ABSTRACT The leguminous inoculation with nodule-inducing bacteria that perform biological nitrogen fixation is a good example of an “eco-friendly agricultural practice”. Bradyrhizobium strains BR 3267 and BR 3262 are recommended for cowpea (Vigna unguiculata) inoculation in Brazil and showed remarkable responses; nevertheless neither strain was characterized at species level, which is our goal in the present work using a polyphasic approach. The strains presented the typical phenotype of Bradyrhizobium with a slow growth and a white colony on yeast extract-mannitol medium. Strain BR 3267 was more versatile in its use of carbon sources compared to BR 3262. The fatty acid composition of BR 3267 was similar to the type strain of Bradyrhizobium yuanmingense; while BR 3262 was similar to Bradyrhizobium elkanii and Bradyrhizobium pachyrhizi. Phylogenetic analyses based on 16S rRNA and three housekeeping genes placed both strains within the genus Bradyrhizobium: strain BR 3267 was closest to B. yuanmingense and BR 3262 to B. pachyrhizi. Genome average nucleotide identity and DNA–DNA reassociation confirmed the genomic identification of B. yuanmingense BR 3267 and B. pachyrhizi BR 3262. The nodC and nifH gene analyses showed that strains BR 3267 and BR 3262 hold divergent symbiotic genes. In summary, the results indicate that cowpea can establish effective symbiosis with divergent bradyrhizobia isolated from Brazilian soils.

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.

ABSTRACT The strain BR 3351T (Bradyrhizobium manausense) was obtained from nodules of cowpea (Vigna unguiculata L. Walp) growing in soil collected from Amazon rainforest. Furthermore, it was observed that the strain has high capacity to fix nitrogen symbiotically in symbioses with cowpea. We report here the draft genome sequence of strain BR 3351T. The information presented will be important for comparative analysis of nodulation and nitrogen fixation for diazotrophic bacteria. A draft genome with 9,145,311 bp and 62.9% of GC content was assembled in 127 scaffolds using 100 bp pair-end Illumina MiSeq system. The RAST annotation identified 8603 coding sequences, 51 RNAs genes, classified in 504 subsystems.

Abstract The strain BR 3267 is a nitrogen-fixing symbiotic bacteria isolated from soil of semi-arid area of Brazilian Northeast using cowpea as the trap plant. This strain is used as commercial inoculant for cowpea and presents high efficient in nitrogen fixation as consequence of its adaptation potential to semi-arid conditions. We report here the draft genome sequence of Bradyrhizobium sp. strain BR 3267, an elite bacterium used as inoculant for cowpea. Whole genome sequencing of BR 3267 using Illumina MiSeq sequencing technology has 55 scaffolds with a total genome size of 7,904,309 bp and C+G 63%. Annotation was added by the RAST prokaryotic genome annotation service and has shown 7314 coding sequences and 52 RNA genes.

Abstract The strain BR 3262 was isolated from nodule of cowpea (Vigna unguiculata L. Walp) growing in soil of the Atlantic Forest area in Brazil and it is reported as an efficient nitrogen fixing bacterium associated to cowpea. Firstly, this strain was assigned as Bradyrhizobium elkanii, however, recently a more detailed genetic and molecular characterization has indicated it could be a Bradyrhizobium pachyrhizi species. We report here the draft genome sequence of B. pachyrhizi strain BR 3262, an elite bacterium used as inoculant for cowpea. The whole genome with 116 scaffolds, 8,965,178 bp and 63.8% of C+G content for BR 3262 was obtained using Illumina MiSeq sequencing technology. Annotation was added by the RAST prokaryotic genome annotation service and shown 8369 coding sequences, 52 RNAs genes, classified in 504 subsystems.

A composição de comunidades e diversidade de fungos micorrízicos arbusculares (FMAs) pode ser influenciada por diversos fatores como o clima, a biota do solo e as plantas hospedeiras. Este estudo objetivou avaliar a influência da revegetação de uma cava de extração de argila com Sesbania virgata (SV) em plantios puro e consorciado com Eucalyptus camaldulensis (EC) e Acacia mangium (AM), na composição e diversidade de fungos micorrízicos arbusculares (FMAs), bem como na quantidade da proteína do solo relacionada à glomalina (PSRG). O delineamento experimental utilizado foi em blocos casualizados com quatro tratamentos (plantio puro de SV - 100SV; consórcio de SV + EC - 50SV:50EC; consórcio de SV + AM - 50SV:50AM; e área degradada com vegetação espontânea - ADVE), com três repetições. A revegetação da cava de extração de argila com SV, em plantios puro ou consorciado, reduziu a abundância de esporos e a dominância de espécies (Índice de Simpson - IDS) e aumentou a riqueza de espécies de FMAs e o índice de diversidade de Shannon e Wiener. Além disso, aumentou a quantidade de proteína do solo relacionada à glomalina, quando comparada a área degradada com vegetação espontânea.

