Proteomic Analysis of Rhizobium favelukesii LPU83 in Response to Acid Stress.

Acid soils constitute a severe problem for leguminous crops mainly through a disturbance in rhizobium-legume interactions. Rhizobium favelukesii-an acid-tolerant rhizobium able to nodulate alfalfa-is highly competitive for nodule occupation under acid conditions but inefficient for biologic nitrogen fixation. In this work, we obtained a general description of the acid-stress response of R. favelukesii LPU83 by means of proteomics by comparing the total proteome profiles in the presence or absence of acid stress by nanoflow ultrahigh-performance liquid chromatography coupled to mass spectrometry. Thus, a total of 336 proteins were identified with a significant differential expression, 136 of which species were significantly overexpressed and 200 underexpressed in acidity. An in silico functional characterization with those respective proteins revealed a complex and pleiotropic response by these rhizobia involving components of oxidative phosphorylation, glutamate metabolism, and peptidoglycan biosynthesis, among other pathways. Furthermore, a lower permeability was evidenced in the acid-stressed cells along with several overexpressed proteins related to γ-aminobutyric acid metabolism, such as the gene product of livK, which gene was mutated. This mutant exhibited an acid-sensitive phenotype in agreement with the proteomics results. We conclude that both the γ-aminobutyric acid metabolism and a modified cellular envelope could be relevant to acid tolerance in R. favelukesii.

Nodule bacteria from the cultured legume Phaseolus dumosus (belonging to the Phaseolus vulgaris cross-inoculation group) with common tropici phenotypic characteristics and symbiovar but distinctive phylogenomic position and chromid.

Phaseolus dumosus is an endemic species from mountain tops in Mexico that was found in traditional agriculture areas in Veracruz, Mexico. P. dumosus plants were identified by ITS sequences and their nodules were collected from agricultural fields or from trap plant experiments in the laboratory. Bacteria from P. dumosus nodules were identified as belonging to the phaseoli-etli-leguminosarum (PEL) or to the tropici group by 16S rRNA gene sequences. We obtained complete closed genomes from two P. dumosus isolates CCGE531 and CCGE532 that were phylogenetically placed within the tropici group but with a distinctive phylogenomic position and low average nucleotide identity (ANI). CCGE531 and CCGE532 had common phenotypic characteristics with tropici type B rhizobial symbionts. Genome synteny analysis and ANI showed that P. dumosus isolates had different chromids and our analysis suggests that chromids have independently evolved in different lineages of the Rhizobium genus. Finally, we considered that P. dumosus and Phaseolus vulgaris plants belong to the same cross-inoculation group since they have conserved symbiotic affinites for rhizobia.

Agronomic efficiency of Rhizobium strains from the Amazon region in common bean / Eficiência agronômica de estirpes de Rhizobium isoladas da Amazônia sobre o feijoeiro-comum

To maximize the contribution of biological nitrogen fixation in common bean, it is necessary to use bacterial strains that are more adapted, competitive, and efficient in the symbiotic process. In this regard, the aim of this study was to evaluate the agronomic efficiency (AE) of three bacterial strains isolated from acid soils with high Al content from the Amazon region in an Argissolo Vermelho Distrófico típico soil (Typic Rhodustults - USDA Classification) from the municipality of Formiga, MG, Brazil. We compared their AE to that of the reference strain CIAT 899T and of two controls without inoculation (one without and another with 80 kg ha-1 of N-urea). The results indicated that inoculation with the strains UFLA 02-100 and UFLA 02-127 provides grain yield equivalent to inoculation with the reference strain and to the control with mineral N. Thus, both have potential for recommendation as inoculants for common bean.(AU)

Para maximizar a contribuição da fixação biológica de nitrogênio no feijoeiro-comum é necessária a utilização de estirpes de bactérias mais adaptadas, competitivas e eficientes no processo simbiótico. Nesse sentido, objetivou-se avaliar, em um Argissolo Vermelho Distrófico típico do município de Formiga-MG, a eficiência agronômica (EA) de três estirpes isoladas de solos ácidos e com alto teor de Al da Amazônia e comparar suas EA à da estirpe referência CIAT 899T e à de dois controles sem inoculação (um sem e outro com 80 kg ha-1 of N-ureia). Os resultados indicaram que a inoculação com as estirpes UFLA 02-100 e UFLA 02-127 propicia rendimento de grãos equivalente ao da estirpe referência e ao do controle com N mineral e que por isso, ambas têm potencial para ser recomendadas como inoculantes para o feijoeiro-comum.(AU)

