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 The use of dark septate fungi (DSE) to promote plant growth can be beneficial to agriculture, and these organisms are important allies in the search for sustainable agriculture practices. This study investigates the contribution of dark septate fungi to the absorption of nutrients by rice plants and their ensuing growth. Four dark septate fungi isolates that were identified by Internal transcribed spacer phylogeny were inoculated in rice seeds (Cv. Piauí). The resulting root colonization was estimated and the kinetic parameters Vmax and Km were calculated from the nitrate contents of the nutrient solution. The macronutrient levels in the shoots, and the NO3--N, NH4+-N, free amino-N and soluble sugars in the roots, sheathes and leaves were measured. The rice roots were significantly colonized by all of the fungi, but in particular, isolate A103 increased the fresh and dry biomass of the shoots and the number of tillers per plant, amino-N, and soluble sugars as well as the N, P, K, Mg and S contents in comparison with the control treatment. When inoculated with isolates A103 and A101, the plants presented lower Km values, indicating affinity increases for NO3--N absorption. Therefore, the A103 Pleosporales fungus presented the highest potential for the promotion of rice plant growth, increasing the tillering and nutrients uptake, especially N (due to an enhanced affinity for N uptake) and P.

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.

O objetivo deste trabalho foi avaliar a contribuição de estirpes de rizóbio, eficientes na fixação de nitrogênio, sobre a produção de grãos de novas cultivares de feijão-caupi, indicadas para cultivo no Semiárido brasileiro, no Submédio do Vale do Rio São Francisco. Dois experimentos foram implantados nos perímetros irrigados de Mandacaru (Juazeiro, BA) e Bebedouro (Petrolina, PE). Os tratamentos consistiram da inoculação isolada de cinco estirpes de rizóbio - BR 3267, BR 3262, INPA 03-11B, UFLA 03-84 (Bradyrhizobiumsp.) e BR 3299T (Microvirga vignae) -, além de um tratamento com nitrogênio e de um controle sem inoculação ou aplicação de N. As seguintes cultivares de feijão-caupi foram avaliadas: BRS Pujante, BRS Tapaihum, BRS Carijó e BRS Acauã. Utilizou-se o delineamento experimental de blocos ao acaso, com quatro repetições. As plantas inoculadas apresentaram produtividade de grãos similar à observada em plantas adubadas com 80 kg ha-1 de N. As cultivares BRS Tapaihum e BRS Pujante destacaram-se quanto à produtividade e ao teor de proteínas nos grãos, quando inoculadas, o que mostra seu potencial para cultivo na região do Submédio do Vale do São Francisco.

The objective of this work was to evaluate the contribution of efficient nitrogen-fixing rhizobial strains to grain yield of new cowpea cultivars, indicated for cultivation in the Brazilian Semiarid region, in the sub-medium of the São Francisco River Valley. Two experiments were set up at the irrigated perimeters of Mandacaru (Juazeiro, state of Bahia) and Bebedouro (Petrolina, state of Pernambuco). The treatments consisted of single inoculation of five rhizobial strains - BR 3267, BR 3262, INPA 03-11B, UFLA 03-84 (Bradyrhizobiumsp.), and BR 3299T(Microvirga vignae) -, besides a treatment with nitrogen and a control without inoculation or N application. The following cowpea cultivars were evaluated: BRS Pujante, BRS Tapaihum, BRS Carijó, and BRS Acauã. A randomized complete block design, with four replicates, was used. Inoculated plants showed similar grain yield to the one observed with plants fertilized with 80 kg ha-1 N. The cultivars BRS Tapaihum and BRS Pujante stood out in grain yield and protein contents when inoculated, showing their potential for cultivation in the sub-medium of the São Francisco River Valley.

