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A group of Gram-negative plant-associated diazotrophic bacteria belonging to the genus Nitrospirillum was investigated, including both previously characterized and newly isolated strains from diverse regions and biomes, predominantly in Brazil. Phylogenetic analysis of 16S rRNA and recA genes revealed the formation of a distinct clade consisting of thirteen strains, separate from the formally recognized species N. amazonense (the closest species) and N. iridis. Comprehensive taxonomic analyses using the whole genomes of four strains (BR 11140T = AM 18T = Y-2T = DSM 2788T = ATCC 35120T, BR 11142T = AM 14T = Y-1T = DSM 2787T = ATCC 35119T, BR 11145 = CBAmC, and BR 12005) supported the division of these strains into two species: N. amazonense (BR 11142 T and BR 12005) and a newly proposed species (BR 11140 T and BR 11145), distinct from N. iridis. The phylogenomic analysis further confirmed the presence of the new Nitrospirillum species. Additionally, MALDI-TOF MS analysis of whole-cell mass spectra provided further evidence for the differentiation of the proposed Nitrospirillum species, separate from N. amazonense. Analysis of chemotaxonomy markers (i.e., genes involved in fatty acid synthesis, metabolism and elongation, phospholipid synthesis, and quinone synthesis) revealed that the new species highlights high similarity and evolutionary convergence with other Nitrospirillum species. This new species exhibited nitrogen fixation ability in vitro, it has similar NifHDK protein phylogeny position with the closest species, lacked denitrification capability, but possessed the nosZ gene, enabling N2O reduction, distinguishing it from the closest species. Despite being isolated from diverse geographic regions, soil types, and ecological niches, no significant phenotypic or physiological differences were observed between the proposed new species and N. amazonense. Based on these findings, a new species, Nitrospirillum viridazoti sp. nov., was classified, with the strain BR 11140T (DSM 2788T, ATCC 35120T) designated as the type strain.
Assuntos
Nitrogênio , Poaceae , Filogenia , RNA Ribossômico 16S/genética , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
As agricultural practices become more sustainable, adopting more sustainable practices will become even more relevant. Searching for alternatives to chemical compounds has been the focus of numerous studies, and bacteriocins are tools with intrinsic biotechnological potential for controlling plant diseases. We continued to explore the biotechnological activity of the bacteriocin Gluconacin from Gluconacetobacter diazotrophicus, PAL5 strain, by investigating this protein's antagonism against important tomato phytopathogens and demonstrating its effectiveness in reducing bacterial spots caused by Xanthomonas perforans. In addition to this pathogen, the bacteriocin Gluconacin demonstrated bactericidal activity in vitro against Ralstonia solanacearum and Pseudomonas syringae pv. tomato, agents that cause bacterial wilt and bacterial spots, respectively. Bacterial spot control tests showed that Gluconacin reduced disease severity by more than 66%, highlighting the biotechnological value of this peptide in ecologically correct formulations.
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Gluconacetobacter diazotrophicus has been the focus of several studies aiming to understand the mechanisms behind this endophytic diazotrophic bacterium. The present study is the first global analysis of the early transcriptional response of exponentially growing G. diazotrophicus to iron, an essential cofactor for many enzymes involved in various metabolic pathways. RNA-seq, targeted gene mutagenesis and computational motif discovery tools were used to define the G. diazotrophicusfur regulon. The data analysis showed that genes encoding functions related to iron homeostasis were significantly upregulated in response to iron limitations. Certain genes involved in secondary metabolism were overexpressed under iron-limited conditions. In contrast, it was observed that the expression of genes involved in Fe-S cluster biosynthesis, flagellar biosynthesis and type IV secretion systems were downregulated in an iron-depleted culture medium. Our results support a model that controls transcription in G. diazotrophicus by fur function. The G. diazotrophicusfur protein was able to complement an E. colifur mutant. These results provide new insights into the effects of iron on the metabolism of G. diazotrophicus, as well as demonstrate the essentiality of this micronutrient for the main characteristics of plant growth promotion by G. diazotrophicus.