Agronomic efficiency of Rhizobium strains from the Amazon region in common bean / Eficiência agronômica de estirpes de Rhizobium isoladas da Amazônia sobre o feijoeiro-comum

To maximize the contribution of biological nitrogen fixation in common bean, it is necessary to use bacterial strains that are more adapted, competitive, and efficient in the symbiotic process. In this regard, the aim of this study was to evaluate the agronomic efficiency (AE) of three bacterial strains isolated from acid soils with high Al content from the Amazon region in an Argissolo Vermelho Distrófico típico soil (Typic Rhodustults - USDA Classification) from the municipality of Formiga, MG, Brazil. We compared their AE to that of the reference strain CIAT 899T and of two controls without inoculation (one without and another with 80 kg ha-1 of N-urea). The results indicated that inoculation with the strains UFLA 02-100 and UFLA 02-127 provides grain yield equivalent to inoculation with the reference strain and to the control with mineral N. Thus, both have potential for recommendation as inoculants for common bean.

Para maximizar a contribuição da fixação biológica de nitrogênio no feijoeiro-comum é necessária a utilização de estirpes de bactérias mais adaptadas, competitivas e eficientes no processo simbiótico. Nesse sentido, objetivou-se avaliar, em um Argissolo Vermelho Distrófico típico do município de Formiga-MG, a eficiência agronômica (EA) de três estirpes isoladas de solos ácidos e com alto teor de Al da Amazônia e comparar suas EA à da estirpe referência CIAT 899T e à de dois controles sem inoculação (um sem e outro com 80 kg ha-1 of N-ureia). Os resultados indicaram que a inoculação com as estirpes UFLA 02-100 e UFLA 02-127 propicia rendimento de grãos equivalente ao da estirpe referência e ao do controle com N mineral e que por isso, ambas têm potencial para ser recomendadas como inoculantes para o feijoeiro-comum.

Expression of the Sinorhizobium meliloti small RNA gene mmgR is controlled by the nitrogen source.

Small non-coding regulatory RNAs (sRNAs) are key players in post-transcriptional regulation of gene expression. Hundreds of sRNAs have been identified in Sinorhizobium meliloti, but their biological function remains unknown for most of them. In this study, we characterized the expression pattern of the gene encoding the 77-nt sRNA MmgR in S. meliloti strain 2011. A chromosomal transcriptional reporter fusion (PmmgR-gfp) showed that the mmgR promoter is active along different stages of the interaction with alfalfa roots. In pure cultures, PmmgR-gfp activity paralleled the sRNA abundance indicating that mmgR expression is primarily controlled at the level of transcriptional initiation. PmmgR-gfp activity was higher during growth in rhizobial defined medium (RDM) than in TY medium. Furthermore, PmmgR-gfp was induced at 60 min after shifting growing cells from TY to RDM medium, i.e. shorter than the cell doubling time. In defined RDM medium containing NO3 (-), both PmmgR-gfp and MmgR level were repressed by the addition of tryptone or single amino acids, suggesting that mmgR expression depends on the cellular nitrogen (N) status. In silico analysis failed to detect conserved motifs upstream the promoter RNA polymerase binding site, but revealed a strongly conserved motif centered at -28 that may be linked to the observed regulatory pattern by the N source.

Efectividad de cepas rizobianas de frijolbajo diferentes regímenes de fósforo / Effectiveness of cowpea rhizobial strains under different phosphorus regimes

En Venezuela, el frijol representa una alternativa a la proteína animal, debido a su alto consumo y valor nutritivo, por ello se ha estimulado la implementación de programas para reactivar la economía de los pequeños y medianos productores, a fin de incrementar su producción y así tener mayor disponibilidad de proteína de alta calidad a bajo costo; de manera que, los estudios encaminados a mejorar su cultivo, son acertados. Se evaluó la efectividad de cepas rizobianas de crecimiento lento (cl) y rápido (cr) en frijol (Vigna unguiculata (L.) Walp.) cultivar TC9-6 en varios regímenes de fósforo (0, 20, 40 y 80 kgP2O5 ha-1), con un diseño experimental de bloques al azar con arreglo factorial. Las plantas se cultivaron en 4 kg de suelo de sabana 45 días y las cepas en caldo de levadura y manitol: 5 (cr: JV91) y 10 (cl: JV94) días. La inoculación (2 ml cada vez) fue aplicada a la siembra y 6 días más tarde. La utilización de fósforo (40-80 kgP2O5 ha-1) incrementó la nodulación (número, peso seco total e individual de nódulos) y favoreció la aparición de nódulos rojos; así mismo, acrecentó el peso de la materia seca, la altura, el número de hojas y la concentración de nitrógeno del vástago. Los valores fueron similares con ambos tipos de cepas (efectividad similar) y para las dos concentraciones (40-80 kgP2O5 ha-1), con las menores estimaciones para 0 y 20 kgP2O5 ha-1. De acuerdo con los resultados las concentraciones de 40 y 80 kgP2O5 ha-1 fueron las más favorables para el crecimiento y la nodulación de frijol.