Devido à variabilidade das leguminosas quanto à eficiência da fixação biológica de nitrogênio (FBN), tem sido enfatizada a importância do melhoramento genético vegetal no favorecimento da fixação do nitrogênio. Este estudo teve como objetivo avaliar a eficiência simbiótica de progenitores de cultivares brasileiras de feijão-caupi e dar suporte científico à inclusão da FBN ao programa de melhoramento genético da cultura do feijão-caupi no Brasil. O experimento foi conduzido em Teresina, Piauí. Utilizou-se delineamento experimental em blocos casualizados em esquema fatorial 6 x 5, com três repetições, sendo seis fontes de N: quatro estirpes de bactérias diazotróficas (BR 3267, BR 3299, BR 3262 e INPA 03-11B), um tratamento com nitrogênio mineral e outro sem inoculação e sem N; e cinco genótipos de feijão-caupi: quatro progenitores (TVu 1190, Pitiúba, CNC 0434 e Alagoano) e uma cultivar (BRS Guariba). Na análise de variância utilizou-se software SAS e as médias foram comparadas pelo teste Tukey. O progenitor Alagoano destacou-se dos demais em eficiência de nodulação, foi superior ao Pitiúba em massa seca de nódulos e diferiu do TVu 1190 em nitrogênio acumulado na parte aérea. Para nitrogênio derivado da fixação biológica verificou-se uma variação média de 49,42 a 65,50%. Nos componentes de produção, o Alagoano se destacou em massa de vagem, comprimento de vagem e massa de 100 sementes. Concluiu-se que os progenitores avaliados são responsivos a FBN e que há indícios de especificidade entre os progenitores e as estirpes de rizóbio. O progenitor Alagoano apresenta resultados mais promissores nos componentes de nodulação e produção.

Among the progenitors, the Alagoano shows most promising results in components of nodulation and production.Due to the variability of leguminous plants as regards their efficiency in the biological fixation of nitrogen (BFN), the importance of plant breeding to improve nitrogen fixation has been emphasized. The objective of this study was to evaluate the symbiotic efficiency of parents of Brazilian cultivars of the cowpea, and give scientific support to the inclusion of BFN in the breeding program for the cowpea crop in Brazil. The experiment was conducted in Teresina, Piauí. An experimental design of randomized blocks in a 5 x 6 factorial arrangement was used, with three replications, consisting of six sources of N: four strains of diazotrophic bacteria (BR 3267, BR 3299, BR 3262 and INPA 03-11B); one treatment with mineral nitrogen and another with no inoculation and no N, and five cowpea genotypes: four parents (TVu 1190, Pitiúba, CNC 0434 and Alagoano) and one cultivar (BRS Guariba). For the variance analysis SAS software was used, and the averages compared by Tukey test. The parent, Alagoano, stood out from the others in nodulation efficiency, was higher than Pitiúba for nodule dry mass, and differed from TVu 1190 in nitrogen accumulation in the shoots. For nitrogen derived from biological fixation there was a mean variation of from 49.42 to 65.50%. As for production components, the Alagoano excelled in pod weight, pod length and 100 seed weight. It was concluded that the evaluated parents are responsive to BFN, and that there is evidence for specificity between the parents and the strains of rhizobia. Of the parents, the Alagoano presents the most promising results for the components of nodulation and production.

O objetivo deste trabalho foi avaliar a compatibilidade do tratamento de sementes com fungicidas e a inoculação com estirpes de Bradyrhizobium em feijão-caupi. Em laboratório, avaliou-se a sobrevivência de células nas sementes da cultivar BRS Guariba, tratadas ou não com fungicidas (fludioxonil, carbendazim, carbendazim + thiram e carboxin + thiram) e inoculadas ou não com Bradyrhizobium (estirpes UFLA3-84, BR 3267, INPA3-11B e BR 3262). Em casa de vegetação, conduziu-se experimento em vasos de Leonard, com os mesmos tratamentos. Foram avaliados: massa de matéria seca da parte aérea, além de número e massa de nódulos 25 dias após a emergência das plantas. No campo, dois experimentos foram conduzidos, tendo-se utilizado a estirpe BR 3262, com aplicação de fungicidas nas sementes: um em área de primeiro cultivo e outro em área cultivada anteriormente com culturas anuais. Avaliaram-se, aos 35 dias, o número de nódulos, a massa de nódulos secos e a massa de matéria seca da parte aérea, e, na colheita, a produtividade de grãos. Os fungicidas não tiveram efeito significativo sobre a sobrevivência de Bradyrhizobium, a nodulação das plantas e o rendimento de grãos, que, em média, foi superior a 1.200 kg ha-1. O tratamento de sementes de feijão-caupi com fungicidas é compatível com a inoculação das estirpes avaliadas.

