RESUMEN
Microbial non-phosphorylative oxidative pathways present promising potential in the biosynthesis of platform chemicals from the hemicellulosic fraction of lignocellulose. An L-arabinonate dehydratase from Rhizobium leguminosarum bv. trifolii catalyzes the rate-limiting step in the non-phosphorylative oxidative pathways, that is, converts sugar acid to 2-dehydro-3-deoxy sugar acid. We have shown earlier that the enzyme forms a dimer of dimers, in which the C-terminal histidine residue from one monomer participates in the formation of the active site of an adjacent monomer. The histidine appears to be conserved across the sequences of sugar acid dehydratases. To study the role of the C-terminus, five variants (H579A, H579F, H579L, H579Q, and H579W) were produced. All variants showed decreased activity for the tested sugar acid substrates, except the variant H579L on D-fuconate, which showed about 20% increase in activity. The reaction kinetic data showed that the substrate preference was slightly modified in H579L compared to the wild-type enzyme, demonstrating that the alternation of the substrate preference of sugar acid dehydratases is possible. In addition, a crystal structure of H579L was determined at 2.4 Šwith a product analog 2-oxobutyrate. This is the first enzyme-ligand complex structure from an IlvD/EDD superfamily enzyme. The binding of 2-oxobutyrate suggests how the substrate would bind into the active site in the orientation, which could lead to the dehydration reaction. KEY POINTS: ⢠Mutation of the last histidine at the C-terminus changed the catalytic activity of L-arabinonate dehydratase from R. leguminosarum bv. trifolii against various C5/C6 sugar acids. ⢠The variant H579L of L-arabinonate dehydratase showed an alteration of substrate preferences compared with the wild type. ⢠The first enzyme-ligand complex crystal structure of an IlvD/EDD superfamily enzyme was solved.
Asunto(s)
Hidroliasas , Rhizobium leguminosarum , Hidroliasas/metabolismo , Hidroliasas/genética , Hidroliasas/química , Especificidad por Sustrato , Rhizobium leguminosarum/enzimología , Rhizobium leguminosarum/genética , Cinética , Dominio Catalítico , Azúcares Ácidos/metabolismo , Histidina/metabolismo , Histidina/química , Histidina/genética , Multimerización de Proteína , Modelos Moleculares , Proteínas Bacterianas/genética , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismoRESUMEN
The species Rhizobium indigoferae and Sinorhizobium kummerowiae were isolated from legume nodules and the 16S rRNA sequences of their respective type strains, CCBAU 71042T and CCBAU 71714T, were highly divergent from those of the other species of the genera Rhizobium and Sinorhizobium, respectively. However, the 16S rRNA gene sequences obtained for strains CCBAU 71042T and CCBAU 71714T several years after description, were different from the original ones, showing 100â% similarity to the type strains of Rhizobium leguminosarum and Sinorhizobium meliloti, respectively. Phylogenetic analyses of two housekeeping genes, recA and atpD, confirmed the high phylogenetic closeness of strains CCBAU 71042T and CCBAU 71714T to the respective type strains of R. leguminosarum and S. meliloti. In the present work, we compared the genomes of the type strains of R. indigoferae and S. kummerowiae available in several culture collections with those of the respective type strains of R. leguminosarum and S. meliloti, some of them obtained in this study. The calculated average nucleotide identity-blast and digital DNA-DNA hybridization values in both cases were higher than those recommended for species differentiation, supporting the proposal for the reclassification of the type strains of R. indigoferae and S. kummerowiae into the species R. leguminosarum and S. meliloti, respectively.