Assuntos
Gluconacetobacter , Ferro , Proteínas de Bactérias/metabolismo , Meios de Cultura/farmacologia , Ferro/metabolismo , TranscriptomaRESUMO
Sugarcane is an economically important crop that is used for the production of fuel ethanol. Diazotrophic bacteria have been isolated from sugarcane tissues, without causing visible plant anatomical changes or disease symptoms. These bacteria can be beneficial to the plant by promoting root growth and an increase in plant yield. Different rates of Biological Nitrogen Fixation (BNF) were observed in different genotypes. The aim of this work was to conduct a comprehensive molecular and physiological analysis of two model genotypes for contrasting BNF efficiency in order to unravel plant genes that are differentially regulated during a natural association with diazotrophic bacteria. A next-generation sequencing of RNA samples from the genotypes SP70-1143 (high-BNF) and Chunee (low-BNF) was performed. A differential transcriptome analysis showed that several pathways were differentially regulated among the two BNF-contrasting genotypes, including nitrogen metabolism, hormone regulation and bacteria recognition. Physiological analyses, such as nitrogenase and GS activity quantification, bacterial colonization, auxin response and root architecture evaluation, supported the transcriptome expression analyses. The differences observed between the genotypes may explain, at least in part, the differences in BNF contributions. Some of the identified genes might be involved in key regulatory processes for a beneficial association and could be further used as tools for obtaining more efficient BNF genotypes.
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ABSTRACT This work aimed to isolate and characterize plant growth promoting rhizobacteria (PGPR) from 10 Paspalum genotypes and evaluate the effect of their inoculation on P. regnellii, P. atratum, and P. malacophyllum genotypes. The bacterial population ranged from undetectable to 107 bacterial cells per gram of fresh matter in the Paspalum genotypes. Initially, we isolated 164 bacteria from rhizospheric soil and roots of the Paspalum genotypes using media N-free LG agar plate, semi-solid NFb, and LGI. The isolates were characterized genetically and physiologically. The sequencing of 16S rRNA showed the presence of many genera, and some are new in association with Paspalum. The most common was Bacillus followed by Rhizobium, Paraburkholderia, Enterobacter, Cupriavidus, Pseudomonas, Dyadobacter and Acinetobacter. Thirty-eight per cent of isolates produced siderophores, 25 % produced solubilized phosphate, and only 9 % produced indolic compounds. Three greenhouse experiments were performed in randomized blocks with six replicates using representative bacterial strains isolated from P. regnellii, P. malacophyllum and P. atratum cv. Pojuca. We also included strain Sp245 (Azospirillum baldaniorum), uninoculated control, and nitrogen control (150 kg N ha1). There was an increase of up to 53 % in shoot dry matter in P. regnellii inoculated with strain Sp245 and the shoots accumulated more N. In contrast, only small effects were observed for the other Paspalum genotypes inoculated with PGPR from the host genotypes. This study shows a high diversity of diazotrophic rhizosphere bacteria and suggests no strain specificity between the bacterial isolates and the Paspalum genotypes.
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This work aimed to isolate and characterize plant growth promoting rhizobacteria (PGPR) from 10 Paspalum genotypes and evaluate the effect of their inoculation on P. regnellii, P. atratum, and P. malacophyllum genotypes. The bacterial population ranged from undetectable to 107 bacterial cells per gram of fresh matter in the Paspalum genotypes. Initially, we isolated 164 bacteria from rhizospheric soil and roots of the Paspalum genotypes using media N-free LG agar plate, semi-solid NFb, and LGI. The isolates were characterized genetically and physiologically. The sequencing of 16S rRNA showed the presence of many genera, and some are new in association with Paspalum. The most common was Bacillus followed by Rhizobium, Paraburkholderia, Enterobacter, Cupriavidus, Pseudomonas, Dyadobacter and Acinetobacter. Thirty-eight per cent of isolates produced siderophores, 25 % produced solubilized phosphate, and only 9 % produced indolic compounds. Three greenhouse experiments were performed in randomized blocks with six replicates using representative bacterial strains isolated from P. regnellii, P. malacophyllum and P. atratum cv. Pojuca. We also included strain Sp245 (Azospirillum baldaniorum), uninoculated control, and nitrogen control (150 kg N ha1). There was an increase of up to 53 % in shoot dry matter in P. regnellii inoculated with strain Sp245 and the shoots accumulated more N. In contrast, only small effects were observed for the other Paspalum genotypes inoculated with PGPR from the host genotypes. This study shows a high diversity of diazotrophic rhizosphere bacteria and suggests no strain specificity between the bacterial isolates and the Paspalum genotypes.