The objective of this work was to evaluate the compatibility of seed treatment with fungicides and Bradyrhizobium strains inoculation for cowpea. Cell survival on seeds of cultivar BRS Guariba treated or not with fungicides (fludioxonil, carbendazim, carbendazim + thiram, and carboxin + thiram) and inoculated or not with Bradyrhizobium (strains UFLA3-84, BR 3267, INPA3-11B, and BR 3262) were evaluated in laboratory. Another experiment was carried out in greenhouse, in Leonard jars, using the same treatments. The following were evaluated: shoot dry matter weight, besides number and dry matter weight of nodules 25 days after plant emergence. Two experiments were performed in the field with the strain BR 3262, with fungicides on the seeds: one in a first cultivation area and the other in an area previously cultivated with annual crops. Nodule number, nodule dry weight, and shoot dry matter weight were evaluated at 35 days, and grain yield at harvest. Fungicides did not have significant effect on Bradyrhizobium survival, plant nodulation, and grain yield, which was, in average, higher than 1,200 kg ha-1. Cowpea seed treatment with fungicides is compatible with the inoculation of the evaluated strains.

This study evaluated 26 pigeonpea rhizobial isolates according to their cultural characteristics, intrinsic antibiotic resistance, salt and temperature tolerance, carbon source utilization and amylolytic activity. The cultural characterization showed that the majority of them presented the ability to acidify the YMA. Among the 27 isolates evaluated, 25 were able to grow when incubated at 42° C and 11 showed tolerance to 3% (w/v) of NaCl in YMA medium. The patterns of carbon sources utilization was very diverse among the isolates. It was observed the capacity of three strains to metabolize all the carbon sources evaluated and a total of 42% of the bacterial isolates was able to grow in the culture medium supplemented with at least, six carbon sources. The carbon sources mannitol (control) and sucrose were metabilized by all isolates evaluated. The profile of intrinsic resistance to antibiotics showed that the isolates were mostly resistant to streptomycin and ampicillin, but susceptible to kanamycin and chloranphenicol. High amylolytic activity of, at least, four isolates was also demonstrated, especially for isolated 47.3b, which showed the highest enzymatic index. These results indicate the metabolic versatility of the pigeonpea rhizobia, and indicates the isolate 47.3b to further studies regarding the amylase production and characterization.

As espécies de mucuna são muito utilizadas como adubos verdes, e poucas informações estão disponíveis a respeito dos rizóbios nativos capazes de nodulá-las. O objetivo deste trabalho foi avaliar a diversidade e a capacidade simbiótica de isolados bacterianos de nódulos de mucuna-cinza (Mucuna pruriens (L.) DC.) e mucuna-anã (Mucuna deeringiana (Bort.) Merr.). As bactérias foram isoladas de nódulos de mucunas cinza e anã cultivadas em vasos com solos de um sistema de produção agroecológica. Foram isoladas 160 bactérias, sendo 80 de mucuna-anã e 80 de mucuna-cinza, que foram autenticadas e selecionadas para avaliação da capacidade simbiótica. A diversidade dos isolados foi avaliada por meio das características culturais em meio de cultura YMA e da técnica de análise de restrição do produto de PCR do gene 16S rDNA. A inoculação de cinco isolados em mucuna-cinza e dois em mucuna-anã apresentou elevada biomassa da parte aérea. A maioria dos isolados apresentou crescimento rápido e acidificou o meio de cultura. A análise de restrição demonstrou que as bactérias isoladas apresentam baixa similaridade com estirpes de referência, sugerindo a existência de isolados pertencentes a novos grupos, capazes de nodular as mucunas anã e cinza.

Mucuna species are widely used as green manure, and little information is available about rhizobia that can nodulate them. The objective of this study was to evaluate the diversity and ability of bacteria isolated from symbiotic nodules of velvet bean (Mucuna pruriens (L.) DC.) and Mucuna deeringiana (Bort.) Merr. From the nodules of either mucuna species, grown in pots with soils from an agroecological system, 80 bacteria were isolated. The selected bacteria were classified and used to assess the symbiotic capacity. The diversity assessment of the isolates was based on cultural characteristics on YMA medium and the technique of restriction analysis of PCR products of 16S rDNA. The inoculation of Mucuna pruriens with five and Mucuna deeringiana with two isolates increased the aboveground biomass. Most isolates grew rapidly and acidified the culture medium. Restriction analysis showed low similarity of the reference strains with the bacterial isolates, suggesting the existence of isolates belonging to new groups that are able to nodulate the two mucuna species.