Asunto(s)
Técnicas de Tipificación Bacteriana , ADN Bacteriano , Filogenia , ARN Ribosómico 16S , Rhizobium leguminosarum , Análisis de Secuencia de ADN , Sinorhizobium meliloti , ARN Ribosómico 16S/genética , ADN Bacteriano/genética , Sinorhizobium meliloti/genética , Sinorhizobium meliloti/clasificación , Rhizobium leguminosarum/genética , Rhizobium leguminosarum/clasificación , Genoma Bacteriano , Rhizobium/clasificación , Rhizobium/genética , Rhizobium/aislamiento & purificación , Nódulos de las Raíces de las Plantas/microbiología , Genes Esenciales , Genes Bacterianos , Hibridación de Ácido NucleicoRESUMEN
BACKGROUND: Mercury (Hg) is highly toxic and has the potential to cause severe health problems for humans and foraging animals when transported into edible plant parts. Soil rhizobia that form symbiosis with legumes may possess mechanisms to prevent heavy metal translocation from roots to shoots in plants by exporting metals from nodules or compartmentalizing metal ions inside nodules. Horizontal gene transfer has potential to confer immediate de novo adaptations to stress. We used comparative genomics of high quality de novo assemblies to identify structural differences in the genomes of nitrogen-fixing rhizobia that were isolated from a mercury (Hg) mine site that show high variation in their tolerance to Hg. RESULTS: Our analyses identified multiple structurally conserved merA homologs in the genomes of Sinorhizobium medicae and Rhizobium leguminosarum but only the strains that possessed a Mer operon exhibited 10-fold increased tolerance to Hg. RNAseq analysis revealed nearly all genes in the Mer operon were significantly up-regulated in response to Hg stress in free-living conditions and in nodules. In both free-living and nodule environments, we found the Hg-tolerant strains with a Mer operon exhibited the fewest number of differentially expressed genes (DEGs) in the genome, indicating a rapid and efficient detoxification of Hg from the cells that reduced general stress responses to the Hg-treatment. Expression changes in S. medicae while in bacteroids showed that both rhizobia strain and host-plant tolerance affected the number of DEGs. Aside from Mer operon genes, nif genes which are involved in nitrogenase activity in S. medicae showed significant up-regulation in the most Hg-tolerant strain while inside the most Hg-accumulating host-plant. Transfer of a plasmid containing the Mer operon from the most tolerant strain to low-tolerant strains resulted in an immediate increase in Hg tolerance, indicating that the Mer operon is able to confer hyper tolerance to Hg. CONCLUSIONS: Mer operons have not been previously reported in nitrogen-fixing rhizobia. This study demonstrates a pivotal role of the Mer operon in effective mercury detoxification and hypertolerance in nitrogen-fixing rhizobia. This finding has major implications not only for soil bioremediation, but also host plants growing in mercury contaminated soils.
Asunto(s)
Transferencia de Gen Horizontal , Mercurio , Operón , Simbiosis , Transcriptoma , Mercurio/metabolismo , Mercurio/toxicidad , Bacterias Fijadoras de Nitrógeno/genética , Bacterias Fijadoras de Nitrógeno/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Regulación Bacteriana de la Expresión Génica , Fijación del Nitrógeno , Rhizobium leguminosarum/genética , Rhizobium leguminosarum/metabolismo , Microbiología del SueloRESUMEN
Rhizobium leguminosarum aspartate aminotransferase (AatA) mutants show drastically reduced symbiotic nitrogen fixation in legume nodules. Whilst AatA reversibly transaminates the two major amino-donor compounds aspartate and glutamate, the reason for the lack of N2 fixation in the mutant has remained unclear. During our investigations into the role of AatA, we found that it catalyses an additional transamination reaction between aspartate and pyruvate, forming alanine. This secondary reaction runs at around 60â% of the canonical aspartate transaminase reaction rate and connects alanine biosynthesis to glutamate via aspartate. This may explain the lack of any glutamate-pyruvate transaminase activity in R. leguminosarum, which is common in eukaryotic and many prokaryotic genomes. However, the aspartate-to-pyruvate transaminase reaction is not needed for N2 fixation in legume nodules. Consequently, we show that aspartate degradation is required for N2 fixation, rather than biosynthetic transamination to form an amino acid. Hence, the enzyme aspartase, which catalyses the breakdown of aspartate to fumarate and ammonia, suppressed an AatA mutant and restored N2 fixation in pea nodules.