Assuntos
Biologia do Solo/análise , Genes Bacterianos , Paspalum/crescimento & desenvolvimento , Paspalum/genéticaRESUMO
This work aimed to isolate and characterize plant growth promoting rhizobacteria (PGPR) from 10 Paspalum genotypes and evaluate the effect of their inoculation on P. regnellii, P. atratum, and P. malacophyllum genotypes. The bacterial population ranged from undetectable to 107 bacterial cells per gram of fresh matter in the Paspalum genotypes. Initially, we isolated 164 bacteria from rhizospheric soil and roots of the Paspalum genotypes using media N-free LG agar plate, semi-solid NFb, and LGI. The isolates were characterized genetically and physiologically. The sequencing of 16S rRNA showed the presence of many genera, and some are new in association with Paspalum. The most common was Bacillus followed by Rhizobium, Paraburkholderia, Enterobacter, Cupriavidus, Pseudomonas, Dyadobacter and Acinetobacter. Thirty-eight per cent of isolates produced siderophores, 25 % produced solubilized phosphate, and only 9 % produced indolic compounds. Three greenhouse experiments were performed in randomized blocks with six replicates using representative bacterial strains isolated from P. regnellii, P. malacophyllum and P. atratum cv. Pojuca. We also included strain Sp245 (Azospirillum baldaniorum), uninoculated control, and nitrogen control (150 kg N ha−1). There was an increase of up to 53 % in shoot dry matter in P. regnellii inoculated with strain Sp245 and the shoots accumulated more N. In contrast, only small effects were observed for the other Paspalum genotypes inoculated with PGPR from the host genotypes. This study shows a high diversity of diazotrophic rhizosphere bacteria and suggests no strain specificity between the bacterial isolates and the Paspalum genotypes.
Assuntos
Pastagens , Paspalum , Bactérias Fixadoras de Nitrogênio/isolamento & purificação , RizosferaRESUMO
Plant growth-promoting bacteria (PGPB) are bacteria that have mechanisms that facilitate plant growth in stress conditions such as drought. The objective of this study was to characterize bacterial strains isolated from bromeliads roots in ironstone outcrops (Urucum Residual Plateau, Mato Grosso do Sul, Brazil) for plant growth-promoting under drought conditions. Firstly, we screened isolates with the presence of 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity. Then, all isolates were tested for tolerance to drought, exopolysaccharides (EPS) production, indole-3-acetic acid (IAA)-producing abilities, phosphate and zinc solubilization, production of catalase and hydrolytic enzymes (amylase, cellulase, and protease). Germination assay and a pot experiment with maize plants submitted to well-watered and drought conditions were performed with the strains most promising (VBN11 and VBE23). Briefly, Bacillus cereus VBE23 showed in vitro higher ACC deaminase activity (3.83 and 2.52 µmol α-KB mg-1 h-1 in non-drought and drought conditions, respectively), tolerance to drought, EPS production and other mechanisms of plant growth promotion: solubilization of phosphate and zinc, ammonia production, catalase activity and production of hydrolytic enzymes (amylase, cellulase, and protease). Inoculation of strain VBE23 in maize seeds submitted to drought conditions showed higher germination concerning uninoculated seeds and inoculated with VBN11. Also, the results indicated that the isolate VBE23 provided higher values of fresh and dry biomass compared to the control of uninoculated treatment and inoculated with VBN11 under drought conditions. This is the first report on the PGPB from ironstone outcrops of Urucum Residual Plateau, Mato Grosso do Sul, Brazil. Thus, this bacterial isolate could be used as a strategy for the facilitation of plant growth in drought environments.