O objetivo deste trabalho foi avaliar a estrutura metabólica e genética de comunidades bacterianas em Latossolo de cerrado sob vegetação nativa ou cultivado em sistema de rotação soja/milho sob preparo convencional e plantio direto. Foram utilizadas microplacas EcoPlate para determinar o perfil e a diversidade metabólica das comunidades bacterianas, e eletroforese em gel com gradiente desnaturante (DGGE) para avaliar a estrutura genética. O teste estatístico de Mantel foi utilizado para avaliar a relação entre a estrutura metabólica e a genética. A comunidade bacteriana sob vegetação nativa apresentou perfil metabólico diferente do encontrado em solos cultivados. No solo cultivado com soja sob preparo convencional, o padrão de utilização das fontes de carbono diferenciou-se dos demais tratamentos. Com base nos resultados de DGGE, a comunidade bacteriana sob vegetação nativa apresentou 35% de similaridade com as de áreas cultivadas. Foram formados grupos distintos de comunidades bacterianas do solo entre as áreas sob preparo convencional e plantio direto. Houve correlação significativa de 62% entre as matrizes geradas pelas microplacas EcoPlate e pela DGGE. Variações no perfil metabólico estão relacionadas às variações na estrutura genética das comunidades bacterianas do solo.

The objective of this work was to evaluate the metabolic and genetic structure of bacterial communities in a cerrado Oxisol under native vegetation or cultivated in a soybean/maize rotation system under conventional tillage and no tillage. EcoPlate microplates were used to determine the metabolic profile and diversity of bacterial communities, and denaturing gradient gel electrophoresis (DGGE) to assess the genetic structure. The Mantel statistic test was used to evaluate the relationship between the metabolic and the genetic structure. The bacterial community under native vegetation showed a different metabolic profile than that of cultivated soils. In the area cultivated with soybean under conventional tillage, the carbon source utilization pattern differed from the other treatments. Based on the DGGE results, the bacterial community under native vegetation showed 35% similarity to the ones found under cultivation. Distinct groups of soil bacterial communities were formed between the areas under conventional tillage and no tillage. A significant correlation of 62% was observed between matrices generated by EcoPlate microplates and by DGGE. Variations in the metabolic profile are related to changes in the genetic structure of soil bacterial communities.

O objetivo deste trabalho foi avaliar a eficiência agronômica da formulação polimérica IPC 2.2 como veículo de inoculação para feijão-caupi. Foi avaliada a sobrevivência de células aos 180 dias de armazenamento. Em experimento de campo, comparou-se a formulação polimérica com a turfosa e a líquida e com adubação nitrogenada de 50 e 80 kg ha-¹ de N e um controle absoluto. A formulação IPC 2.2 manteve concentração de células superior a 109 g-1 após 180 dias e, em campo, proporcionou produtividade de grãos superior à do controle absoluto e igual à dos tratamentos nitrogenados e veículos turfoso e líquido.

The objective of this work was to evaluate the agronomic efficiency of the polymer formulation IPC 2.2 as a carrier for inoculation in cowpea. Cell survival at 180 days storage was evaluated. The polymer formulation was compared with peat and liquid carriers and with nitrogen rates of 50 and 80 kg ha-1 N and absolute control, in a field experiment. The formulation IPC 2.2 kept a cell concentration higher than 10(9) g-1, after 180 days and, in the field experiment, it provided a yield higher than that of the treatment absolute and equal to those of the nitrogen treatments and to the peat and liquid carriers.