Asunto(s)
Aspartato Aminotransferasas , Ácido Aspártico , Fijación del Nitrógeno , Pisum sativum , Rhizobium leguminosarum , Nódulos de las Raíces de las Plantas , Rhizobium leguminosarum/genética , Rhizobium leguminosarum/metabolismo , Rhizobium leguminosarum/enzimología , Ácido Aspártico/metabolismo , Pisum sativum/microbiología , Nódulos de las Raíces de las Plantas/microbiología , Aspartato Aminotransferasas/metabolismo , Aspartato Aminotransferasas/genética , Especificidad por Sustrato , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/genética , Simbiosis , MutaciónRESUMEN
The bean yellow mosaic virus (BYMV) is one of the most serious economic diseases affecting faba bean crop production. Rhizobium spp., well known for its high nitrogen fixation capacity in legumes, has received little study as a possible biocontrol agent and antiviral. Under greenhouse conditions, foliar application of molecularly characterized Rhizobium leguminosarum bv. viciae strain 33504-Borg201 to the faba bean leaves 24 h before they were infected with BYMV made them much more resistant to the disease while also lowering its severity and accumulation. Furthermore, the treatment promoted plant growth and health, as evidenced by the increased total chlorophyll (32.75 mg/g f.wt.) and protein content (14.39 mg/g f.wt.), as well as the improved fresh and dry weights of the plants. The protective effects of 33504-Borg201 greatly lowered the levels of hydrogen peroxide (H2O2) (4.92 µmol/g f.wt.) and malondialdehyde (MDA) (173.72 µmol/g f.wt.). The antioxidant enzymes peroxidase (1.58 µM/g f.wt.) and polyphenol oxidase (0.57 µM/g f.wt.) inhibited the development of BYMV in plants treated with 33504-Borg201. Gene expression analysis showed that faba bean plants treated with 33504-Borg201 had higher amounts of pathogenesis-related protein-1 (PR-1) (3.28-fold) and hydroxycinnamoyl-CoA quinate hydroxycinnamoyltransferase (4.13-fold) than control plants. These findings demonstrate the potential of 33,504-Borg201 as a cost-effective and eco-friendly method to protect faba bean plants against BYMV. Implementing this approach could help develop a simple and sustainable strategy for protecting faba bean crops from the devastating effects of BYMV.
Asunto(s)
Enfermedades de las Plantas , Hojas de la Planta , Rhizobium leguminosarum , Vicia faba , Rhizobium leguminosarum/genética , Rhizobium leguminosarum/crecimiento & desarrollo , Rhizobium leguminosarum/fisiología , Vicia faba/virología , Vicia faba/microbiología , Enfermedades de las Plantas/microbiología , Enfermedades de las Plantas/virología , Enfermedades de las Plantas/prevención & control , Hojas de la Planta/microbiología , Hojas de la Planta/virología , Resistencia a la Enfermedad , Peróxido de Hidrógeno/metabolismoRESUMEN
Bacterial persistence in the rhizosphere and colonization of root niches are critical for the establishment of many beneficial plant-bacteria interactions including those between Rhizobium leguminosarum and its host legumes. Despite this, most studies on R. leguminosarum have focused on its symbiotic lifestyle as an endosymbiont in root nodules. Here, we use random barcode transposon sequencing to assay gene contributions of R. leguminosarum during competitive growth in the rhizosphere and colonization of various plant species. This facilitated the identification of 189 genes commonly required for growth in diverse plant rhizospheres, mutation of 111 of which also affected subsequent root colonization (rhizosphere progressive), and a further 119 genes necessary for colonization. Common determinants reveal a need to synthesize essential compounds (amino acids, ribonucleotides, and cofactors), adapt metabolic function, respond to external stimuli, and withstand various stresses (such as changes in osmolarity). Additionally, chemotaxis and flagella-mediated motility are prerequisites for root colonization. Many genes showed plant-specific dependencies highlighting significant adaptation to different plant species. This work provides a greater understanding of factors promoting rhizosphere fitness and root colonization in plant-beneficial bacteria, facilitating their exploitation for agricultural benefit.