Assuntos
Carbono-Carbono Liases , Secas , Bactérias/genética , Brasil , Raízes de Plantas , Microbiologia do SoloRESUMO
Bacteria of the genus Bacillus can colonize endophytically and benefit several crops including the control of some pest orders. In view of the benefits provided by these microorganisms and in order to find out an efficient biotechnological control for the giant borer, our interest in studying the microorganisms in symbiosis with sugarcane and the giant borer has arisen, since there is no efficient chemical or biological control method for this pest. Therefore, endophytic Bacillus strains were isolated from three sugarcane niches (apoplast fluid, central internode cylinder and roots) and also from the giant borer larvae living inside sugarcane varieties grown in the Northeast region of Brazil. The taxonomical characterization (16S rRNA) of 157 Gram-positive isolates showed that 138 strains belonged to the Bacillus genus. The most representative species were phylogenetically closely related to B. megaterium (11.5%) followed by B. safensis (10.8%), B. cereus (8.9%), B. oleronius (8.9%), B. amyloliquefaciens (7.0%), and B. pacificus (6.4%). BOX-PCR analyses showed very distinct band pattern profiles suggesting a great diversity of Bacillus species within the sugarcane niches and the digestive tract, while the B. cereus group remained very closely clustered in the dendrogram. According to XRE biomarker analysis, eleven strains (FORCN005, 007, 008, 011, 012, 014, 067, 076, 092, 093, and 135) correspond to B. thuringiensis species. Additional studies using conserved genes (glp, gmk, pta, and tpi) indicated that most of these strains were phylogenetically closely related to B. thuringiensis and may be considered different subspecies. In conclusion, this study suggests that the culturable Bacillus species are greatly diversified within the plant niches and showed Bacillus species in the digestive tract of the giant borer for the first time. These results open new perspectives to understand the role and functions played by these microorganisms in symbiosis with this pest and also the possibility of developing an efficient biological control method for the giant borer using strains identified as the B. thuringiensis species.
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Bradyrhizobium spp. are well known to mediate biological nitrogen fixation (BNF) as microsymbionts inhabiting nodules on leguminous plants. However, they may also contribute to plant growth via free-living N2 fixation (FLNF) in association with non-legumes. Notably, several Bradyrhizobium strains from sugarcane roots display FLNF activity. Among them, Bradyrhizobium sacchari is a legume symbiotic species, whereas strains AG48 and M12 are non-symbiotic. In the present study, a phylogenomic approach was applied to study peculiarities of these and other Bradyrhizobium strains with respect to N fixation (nif) gene content in order to reveal genetic features that enable FNLF in Bradyrhizobium spp. All FLNF strains carry an ancestral 'non-symbiotic' nif-gene cluster (NSC). B. sacchari also contains a second 'symbiotic' nif-gene cluster (SC), a characteristic observed in only three of 156 evaluated genomes. B. sacchari stood out and presented a high level of sequence divergence between individual nif-gene homologues and we discuss scenarios for the evolutionary origin of these clusters. The transcript level of NSC nifH gene increased during FLNF, when compared to symbiotic conditions. The data suggest that sugarcane roots harbor diverse Bradyrhizobium spp. that are genetically adapted to a dynamic environment where leguminous and non-leguminous host plants are alternately available.
Assuntos
Bradyrhizobium , Fabaceae , Saccharum , Bradyrhizobium/genética , DNA Bacteriano/genética , Família Multigênica , Fixação de Nitrogênio/genética , Filogenia , RNA Ribossômico 16S/genética , Nódulos Radiculares de Plantas , Simbiose/genéticaRESUMO
BACKGROUND: Inoculation with Gluconacetobacter diazotrophicus has shown to influence root development in red rice plants, and more recently, the induced systemic tolerance (IST) response to drought was also demonstrated. The goal of this study was to evaluate the inoculation effect of G. diazotrophicus strain Pal5 on the amelioration of drought stress and root development in red rice (Oryza sativa L.). METHODS: The experimental treatments consist of red rice plants inoculated with and without strain Pal5 in presence and absence of water restriction. Physiological, biochemical, and molecular analyses of plant roots were carried out, along with measurements of growth and biochemical components. RESULTS: The plants showed a positive response to the bacterial inoculation, with root growth promotion and induction of tolerance to drought. An increase in the root area and higher levels of osmoprotectant solutes were observed in roots. Bacterial inoculation increased the drought tolerance and positively regulated certain root development genes against the water deficit in plants. CONCLUSION: G. diazotrophicus Pal5 strain inoculation favored red rice plants by promoting various root growth and developmental mechanisms against drought stress, enabling root development and improving biochemical composition.