The community composition and diversity of arbuscular mycorrhizal fungi (AMF) can be affected by several factors, including the climate, soil biota and host plants. This study aimed to evaluate the influence of Sesbania virgata (SV) revegetation in pure and intercropped with plantation Eucalyptus camaldulensis (EC) and Acacia mangium (AM), in the composition and diversity of arbuscular mycorrhizal fungi (AMF), and the amount glomalin-related soil protein (GRSP) in an area damaged by clay extraction. The experimental design was a randomized block with four treatments (planting pure SV - 100SV); consortium of SV + EC - 50SV: 50EC; consortium AM SV + - 50SV: 50AM) and degraded area with natural vegetation - ADVE) and three replications. Revegetation of mining digging clay with SV in pure or intercropped planting reduced the abundance of spores and increased species diversity of arbuscular mycorrhizal fungi. Furthermore, increased the amount of glomalin-related soil protein compared with the degraded area and spontaneous vegetation.

RESUMO Esse estudo teve como objetivo avaliar a influência da revegetação com Eucalyptus camaldulensis e Acacia mangium, em plantio puro e consorciado, na composição e diversidade de fungos micorrízicos arbusculares (FMAs), bem como na quantidade da proteína do solo relacionada à glomalina (PSRG) em uma área degradada pela extração de argila. O delineamento experimental utilizado foi o de blocos casualizados com quatro tratamentos (plantios puros de Eucalyptus camaldulensis e Acacia mangium; consórcio de Eucalyptus camaldulensis + Acacia mangium; e área degradada com vegetação espontânea - ADVE) e três repetições, onde, em cada uma das parcelas, foram coletadas amostras de solo na camada 0-5 cm. Os esporos de FMA foram extraídos e taxonomicamente identificados. Analisou-se a densidade relativa, a frequência de cada espécie e os índices de Shannon-Wiener, Pielou e de Simpson. A PSRG (glomalina total - GT e glomalina facilmente extraível - GFE) foi extraída com citrato de sódio e quantificada pelo método de Bradford. A abundância de FMAs foi maior na área degradada com vegetação espontânea quando comparada aos plantios, em contrapartida apresentou baixa diversidade de espécies. As áreas de eucalipto em monocultivo mostraram menor índice de diversidade de FMAs em relação às áreas de eucalipto consorciadas com a acácia. Glomus e Acaulospora foram os gêneros de FMAs que apresentaram o maior número de espécies. A PSRG foi estreitamente correlacionada com o C e o N do solo, tendo sido observada em maiores quantidades nos plantios, em relação à ADVE. A revegetação da cava de extração de argila promoveu redução na esporulação dos FMAs, enquanto a diversidade e a quantidade de PSRG foram aumentadas.

ABSTRACT The objective of this study was to evaluate the influence of revegetation with Eucalyptus camaldulensis and Acacia mangium in pure and mixed stands in the composition and mycorrhizal fungi diversity (AMF), as well as in the production of glomalin-related soil protein (GRSP) of an area degraded by clay extraction. The experimental design used was randomized complete block with four treatments (pure stands Eucalyptus camaldulensis and Acacia mangium; mixed Eucalyptus camaldulensis + Acacia mangium; and covered with spontaneous vegetation - ADVE) and three replications. Soil samples were collected at 0-5 cm soil layer in each plot. The spores were extracted and taxonomically identified. Relative density, frequency of each species and the Shannon-Wiener, Pielou and Simpson indexes were analyzed. The GRSP (total glomalin - TG and easily extractable glomalin - EEG) was extracted with sodium citrate and quantified by the Bradford method. Abundance of AMF was higher in the degraded areas covered by weeds (spontaneous vegetation) compared to plantations; however, it showed lower species diversity. The areas of eucalypt monoculture showed a lower level of AMF diversity in relation to areas of eucalypt intercropped with Acacia. The genera Glomus and Acaulospora were the AMF, with the largest number of species. The GRSP was closely correlated with soil C and N, which observed in greater amounts in plantations in relation to the sites covered with spontaneous vegetation. Revegetation of clay extraction site promoted the reduction of AMF sporulation, while the diversity and amount GRSP increased.

O objetivo deste trabalho foi avaliar o efeito isolado ou simultâneo dos estresses hídrico e térmico na expressão gênica em nódulos de feijão-caupi. A bactéria Bradyrhizobium japonicum (estirpe BR 3267) foi inoculada em sementes de feijão-caupi da cultivar IPA 206 e, 35 dias após a germinação, as plantas foram submetidas a diferentes regimes de disponibilidade hídrica e a estresse térmico, em casa de vegetação. Para a identificação dos genes diferencialmente expressos, foi utilizada a técnica de cDNA-AFLP, tendo-se isolado 67 fragmentos derivados de transcritos (FDTs) diferencialmente expressos. Após o sequenciamento dos FDTs e das análises de similaridade, com uso do programa Blastx, foram identificados 14 genes diferencialmente expressos envolvidos em diferentes processos metabólicos. O padrão de expressão de seis genes sob estresse abiótico foi confirmado por RT-qPCR, e observou-se indução de genes pertencentes a diferentes categorias funcionais, como biossíntese de ácido abscísico, sinalização celular, transportador de prolina e biossíntese de lipídeos de membranas. A expressão desses genes indica sua participação em processos relacionados à proteção dos nódulos ao estresse abiótico.