Asunto(s)
Raíces de Plantas , Rhizobium leguminosarum , Rizosfera , Simbiosis , Raíces de Plantas/microbiología , Rhizobium leguminosarum/genética , Rhizobium leguminosarum/crecimiento & desarrollo , Rhizobium leguminosarum/fisiología , Fabaceae/microbiología , Fabaceae/crecimiento & desarrollo , Microbiología del SueloRESUMEN
Lentil is one of the most important legumes cultivated in various provinces of Iran. However, there is limited information about the symbiotic rhizobia of lentils in this country. In this study, molecular identification of lentil-nodulating rhizobia was performed based on 16S-23S rRNA intergenic spacer (IGS) and recA, atpD, glnII, and nodC gene sequencing. Using PCR-RFLP analysis of 16S-23S rRNA IGS, a total of 116 rhizobia isolates were classified into 20 groups, leaving seven strains unclustered. Phylogenetic analysis of representative isolates revealed that the rhizobia strains belonged to Rhizobium leguminosarum and Rhizobium laguerreae, and the distribution of the species is partially related to geographical location. Rhizobium leguminosarum was the dominant species in North Khorasan and Zanjan, while R. laguerreae prevailed in Ardabil and East Azerbaijan. The distribution of the species was also influenced by agroecological climates; R. leguminosarum thrived in cold semiarid climates, whereas R. laguerreae adapted to humid continental climates. Both species exhibited equal dominance in the Mediterranean climate, characterized by warm, dry summers and mild, wet winters, in Lorestan and Kohgiluyeh-Boyer Ahmad provinces.
Asunto(s)
ADN Bacteriano , Lens (Planta) , Filogenia , Rhizobium , Lens (Planta)/microbiología , Irán , Rhizobium/genética , Rhizobium/clasificación , Rhizobium/aislamiento & purificación , ADN Bacteriano/genética , ARN Ribosómico 16S/genética , Clima , ADN Espaciador Ribosómico/genética , Polimorfismo de Longitud del Fragmento de Restricción , Análisis de Secuencia de ADN , ARN Ribosómico 23S/genética , Rhizobium leguminosarum/genética , Rhizobium leguminosarum/clasificación , Rhizobium leguminosarum/aislamiento & purificación , Simbiosis , Proteínas Bacterianas/genética , Reacción en Cadena de la PolimerasaRESUMEN
Red clover (Trifolium pratense L.) is a forage legume cultivated worldwide. This plant is capable of establishing a nitrogen-fixing symbiosis with Rhizobium leguminosarum symbiovar trifolii strains. To date, no comparative analysis of the symbiotic properties and heterogeneity of T. pratense microsymbionts derived from two distinct geographic regions has been performed. In this study, the symbiotic properties of strains originating from the subpolar and temperate climate zones in a wide range of temperatures (10-25 °C) have been characterized. Our results indicate that all the studied T. pratense microsymbionts from two geographic regions were highly efficient in host plant nodulation and nitrogen fixation in a wide range of temperatures. However, some differences between the populations and between the strains within the individual population examined were observed. Based on the nodC and nifH sequences, the symbiotic diversity of the strains was estimated. In general, 13 alleles for nodC and for nifH were identified. Moreover, 21 and 61 polymorphic sites in the nodC and nifH sequences were found, respectively, indicating that the latter gene shows higher heterogeneity than the former one. Among the nodC and nifH alleles, three genotypes (I-III) were the most frequent, whereas the other alleles (IV-XIII) proved to be unique for the individual strains. Based on the nodC and nifH allele types, 20 nodC-nifH genotypes were identified. Among them, the most frequent were three genotypes marked as A (6 strains), B (5 strains), and C (3 strains). Type A was exclusively found in the temperate strains, whereas types B and C were identified in the subpolar strains. The remaining 17 genotypes were found in single strains. In conclusion, our data indicate that R. leguminosarum sv. trifolii strains derived from two climatic zones show a high diversity with respect to the symbiotic efficiency and heterogeneity. However, some of the R. leguminosarum sv. trifolii strains exhibit very good symbiotic potential in the wide range of the temperatures tested; hence, they may be used in the future for improvement of legume crop production.