Assuntos
Gluconacetobacter/fisiologia , Oryza/microbiologia , Raízes de Plantas/microbiologia , Secas , Oryza/crescimento & desenvolvimento , Raízes de Plantas/crescimento & desenvolvimento , Estresse Fisiológico , Simbiose , Água/metabolismoRESUMO
Nine bacterial strains were previously isolated in association with pinewood nematode (PWN) from wilted pine trees. They proved to be nematicidal in vitro, and one of the highest activities, with potential to control PWN, was showed by Serratia sp. M24T3. Its ecology in association with plants remains unclear. This study aimed to evaluate the ability of strain M24T3 to colonize the internal tissues of the model plant Arabidopsis thaliana using confocal microscopy. Plant growth-promoting bacteria (PGPB) functional traits were tested and retrieved in the genome of strain M24T3. In greenhouse conditions, the bacterial effects of all nematicidal strains were also evaluated, co-inoculated or not with Bradyrhizobium sp. 3267, on Vigna unguiculata fitness. Inoculation of strain M24T3 increased the number of A. thaliana lateral roots and the confocal analysis confirmed effective bacterial colonization in the plant. Strain M24T3 showed cellulolytic activity, siderophores production, phosphate and zinc solubilization ability, and indole acetic acid production independent of supplementation with L-tryptophan. In the genome of strain M24T3, genes involved in the interaction with the plants such as 1-aminocyclopropane-1-carboxylate (ACC) deaminase, chitinolytic activity, and quorum sensing were also detected. The genomic organization showed ACC deaminase and its leucine-responsive transcriptional regulator, and the activity of ACC deaminase was 594.6 nmol α-ketobutyrate µg protein-1 µl-1. Strain M24T3 in co-inoculation with Bradyrhizobium sp. 3267 promoted the growth of V. unguiculata. In conclusion, this study demonstrated the ability of strain M24T3 to colonize other plants besides pine trees as an endophyte and displays PGPB traits that probably increased plant tolerance to stresses.
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Arabidopsis/microbiologia , Nematoides/microbiologia , Serratia/fisiologia , Animais , Antibiose , Arabidopsis/crescimento & desenvolvimento , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Carbono-Carbono Liases/genética , Carbono-Carbono Liases/metabolismo , Pinus/parasitologia , Doenças das Plantas/parasitologia , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/microbiologia , Percepção de Quorum , Serratia/enzimologia , Serratia/genética , Serratia/isolamento & purificação , Vigna/crescimento & desenvolvimento , Vigna/microbiologiaRESUMO
The stalk apoplast fluid of sugarcane contains different sugars, organic acids and amino acids that may supply the demand for carbohydrates by endophytic bacteria including diazotrophs P. tropica (syn. B. tropica) strain Ppe8, isolated from sugarcane, is part of the bacterial consortium recommended as inoculant to sugarcane. However, little information has been accumulated regarding this plant-bacterium interaction considering that it colonizes internal sugarcane tissues. Here, we made use of the RNA-Seq transcriptomic analysis to study the influence of sugarcane stalk apoplast fluid on Ppe8 gene expression. The bacterium was grown in JMV liquid medium (100 ml), divided equally and then supplemented with 50 ml of fresh JMV medium or 50 ml of apoplast fluid extracted from sugarcane variety RB867515. Total RNA was extracted 2 hours later, the rRNAs were depleted and mRNAs used to construct libraries to sequence the fragments using Ion Torrent technology. The mapping and statistical analysis were carried out with CLC Genomics Workbench software. The RNA-seq data was validated by RT-qPCR using the reference genes fliP1, paaF, and groL. The data analysis showed that 544 genes were repressed and 153 genes were induced in the presence of apoplast fluid. Genes that induce plant defense responses, genes related to chemotaxis and movements were repressed in the presence of apoplast fluid, indicating that strain Ppe8 recognizes the apoplast fluid as a plant component. The expression of genes involved in bacterial metabolism was regulated (up and down), suggesting that the metabolism of strain Ppe8 is modulated by the apoplast fluid. These results suggest that Ppe8 alters its gene expression pattern in the presence of apoplast fluid mainly in order to use compounds present in the fluid as well as to avoid the induction of plant defense mechanisms. This is a pioneer study showing the role played by the sugarcane apoplast fluid on the global modulation of genes in P. tropica strain Ppe8.
