 5 closely related type strains of the genus Lysinibacillus revealed that this strain formed a distinct monophyletic clade with the nearest neighbour being Lysinibacillus boronitolerans. DNA-DNA relatedness studies using in silico DNA-DNA hybridizations (DDH) showed relationships for the new strain were below the species threshold of 70%. Based upon the consensus of phylogenetic and phenotypic analyses, we conclude that this strain represents a novel species within the genus Lysinibacillus, for which the name Lysinibacillus capsici sp. nov. is proposed, with type strain PB300T (= NRRL B-65515T, = CCUG 72241T).


Assuntos
Bacillaceae/classificação , Bacillaceae/isolamento & purificação , Microbiologia do Solo , Bacillaceae/química , Bacillaceae/genética , Técnicas de Tipagem Bacteriana , Composição de Bases , Capsicum/crescimento & desenvolvimento , Parede Celular/química , Análise por Conglomerados , Citosol/química , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Ácidos Graxos/análise , Hibridização de Ácido Nucleico , Peptidoglicano/análise , Filogenia , RNA Ribossômico 16S/genética , Rizosfera , Análise de Sequência de DNA
19.
Antonie Van Leeuwenhoek ; 112(6): 897-903, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30788630

RESUMO

A novel Gram-stain positive, short rod, forming sub-terminal endospores of ellipsoidal shape, halophilic, alkaliphilic and aerobic bacterium, designated strain KQ-12T, was isolated from a saline-alkaline lake in China, and characterised by a polyphasic taxonomic approach. The isolate grew at 4-40 °C (optimum, 25 °C), at pH 8.0-10.0 (pH 9.0) and in the presence of 0-16% (w/v) NaCl (8%). 16S rRNA gene sequence similarity of KQ-12T to species in the genera Salipaludibacillus ranged from 96.6 to 98.1%. Phylogenetic trees indicated that the strain should be assigned to the genus Salipaludibacillus. The polar lipids of KQ-12T were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, and an unidentified phospholipid and its major cellular fatty acids were anteiso-C15:0, anteiso-C17:0, iso-C15:0, and C16:0. The isoprenoid quinone was MK-7. These key chemotaxonomic properties also confirmed the affiliation of the strain to the genus Salipaludibacillus. However, some physiological, biochemical properties, low average nucleotide identity and low digital DNA-DNA hybridization relatedness values enabled the strain to be differentiated from closely related species of the genus Salipaludibacillus. Thus, KQ-12T can be classified as a novel species in the genus Salipaludibacillus, for which the name Salipaludibacillus keqinensis sp. nov. is proposed. The type strain is KQ-12T ( =  ACCC 60430T   =  KCTC 33935T).


Assuntos
Bacillaceae/isolamento & purificação , Lagos/microbiologia , Bacillaceae/classificação , Bacillaceae/genética , Bacillaceae/metabolismo , Técnicas de Tipagem Bacteriana , China , DNA Bacteriano/genética , Ácidos Graxos/química , Ácidos Graxos/metabolismo , Lagos/análise , Filogenia , RNA Ribossômico 16S/genética , Cloreto de Sódio/análise , Cloreto de Sódio/metabolismo
20.
Appl Environ Microbiol ; 85(7)2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-30683748

RESUMO

Permafrost hosts a community of microorganisms that survive and reproduce for millennia despite extreme environmental conditions, such as water stress, subzero temperatures, high salinity, and low nutrient availability. Many studies focused on permafrost microbial community composition use DNA-based methods, such as metagenomics and 16S rRNA gene sequencing. However, these methods do not distinguish among active, dead, and dormant cells. This is of particular concern in ancient permafrost, where constant subzero temperatures preserve DNA from dead organisms and dormancy may be a common survival strategy. To circumvent this, we applied (i) LIVE/DEAD differential staining coupled with microscopy, (ii) endospore enrichment, and (iii) selective depletion of DNA from dead cells to permafrost microbial communities across a Pleistocene permafrost chronosequence (19,000, 27,000, and 33,000 years old). Cell counts and analysis of 16S rRNA gene amplicons from live, dead, and dormant cells revealed how communities differ between these pools, how they are influenced by soil physicochemical properties, and whether they change over geologic time. We found evidence that cells capable of forming endospores are not necessarily dormant and that members of the class Bacilli were more likely to form endospores in response to long-term stressors associated with permafrost environmental conditions than members of the Clostridia, which were more likely to persist as vegetative cells in our older samples. We also found that removing exogenous "relic" DNA preserved within permafrost did not significantly alter microbial community composition. These results link the live, dead, and dormant microbial communities to physicochemical characteristics and provide insights into the survival of microbial communities in ancient permafrost.IMPORTANCE Permafrost soils store more than half of Earth's soil carbon despite covering ∼15% of the land area (C. Tarnocai et al., Global Biogeochem Cycles 23:GB2023, 2009, https://doi.org/10.1029/2008GB003327). This permafrost carbon is rapidly degraded following a thaw (E. A. G. Schuur et al., Nature 520:171-179, 2015, https://doi.org/10.1038/nature14338). Understanding microbial communities in permafrost will contribute to the knowledge base necessary to understand the rates and forms of permafrost C and N cycling postthaw. Permafrost is also an analog for frozen extraterrestrial environments, and evidence of viable organisms in ancient permafrost is of interest to those searching for potential life on distant worlds. If we can identify strategies microbial communities utilize to survive in permafrost, it may yield insights into how life (if it exists) survives in frozen environments outside of Earth. Our work is significant because it contributes to an understanding of how microbial life adapts and survives in the extreme environmental conditions in permafrost terrains.


Assuntos
Microbiota/fisiologia , Pergelissolo/microbiologia , Microbiologia do Solo , Solo/química , Actinobacteria/genética , Actinobacteria/isolamento & purificação , Alaska , Bacillaceae/genética , Bacillaceae/isolamento & purificação , Carbono/metabolismo , Clostridiaceae/genética , Clostridiaceae/isolamento & purificação , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , Ecologia , Congelamento , Metagenômica , Microbiota/genética , Filogenia , RNA Ribossômico 16S/genética , Esporos Bacterianos/fisiologia , Temperatura
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