Asunto(s)
Fabaceae , Rhizobium leguminosarum , Rhizobium , Trifolium , Rhizobium leguminosarum/genética , Simbiosis/genética , Fabaceae/genética , Trifolium/genética , Fijación del Nitrógeno , Filogenia , Rhizobium/genética , ADN Bacteriano/genéticaRESUMEN
Motility and chemotaxis are crucial processes for soil bacteria and plant-microbe interactions. This applies to the symbiotic bacterium Rhizobium leguminosarum, where motility is driven by flagella rotation controlled by two chemotaxis systems, Che1 and Che2. The Che1 cluster is particularly important in free-living motility prior to the establishment of the symbiosis, with a che1 mutant delayed in nodulation and reduced in nodulation competitiveness. The Che2 system alters bacteroid development and nodule maturation. In this work, we also identified 27 putative chemoreceptors encoded in the R. leguminosarum bv. viciae 3841 genome and characterized its motility in different growth conditions. We describe a metabolism-based taxis system in rhizobia that acts at high concentrations of dicarboxylates to halt motility independent of chemotaxis. Finally, we show how PTSNtr influences cell motility, with PTSNtr mutants exhibiting reduced swimming in different media. Motility is restored by the active forms of the PTSNtr output regulatory proteins, unphosphorylated ManX and phosphorylated PtsN. Overall, this work shows how rhizobia typify soil bacteria by having a high number of chemoreceptors and highlights the importance of the motility and chemotaxis mechanisms in a free-living cell in the rhizosphere, and at different stages of the symbiosis.
Asunto(s)
Rhizobium leguminosarum , Rhizobium , Simbiosis , Proteínas Bacterianas/metabolismo , Rhizobium leguminosarum/genética , Rhizobium leguminosarum/metabolismo , SueloRESUMEN
High temperature is one of the most important factors limiting legume productivity. We have previously shown the induction of senescence in the apical part of nodules of the pea SGE line, formed by Rhizobium leguminosarum bv. viciae strain 3841, when they were exposed to elevated temperature (28 °C). In this study, we analyzed the potential involvement of abscisic acid (ABA), ethylene, and gibberellins in apical senescence in pea nodules under elevated temperature. Immunolocalization revealed an increase in ABA and 1-aminocyclopropane-1-carboxylic acid (ACC, the precursor of ethylene biosynthesis) levels in cells of the nitrogen fixation zone in heat-stressed nodules in 1 day of exposure compared to heat-unstressed nodules. Both ABA and ethylene appear to be involved in the earliest responses of nodules to heat stress. A decrease in the gibberellic acid (GA3) level in heat-stressed nodules was observed. Exogenous GA3 treatment induced a delay in the degradation of the nitrogen fixation zone in heat-stressed nodules. At the same time, a decrease in the expression level of many genes associated with nodule senescence, heat shock, and defense responses in pea nodules treated with GA3 at an elevated temperature was detected. Therefore, apical senescence in heat-stressed nodules is regulated by phytohormones in a manner similar to natural senescence. Gibberellins can be considered as negative regulators, while ABA and ethylene can be considered positive regulators.
Asunto(s)
Reguladores del Crecimiento de las Plantas , Rhizobium leguminosarum , Giberelinas , Temperatura , Rhizobium leguminosarum/genética , Etilenos , Ácido Abscísico/metabolismo , Fijación del Nitrógeno/genética , Simbiosis/fisiologíaRESUMEN
Despite global warming, the influence of heat on symbiotic nodules is scarcely studied. In this study, the effects of heat stress on the functioning of nodules formed by Rhizobium leguminosarum bv. viciae strain 3841 on pea (Pisum sativum) line SGE were analyzed. The influence of elevated temperature was analyzed at histological, ultrastructural, and transcriptional levels. As a result, an unusual apical pattern of nodule senescence was revealed. After five days of exposure, a senescence zone with degraded symbiotic structures was formed in place of the distal nitrogen fixation zone. There was downregulation of various genes, including those associated with the assimilation of fixed nitrogen and leghemoglobin. After nine days, the complete destruction of the nodules was demonstrated. It was shown that nodule recovery was possible after exposure to elevated temperature for 3 days but not after 5 days (which coincides with heat wave duration). At the same time, the exposure of plants to optimal temperature during the night leveled the negative effects. Thus, the study of the effects of elevated temperature on symbiotic nodules using a well-studied pea genotype and Rhizobium strain led to the discovery of a novel positional response of the nodule to heat stress.