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Burkholderiaceae/genética , Burkholderiaceae/metabolismo , Endófitos/genética , Endófitos/metabolismo , Saccharum/metabolismo , Saccharum/microbiologia , Aminoácidos/metabolismo , Transporte Biológico Ativo , Metabolismo dos Carboidratos , Movimento Celular/genética , Parede Celular/genética , Quimiotaxia/genética , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Genes Bacterianos , Estruturas Vegetais/metabolismo , Estruturas Vegetais/microbiologia , Transdução de SinaisRESUMO
Paraburkholderia tropica (syn Burkholderia tropica) are nitrogen-fixing bacteria commonly found in sugarcane. The Paraburkholderia tropica strain Ppe8 is part of the sugarcane inoculant consortium that has a beneficial effect on yield. Here, we report a draft genome sequence of this strain elucidating the mechanisms involved in its interaction mainly with Poaceae. A genome size of approximately 8.75 Mb containing 7844 protein coding genes distributed in 526 subsystems was de novo assembled with ABySS and annotated by RAST. Genes related to the nitrogen fixation process, the secretion systems (I, II, III, IV, and VI), and related to a variety of metabolic traits, such as metabolism of carbohydrates, amino acids, vitamins, and proteins, were detected, suggesting a broad metabolic capacity and possible adaptation to plant association.(AU)
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Abstract Paraburkholderia tropica (syn Burkholderia tropica) are nitrogen-fixing bacteria commonly found in sugarcane. The Paraburkholderia tropica strain Ppe8 is part of the sugarcane inoculant consortium that has a beneficial effect on yield. Here, we report a draft genome sequence of this strain elucidating the mechanisms involved in its interaction mainly with Poaceae. A genome size of approximately 8.75 Mb containing 7844 protein coding genes distributed in 526 subsystems was de novo assembled with ABySS and annotated by RAST. Genes related to the nitrogen fixation process, the secretion systems (I, II, III, IV, and VI), and related to a variety of metabolic traits, such as metabolism of carbohydrates, amino acids, vitamins, and proteins, were detected, suggesting a broad metabolic capacity and possible adaptation to plant association.
Assuntos
Genoma Bacteriano , Burkholderiaceae/genética , Endófitos/genética , Proteínas de Bactérias/genética , Análise de Sequência de DNA , Biologia Computacional , Saccharum/microbiologia , Burkholderiaceae/isolamento & purificação , Redes e Vias Metabólicas/genética , Anotação de Sequência Molecular , Endófitos/isolamento & purificaçãoRESUMO
Nitrospirillum amazonense is a nitrogen-fixing bacterium that shows potential to promote plant growth when inoculated into sugarcane and rice plants. This microorganism has been the subject of biochemical and genetic characterization to elucidate important functions related to host plant interaction and growth promotion, including the determination of draft genome sequences of two strains, Y2 and CBAmC, the second of which is the aim of the present study. CBAmC has been isolated from sugarcane (Saccharum spp.), and is currently used in a sugarcane consortium inoculant with four other nitrogen-fixing bacterial strains. The present paper describes a significant improvement in the genome sequence and assembly for the N. amazonense strain CBAmC, and determination for the first time of a complete genome sequence for this bacterial species, using PacBio technology. The analysis of the genomic data obtained allowed the discovery of genes coding for metabolic pathways and cellular structures that may be determinant for the success of the bacterial establishment and colonization into the host sugarcane plant, besides conferring important characteristics to the inoculant. These include genes for the use of sucrose and N-glycans, biosynthesis of autoinducer molecules, siderophore production and acquisition, auxin and polyamine biosynthesis, flagellum, σ-fimbriae, a variety of secretion systems, and a complete denitrification system. Concerning genes for nitrogenase and auxiliary proteins, it was possible to corroborate literature data that in N. amazonense these probably had originated from horizontal gene transfer, from bacteria of the Rhizobiales order. The complete genomic sequence of the CBAmC strain of N. amazonense revealed that the bacterium harbors four replicons, including three chromosomes and one chromid, a profile that coincides with that of other two strains, according to literature data, suggesting that as a replicon pattern for the species. Finally, results of phylogenomic analyses in this work support the recent reclassification of the species, separating it from the Azospirillum genus. More importantly, results of the present work shall guide subsequent studies on strain CBAmC as well as the development of a sugarcane inoculant.
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Genoma Bacteriano , Fixação de Nitrogênio , Caules de Planta/microbiologia , Rhodospirillaceae/genética , Saccharum/microbiologia , Rhodospirillaceae/isolamento & purificaçãoRESUMO
Paraburkholderia tropica (syn Burkholderia tropica) are nitrogen-fixing bacteria commonly found in sugarcane. The Paraburkholderia tropica strain Ppe8 is part of the sugarcane inoculant consortium that has a beneficial effect on yield. Here, we report a draft genome sequence of this strain elucidating the mechanisms involved in its interaction mainly with Poaceae. A genome size of approximately 8.75Mb containing 7844 protein coding genes distributed in 526 subsystems was de novo assembled with ABySS and annotated by RAST. Genes related to the nitrogen fixation process, the secretion systems (I, II, III, IV, and VI), and related to a variety of metabolic traits, such as metabolism of carbohydrates, amino acids, vitamins, and proteins, were detected, suggesting a broad metabolic capacity and possible adaptation to plant association.