The objective of this work was to evaluate the isolated or concomitant effect of drought and heat stresses on the gene expression of cowpea nodules. The bacterium Bradyrhizobium japonicum (strain BR 3267) was inoculated in cowpea seeds from the cultivar IPA 206 and, 35 days after germination, the plants were subjected to different water availability regimes and to heat stress under greenhouse conditions. To identify differentially expressed genes, the cDNA-AFLP technique was used, and 67 differentially expressed transcript-derived fragments (TDFs) were isolated. After TDFs sequencing and similarity analyses, using the Blastx program, 14 differentially expressed genes were identified, which are involved in different metabolic processes. The expression pattern of six genes under abiotic stress conditions was confirmed by RT-qPCR, and the induction of different functional gene categories, such as abscisic acid biosynthesis, cell signaling, proline transporter, and lipid membrane biosynthesis, was observed. The expression of these genes indicates their participation in processes related to nodule protection under abiotic stresses.

MADS-box genes encode a family of transcription factors which control diverse developmental processes in flowering plants ranging from root to flower and fruit development. Members of the MADS-box gene family share a highly conserved sequence of approximately 180 nucleotides that encodes a DNA-binding domain. We used bioinformatics tools to investigate the information generated by the Eucalyptus Expressed Sequence Tag (FORESTs) genome project in order to identify and annotate MADS-box genes. The comparative phylogenetic analysis of the Eucalyptus MADS-box genes with Arabidopsis homologues allowed us to group them into one of the well-known subfamilies. Trends in gene expression of these putative Eucalyptus MADS-box genes were investigated by hierarchical clustering analysis. Among 24 MADS-box genes identified by our analysis, 12 are expressed in vegetative organs. Out of these, five are expressed predominately in wood. Understanding of the molecular mechanisms performed by MADS-box proteins underlying Eucalyptus growth, development and stress reactions would provide important insights into tree development and could reveal means by which tree characteristics could be modified for the improvement of industrial properties.

As plantas respondem ao estresse provocado por temperaturas elevadas através da síntese de um grupo de proteínas denominadas proteínas de choque térmico (Heat Shock Proteins-HSPs) que estão associadas com a obtenção de termotolerância. Nesse estudo, foi avaliada a indução de proteínas de choque térmico de baixo peso molecular (sHSPs) em oito cultivares de feijoeiro através de "Northern blot" utilizando o cDNA da W HSP 16,9 como uma sonda heteróloga. Caupi foi utilizado como controle, uma vez que, ao contrário do feijoeiro, é uma espécie que se apresenta bem adaptada às condições de temperaturas elevadas como as observadas no Semi-árido brasileiro. Após um período de crescimento, as plantas foram submetidas a um período de duas horas de choque térmico a 40 ºC. Todas as variedades avaliadas foram capazes de induzir mRNAs para sHSP que hibridizou com a sonda W HSP 16,9. Entretanto, foi observada uma diferença significativa no padrão de indução entre as diferentes variedades. Os níveis de mRNA e sHSPs induzidos em caupi após o choque térmico foram bem maiores que os observados para as variedades de feijoeiro. Além disso, a expressão de sHSPs foi também avaliada em relação ao acúmulo de proteínas através da análise de "Western-blot" para caupi e as variedades de feijoeiro IPA 7 e Negro Argel. O padrão de proteínas observado confirma que as sHSPs são diferencialmente expressas em diferentes variedades de feijoeiro de acordo com a tolerância ao estresse térmico.

Plants respond to temperature stress by synthesizing a set of heat shock proteins (HSPs), which may be responsible for the acquisition of thermotolerance. In this study, the induction of small HSPs (sHSPs) in eight common bean varieties was evaluated by Northern blot analysis using the W HSP 16.9 cDNA as heterologous probe. Cowpea was used, as a positive control since this plant, as opposed to common bean, is known to grow well under high temperature regimes such as that found in the Brazilian semi-arid region. After the growth period, the plants were submitted to two h of heat shock at 40 ºC. All varieties tested were able to induce sHSP mRNAs that hybridized with W HSP 16.9 probe. However, significant kinetic differences were found when comparing different varieties. SHSP mRNA levels induced after heat shock in cowpea was higher than the levels observed on the bean varieties displaying the highest expression of these proteins. Besides, the sHSP expression was also assessed at the protein accumulation level by Western-blot analysis for cowpea and both IPA 7 and Negro Argel varieties of bean plants. The revealed protein pattern confirmed that sHSPs are differentially expressed in distinct varieties of common bean according their heat stress tolerance.