Asunto(s)
Rhizobium leguminosarum , Rhizobium , Pisum sativum , Temperatura , Rhizobium leguminosarum/genética , Rhizobium/genética , Fijación del Nitrógeno/genética , Simbiosis/fisiologíaRESUMEN
Symbiotic nitrogen fixation (SNF) by rhizobium, a Gram-negative soil bacterium, is an essential component in the nitrogen cycle and is a sustainable green way to maintain soil fertility without chemical energy consumption. SNF, which results from the processes of nodulation, rhizobial infection, bacteroid differentiation and nitrogen-fixing reaction, requires the expression of various genes from both symbionts with adaptation to the changing environment. To achieve successful nitrogen fixation, rhizobia and their hosts cooperate closely for precise regulation of symbiotic genes, metabolic processes and internal environment homeostasis. Many researches have progressed to reveal the ample information about regulatory aspects of SNF during recent decades, but the major bottlenecks regarding improvement of nitrogen-fixing efficiency has proven to be complex. In this mini-review, we summarize recent advances that have contributed to understanding the rhizobial regulatory aspects that determine SNF efficiency, focusing on the coordinated regulatory mechanism of symbiotic genes, oxygen, carbon metabolism, amino acid metabolism, combined nitrogen, non-coding RNAs and internal environment homeostasis. Unraveling regulatory determinants of SNF in the nitrogen-fixing protagonist rhizobium is expected to promote an improvement of nitrogen-fixing efficiency in crop production.
Asunto(s)
Fabaceae , Rhizobium leguminosarum , Rhizobium , Rhizobium/metabolismo , Rhizobium leguminosarum/genética , Fijación del Nitrógeno/fisiología , Nódulos de las Raíces de las Plantas/microbiología , Simbiosis/fisiología , Bacterias Gramnegativas , Nitrógeno/metabolismo , Suelo , Fabaceae/microbiologíaRESUMEN
While shaping of plant microbiome composition through 'host filtering' is well documented in legume-rhizobium symbioses, it is less clear to what extent different varieties and genotypes of the same plant species differentially influence symbiont community diversity and composition. Here, we compared how clover host varieties and genotypes affect the structure of Rhizobium populations in root nodules under conventional field and controlled greenhouse conditions. We first grew four Trifolium repens (white clover) F2 crosses and one variety in a conventional field trial and compared differences in root nodule Rhizobium leguminosarum symbiovar trifolii (Rlt) genotype diversity using high-throughput amplicon sequencing of chromosomal housekeeping (rpoB and recA) genes and auxiliary plasmid-borne symbiosis genes (nodA and nodD). We found that Rlt nodule diversities significantly differed between clover crosses, potentially due to host filtering. However, variance in Rlt diversity largely overlapped between crosses and was also explained by the spatial distribution of plants in the field, indicative of the role of local environmental conditions for nodule diversity. To test the effect of host filtering, we conducted a controlled greenhouse trial with a diverse Rlt inoculum and several host genotypes. We found that different clover varieties and genotypes of the same variety selected for significantly different Rlt nodule communities and that the strength of host filtering (deviation from the initial Rhizobium inoculant composition) was positively correlated with the efficiency of symbiosis (rate of plant greenness colouration). Together, our results suggest that selection by host genotype and local growth conditions jointly influence white clover Rlt nodule diversity and community composition.
Asunto(s)
Rhizobium leguminosarum , Rhizobium , Trifolium , Trifolium/genética , Medicago/genética , Rhizobium leguminosarum/genética , Simbiosis/genética , PlantasRESUMEN
The Pss-I region of Rhizobium leguminosarum bv. trifolii TA1 comprises more than 20 genes coding for glycosyltransferases, modifying enzymes, and polymerization/export proteins, altogether determining the biosynthesis of symbiotically relevant exopolysaccharides. In this study, the role of homologous PssG and PssI glycosyltransferases in exopolysaccharide subunit synthesis were analyzed. It was shown that the glycosyltransferase-encoding genes of the Pss-I region were part of a single large transcriptional unit with potential downstream promoters activated in specific conditions. The ΔpssG and ΔpssI mutants produced significantly lower amounts of the exopolysaccharide, while the double deletion mutant ΔpssIΔpssG produced no exopolysaccharide. Complementation of double mutation with individual genes restored exopolysaccharide synthesis, but only to the level similar to that observed for the single ΔpssI or ΔpssG mutants, indicating that PssG and PssI serve complementary functions in the process. PssG and PssI interacted with each other in vivo and in vitro. Moreover, PssI displayed an expanded in vivo interaction network comprising other GTs involved in subunit assembly and polymerization/export proteins. PssG and PssI proteins were shown to interact with the inner membrane through amphipathic helices at their C-termini, and PssG also required other proteins involved in exopolysaccharide synthesis to localize in the membrane protein fraction.