In Venezuela, cowpea is an alternative to animal protein due to its high consumption and nutritious value, so it has stimulated the implementation of programs to reactivate the small and medium producers economy, in order to increase its production and to have major high quality protein availability at low cost; so that, the studies carry on to improve its cultivation, are well-aimed. The effectiveness of slow (sg) and fast (fg) growing rhizobial strains was evaluated in cowpea (Vigna unguiculata (L.) Walp) cultivar TC9-6 at various phosphorus regimes (0, 20, 40 and 80 kgP2O5 ha-1): randomized block design with factorial arrangement. Plants were cultivated in 4 kg savannah soil: 45 days, and the strains in yeast and mannitol broth: 5 (fg: JV91) and 10 (sg: JV94) days. The inoculation (2 ml each time) was applied at sowing time and 6 days later. Phosphorus utilization (40-80 kgP2O5 ha-1) increased nodulation (nodule number, total and individual dry weight) and favoured nodule red colour appearance; also, incremented shoot dry matter weight, height, leaves number and nitrogen concentration. Values were similar with both strain types (similar effectiveness) and to the two doses (40-80 kgP2O5 ha-1), with lower estimations to 0 and 20 kgP2O5 ha-1. Accordingly with the results, the doses of 40 and 80 kgP2O5 ha-1 were the most favourable to cowpea growth and nodulation.

[Lipopolysaccharides of rhizobiaceae: structure and biosynthesis]. / Lipopolisacáridos de Rhizobiaceae: estructura y biosíntesis.

The lipopolysaccharides (LPS) are major components of the outer membrane of Gram negative bacteria and, because of their location, are important mediators in the interaction between these bacteria and their environment and other organisms. The alpha-Proteobacterial family Rhizobiaceae includes the rhizobia and agrobacteria, microorganisms which establish symbiotic or parasitic relationships with plants. Mutants deficient in LPS biosynthesis show anomalous interactions with their hosts. The agronomical relevance of the relationship between rhizobia and agrobacteria with plants has promoted a large number of studies on the LPS from these bacteria. The complete structures of one or several domains of LPS from Rhizobiaceae have been determined in the last years. Additionally, several metabolic steps in the biosynthesis of these molecules have been elucidated. This review aims at the description of the more recent findings on the structure and biosynthesis of LPS in Rhizobium, Sinorhizobium and Agrobacterium.

Fused RolA protein enhances beta-glucoronidase activity 50-fold: implication for RolA mechanism of action.

We report that the plant oncoprotein RolA from Agrobacterium rhizogenes acts to stabilize beta-glucoronidase (Gus) when the two proteins are expressed as a fusion protein in transformed tobacco. The observed 50-fold increase of Gus activity was shown to be related to protein accumulation, with no significant changes in mRNA abundance, kinetic properties of the enzyme and thermostability. The entire RolA sequence is essential to achieve the full effect since both the N-terminal region, spanning a putative reverse signal-anchor and nuclear targeting sequences, or the contiguous C-terminal portion were shown to increase Gus activity only 10-fold. A possible interference of RolA in the protein degradation pathway regulated by auxin is discussed.

A structural model for the rolA protein and its interaction with DNA.

The study of the plant oncogene rolA has been hampered by a lack of structural information. Here we show that, despite a lack of significant sequence similarity to proteins of known structure, the rolA sequence adopts a known fold; that of the papillomavirus E2 DNA-binding domain. This fold is reliably identified by modern threading programs, which consider predicted secondary structure, but not by others. Although the rolA sequence is only around 16% identical to those of the available template structures, a structural model could be built that performed well against protein structure verification programs. The adopted strategy involved alignment corrections, justified by multiple model building and evaluation, with particular attention paid to the hydrophobic core residues. We find that rolA protein is predicted to resemble the template proteins in two key aspects; existence as a dimer and ability to bind DNA. rolA protein has recently been shown experimentally to possess DNA binding ability. This model predicts Lys 24 and Arg 27 to be involved in sequence-specific interactions and eight other residues to hydrogen-bond phosphate groups of the DNA.