O objetivo deste trabalho foi avaliar a eficiência agronômica de estirpes de rizóbio para inoculação emcampo,dacultivarBRS Guaribadefeijão-caupi.Os experimentosforamrealizadosemduasáreasemSanta Luzia do Paruá, MA, na região de Pré-Amazônia, em 2009. Foram testadas as estirpes de Bradyrhizobium BR 3299, BR 3262, e INPA 03-11B, um controle absoluto e um tratamento nitrogenado (80 kg ha-1 de N). As avaliações foram realizadas aos 30 e 50 dias após a emergência (DAE) das plantas. Aos 65 DAE, foi avaliada a produtividade de grãos. Foram avaliados número e massa de matéria seca de nódulos, massa de matéria seca da parte aérea, eficiência relativa e acúmulo de N na parte aérea. As estirpes proporcionaram maior número e massa de matéria seca de nódulos, bem como maior produtividade em comparação ao controle absoluto, sem inoculação e sem ureia, nas duas áreas. Para massa de matéria seca da parte aérea, eficiência relativa e acúmulo de N na parte aérea, a estirpe BR 3299 diferiu significativamente do controle absoluto, aos 30 DAE, em uma das propriedades. O feijão-caupi responde positivamente à inoculação com as estirpes, especialmentea BR 3299.

The objective of this work was to evaluate the agronomic efficiency of rhizobia strains for inoculation, in field conditions, of the cowpea BRS Guariba cultivar. Experiments were carried out in two areas in Santa Luzia do Paruá, Maranhão state, Brazil, in the Pre-Amazon region, in 2009. The BR 3299, BR 3262, INPA03-11B Bradyrhizobium strains, an absolute control, and a nitrogen treatment (80 kg ha-1 N) weretested. At30 and 50 days after emergence (DAE) of plants assesments were made. At 65 DAE, grainyield was evaluated. The number and dry weight of nodules, shoot dry mass, relative efficiency, and N accumulation in shoots were evaluated. All strains provided higher values for number and dry weight of nodules, and higher productivity in comparsion to the absolute control, without inoculation and urea, in the two areas. Regarding shoot dry matter, relative efficiency, and N accumulation in shoots, BR 3299 differed significantly from the absolute control, at 30 DAE, in one of the farms. Cowpea yield responds positively to inoculation with strains, especially strain BR 3299.

O objetivo deste trabalho foi avaliar a produção e o comportamento reológico de exopolissacarídeos (EPS) produzidos por rizóbios isolados de nódulos de guandu [Cajanus cajan (L.) Millsp.], e a similaridade desses isolados, pela técnica de análise de restrição do DNA ribossomal amplificado. As bactérias foram cultivadas em meio YM líquido e os EPS foram obtidos por precipitação com etanol gelado a partir do caldo de cultivo. Em seguida, eles foram recuperados por centrifugação, secados a vácuo, pesados e ressuspensos em água para avaliações reológicas. Os três isolados avaliados apresentaram diferenças na produção e na eficiência relativa da produção de EPS, com destaque para o isolado 8.1c. Os EPS demonstraram comportamento não newtoniano e pseudoplástico, porém também apresentaram diferenças na viscosidade aparente em uma mesma taxa de cisalhamento. Na taxa de cisalhamento de 1 s-1, os três isolados foram diferentes, enquanto na taxa de cisalhamento de 40 s-1, os isolados 53.5 e 30.6a2 foram iguais e diferiram do isolado 8.1c. A similaridade dos isolados foi condizente com os resultados de reologia dos EPS, em que bactérias que sintetizaram EPS com menor viscosidade aparente apresentaram maior similaridade

The objective of this work was to evaluate the production and the rheological behavior of exopolysaccharides (EPS) produced by pigeonpea [Cajanus cajan (L.) Millsp.] rhizobia isolates, as well as to evaluate the similarity of these isolates through amplified ribosomal DNA restriction analysis technique. The bacteria were cultured in YM liquid media and the EPS were precipited with cold ethanol from culture broth. After that, they were vacuum dried, weighted, stored and resuspended in water to rheological evaluations. The three evaluated isolates showed differences on the production and productivity of EPS, and the isolate 8.1c stood out, with higher relative efficiency. The evaluated EPS showed non-newtonian and pseudoplastic behavior, however they also presented differences as to the apparent viscosity at the same shear rate. At the shear rate of 1 s-1 all isolates were different, while at the shear rate of 40 s-1 the isolates 53.5 and 30.6a2 were the same, differing from the isolate 8.1c. Genetic similarity of the isolates was in agreement with the rheology results, where bacteria that synthesized EPS with lower apparent viscosity presented the higher similarity