Asunto(s)
Rhizobium leguminosarum , Rhizobium leguminosarum/genética , Glicosiltransferasas/metabolismo , Mutación , Fijación del Nitrógeno/genética , Polisacáridos Bacterianos/metabolismo , Proteínas Bacterianas/metabolismo , SimbiosisRESUMEN
Although rhizobia that establish a nitrogen-fixing symbiosis with legumes are also known to promote growth in non-legumes, studies on rhizobial associations with wheat roots are scarce. We searched for Rhizobium leguminosarum symbiovar viciae (Rlv) strains naturally competent to endophytically colonize wheat roots. We isolated 20 strains from surface-sterilized wheat roots and found a low diversity of Rlv compared to that observed in the Rlv species complex. We tested the ability of a subset of these Rlv for wheat root colonization when co-inoculated with other Rlv. Only a few strains, including those isolated from wheat roots, and one strain isolated from pea nodules, were efficient in colonizing roots in co-inoculation conditions, while all the strains tested in single strain inoculation conditions were found to colonize the surface and interior of roots. Furthermore, Rlv strains isolated from wheat roots were able to stimulate root development and early arbuscular mycorrhizal fungi colonization. These responses were strain and host genotype dependent. Our results suggest that wheat can be an alternative host for Rlv; nevertheless, there is a strong competition between Rlv strains for wheat root colonization. In addition, we showed that Rlv are endophytic wheat root bacteria with potential ability to modify wheat development.
Asunto(s)
Rhizobium leguminosarum , Rhizobium , Rhizobium leguminosarum/genética , Endófitos/genética , Triticum , Filogenia , Simbiosis/genética , Bacterias/genética , Nódulos de las Raíces de las Plantas/microbiologíaRESUMEN
Rhizobia are soil-borne bacteria forming symbiotic associations with legumes and fixing atmospheric dinitrogen. The nitrogen-fixation potential depends on the type of host plants and microsymbionts as well as environmental factors that affect the distribution of rhizobia. In this study, we compared genetic diversity of bacteria isolated from root nodules of Trifolium pratense grown in two geographical regions (Tromsø, Norway and Lublin, Poland) located in distinct climatic (subpolar and temperate) zones. To characterize these isolates genetically, three PCR-based techniques (ERIC, BOX, and RFLP of the 16S-23S rRNA intergenic spacer), 16S rRNA sequencing, and multi-locus sequence analysis of chromosomal house-keeping genes (atpD, recA, rpoB, gyrB, and glnII) were done. Our results indicate that a great majority of the isolates are T. pratense microsymbionts belonging to Rhizobium leguminosarum sv. trifolii. A high diversity among these strains was detected. However, a lower diversity within the population derived from the subpolar region in comparison to that of the temperate region was found. Multi-locus sequence analysis showed that a majority of the strains formed distinct clusters characteristic for the individual climatic regions. The subpolar strains belonged to two (A and B) and the temperate strains to three R. leguminosarum genospecies (B, E, and K), respectively.