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Burkholderiaceae/genética , Endófitos/genética , Genoma Bacteriano , Proteínas de Bactérias/genética , Burkholderiaceae/isolamento & purificação , Biologia Computacional , Endófitos/isolamento & purificação , Redes e Vias Metabólicas/genética , Anotação de Sequência Molecular , Saccharum/microbiologia , Análise de Sequência de DNARESUMO
Abstract Paraburkholderia tropica (syn Burkholderia tropica) are nitrogen-fixing bacteria commonly found in sugarcane. The Paraburkholderia tropica strain Ppe8 is part of the sugarcane inoculant consortium that has a beneficial effect on yield. Here, we report a draft genome sequence of this strain elucidating the mechanisms involved in its interaction mainly with Poaceae. A genome size of approximately 8.75 Mb containing 7844 protein coding genes distributed in 526 subsystems was de novo assembled with ABySS and annotated by RAST. Genes related to the nitrogen fixation process, the secretion systems (I, II, III, IV, and VI), and related to a variety of metabolic traits, such as metabolism of carbohydrates, amino acids, vitamins, and proteins, were detected, suggesting a broad metabolic capacity and possible adaptation to plant association.
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Quantitative reverse transcription PCR (RT-qPCR) is an important tool for evaluating gene expression. However, this technique requires that specific internal normalizing genes be identified for different experimental conditions. To date, no internal normalizing genes are available for validation of data analyses for Herbaspirillum rubrisubalbicans strain HCC103, an endophyte that is part of the sugarcane consortium inoculant. This work seeks to identify and evaluate suitable reference genes for gene expression studies in HCC103 grown until middle log phase in sugarcane juice obtained from four sugarcane varieties or media with three different carbon sources. The mRNA levels of five candidate genes (rpoA, gyrA, dnaG, recA and gmK) and seven target genes involved in carbon metabolism (acnA, fbp, galE, suhB, wcaA, ORF_0127.0101 and _0127.0123) were quantified by RT-qPCR. Analysis of expression stability of these genes was carried out using geNorm and Normfinder software. The results indicated that the HCC103 dnaG and gyrA genes are the most stable and showed adequate relative expression level changes among the different sugarcane juices. The highest expression level was seen for ORF_0127.0101, which encodes a sugar transporter, in juice from sugarcane variety RB867515 and glucose as the carbon source. The suhB gene, encoding SuhB inositol monophosphatase, had a higher relative expression level on 0.5% glucose, 100% sugarcane juice from variety RB867515 and 0.5% aconitate. Together the results suggest that dnaG and gyrA genes are suitable as reference genes for RT-qPCR analysis of strain HCC103 and that juice from different sugarcane varieties modulates the expression of key genes involved in carbon metabolism.
Assuntos
Carbono/metabolismo , Sucos de Frutas e Vegetais , Genes Bacterianos , Herbaspirillum/efeitos dos fármacos , Herbaspirillum/fisiologia , Saccharum/química , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Estabilidade de RNA , Reação em Cadeia da Polimerase em Tempo Real , TranscriptomaRESUMO
Members of the genus Bradyrhizobium are well-known as nitrogen-fixing microsymbionts of a wide variety of leguminous species, but they have also been found in different environments, notably as endophytes in non-legumes such as sugarcane. This study presents a detailed polyphasic characterization of four Bradyrhizobium strains (type strain BR 10280T), previously isolated from roots of sugarcane in Brazil. 16S rRNA sequence analysis, multilocus sequence analysis (MLSA) and analysis of the 16S-23S rRNA internal transcribed spacer showed that these strains form a novel clade close to, but different from B. huanghuaihaiense strain CCBAU 23303T. Average nucleotide identity (ANI) analyses confirmed that BR 10280T represents a novel species. Phylogenetic analysis based on nodC gene sequences also placed the strains close to CCBAU 23303T, but different from this latter strain, the sugarcane strains did not nodulate soybean, although they effectively nodulated Vigna unguiculata, Cajanus cajan and Macroptilium atropurpureum. Physiological traits are in agreement with the placement of the strains in the genus Bradyrhizobium as a novel species for which the name Bradyrhizobium sacchari sp. nov. is proposed.