Asunto(s)
Rhizobium leguminosarum , Rhizobium , Trifolium , ADN Bacteriano/genética , Filogenia , Polimorfismo de Longitud del Fragmento de Restricción , ARN Ribosómico 16S/genética , Rhizobium/genética , Rhizobium leguminosarum/genética , Nódulos de las Raíces de las Plantas/microbiología , Análisis de Secuencia de ADN , Simbiosis/genética , Trifolium/genética , Trifolium/microbiologíaRESUMEN
Acacia saligna is an invasive alien species that has the ability to establish symbiotic relationships with rhizobia. In the present study, genotypic and symbiotic diversity of native rhizobia associated with A. saligna in Tunisia were studied. A total of 100 bacterial strains were selected and three different ribotypes were identified based on rrs PCR-RFLP analysis. Sequence analyses of rrs and four housekeeping genes (recA, atpD, gyrB and glnII) assigned 30 isolates to four putative new lineages and a single strain to Sinorhizobium meliloti. Thirteen slow-growing isolates representing the most dominant IGS (intergenic spacer) profile clustered distinctly from known rhizobia species within Bradyrhizobium with the closest related species being Bradyrhizobium shewense and Bradyrhizobium niftali, which had 95.17% and 95.1% sequence identity, respectively. Two slow-growing isolates, 1AS28L and 5AS6L, had B. frederekii as their closest species with a sequence identity of 95.2%, an indication that these strains could constitute a new lineage. Strains 1AS14I, 1AS12I and 6AS6 clustered distinctly from known rhizobia species but within the Rhizobium leguminosarum complex (Rlc) with the most closely related species being Rhizobium indicum with 96.3% sequence identity. Similarly, the remaining 11 strains showed 96.9 % and 97.2% similarity values with R. changzhiense and R. indicum, respectively. Based on nodC and nodA phylogenies and cross inoculation tests, these 14 strains of Rlc species clearly diverged from strains of Sinorhizobium and Rlc symbiovars, and formed a new symbiovar for which the name sv. "salignae" is proposed. Bacterial strains isolated in this study that were taxonomically assigned to Bradyrhizobium harbored different symbiotic genes and the data suggested a new symbiovar, for which sv. "cyanophyllae" is proposed. Isolates formed effective nodules on A. saligna.
Asunto(s)
Acacia , Bradyrhizobium , Rhizobium leguminosarum , Rhizobium , ADN Bacteriano/genética , Filogenia , ARN Ribosómico 16S/genética , Rhizobium leguminosarum/genética , Nódulos de las Raíces de las Plantas/microbiología , Análisis de Secuencia de ADN , Simbiosis/genética , TúnezRESUMEN
Nitrogen uptake in legumes is facilitated by bacteria such as Rhizobium leguminosarum. For this bacterium, gene expression data are available, but functional gene annotation is less well developed than for other model organisms. More annotations could lead to a better understanding of the pathways for growth, plant colonization, and nitrogen fixation in R. leguminosarum. In this study, we present a pipeline that combines novel scores from gene coexpression network analysis in a principled way to identify the genes that are associated with certain growth conditions or highly coexpressed with a predefined set of genes of interest. This association may lead to putative functional annotation or to a prioritized list of genes for further study.
Asunto(s)
Rhizobium leguminosarum , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Fijación del Nitrógeno/genética , Rhizobium leguminosarum/genética , Rhizobium leguminosarum/metabolismoRESUMEN
Rhizobium leguminosarum has been widely used as a model to study nodule biochemistry, its genomic sequence has been published. We screened the Rhizobium leguminosarum bv. viciae 3841 genome sequence using a bioinformatics analysis for discovering potential small non-coding RNAs. One of these identified non-coding RNAs, cis-encoded antisense RLS1, was found to affect the symbiotic nodulation and nitrogen fixation. The mature form of RLS1 was 258 nt of non-coding RNA, its disruption mutant strain (â³RLS1) caused that the nodulation stages were delayed dramatically and the total number of nodules decreased, leading to a 25% reduction in the total amount of nitrogen fixed in the symbiotic system of Rhizobium- Pisum sativum, compared with wild-type strain. RLS1 targets an ABC transporter mRNA, bind to Hfq in vitro, and to be stable in the absence of Hfq. Further analysis showed that Hfq is not required for interactions between RLS1 and its target mRNAs. â³RLS1 strain exhibited that its production of extracellular polysaccharide (EPS) was over three times higher than in wild-type strain. The findings suggest that RLS1 might affect nodulation by participating in the regulatory network for EPS accurate secretion, playing a pivotal role in the infection process and in root nodule formation.