Resumen La diversidad de aislamientos rizobianos en el Bosque Seco Tropical de Brasil (BST) (Caatinga) ha sido ampliamente estudiada debido a la gran importancia de encontrar nuevas especies de bacterias con capacidad para fijar nitrógeno en leguminosas arbóreas, entre ellas, Mmosa tenuiflora (Willd.) Poir., Piptadenia stipulacea (Benth.) Ducke y Mimosa caesalpiniifolia Benth. El bioma del BST es importante en Brasil, por tanto, el conocimiento de comunidades microbianas que hacen simbiosis con plantas autóctonas ayuda en la comprensión de la interacción planta-microorganismos, así como en la optimización de procesos biológicos que pueden mejorar el sistema de cultivo en un área como ésta, rica en diversidad y que ha sido poco estudiada. Este estudio se realizó en tres municipios del Sertáo de Pernambuco y Paraíba, con las especies antes citadas. Para el efecto, se realizaron las mediciones siguientes: aislamiento de rizobios y su caracterización fenotípica y molecular con el uso de marcadores de fingerprint como BOX, REP, ERIC y BOX-PCR; y confirmación de los aislamientos, utilizando la técnica de amplificación por dúplex-PCR con los genes niJH y nodC. Los resultados mostraron que dada la variación encontrada en la amplificación de los genes niJH y nodC, la técnica duplex-PCR puede mostrar resultados falsos positivos, ya que estos genes tienen un polimorfismo muy grande. La falta de conocimiento de los aislamientos que hacen simbiosis con estas plantas ayuda, aún más, a concluir que esta técnica de autenticación todavía no puede ser aplicada a todos los aislados nodulantes de leguminosas. En cuanto a los análisis realizados con marcadores de fingerprint (huellas digitales), se observó que todos eran muy eficientes en la capacidad de distinguir especies y que cuanto mayor es el número de marcadores utilizados, más seguro es el conocimiento sobre la taxonomía y diversidad de rizobios.

Abstract The diversity of rhizobial isolates in tropical dry forests (TDF) has been studied due to the great importance of finding new species of bacteria capable of fixing nitrogen in tree legumes. In the Brazilian TDF (caatinga), Leguminosae is the most important family of plants, so the knowledge of microbial communities that make symbioses with native plants can help in the understanding of the interaction plant-microorganisms, as well as in the optimization of biological processes that can improve the cultivation system in an area so rich and understudied. In this study was determined the characteristics of rhizobia isolated from nodules of Mimosa tenuiflora (Willd.) Poir., Piptadenia stipulacea (Benth.) Ducke and Mimosa caesalpiniifolia Benth, grown in soils collected under caatinga vegetation. The phenotypic and molecular characterization of the isolates, with use of fingerprint markers such as BOX, REP, ERIC and BOX-PCR were performed. Amplification technique by duplex PCR with the niJH and nodC genes was used for the authentication of isolates. The results showed that given the variation found in the amplification of the niJH and nodC genes, the duplex PCR technique can show false-positive results, as these genes have a very large polymorphism. The lack of knowledge of isolates that make symbioses with these plants further help to conclude that this authentication technique cannot yet be applied to all legume-nodulating isolates. Regarding the analyses made with fingerprint markers, it was observed that all were very efficient in the ability to distinguish species and that the greater the number of markers used, the safer the knowledge on the taxonomy and diversity of rhizobia.

Abstract Erythrina velutina ("mulungu") is a legume tree from Caatinga that associates with rhizobia but the diversity and symbiotic ability of "mulungu" rhizobia are poorly understood. The aim of this study was to characterize "mulungu" rhizobia from Caatinga. Bacteria were obteined from Serra Talhada and Caruaru in Caatinga under natural regeneration. The bacteria were evaluated to the amplification of nifH and nodC and to metabolic characteristics. Ten selected bacteria identified by 16S rRNA sequences. They were tested in vitro to NaCl and temperature tolerance, auxin production and calcium phosphate solubilization. The symbiotic ability were assessed in an greenhouse experiment. A total of 32 bacteria were obtained and 17 amplified both symbiotic genes. The bacteria showed a high variable metabolic profile. Bradyrhizobium (6), Rhizobium (3) and Paraburkholderia (1) were identified, differing from their geographic origin. The isolates grew up to 45 °C to 0.51 mol L-1 of NaCl. Bacteria which produced more auxin in the medium with l-tryptophan and two Rhizobium and one Bradyrhizobium were phosphate solubilizers. All bacteria nodulated and ESA 90 (Rhizobium sp.) plus ESA 96 (Paraburkholderia sp.) were more efficient symbiotically. Diverse and efficient rhizobia inhabit the soils of Caatinga dry forests, with the bacterial differentiation by the sampling sites.

ABSTRACT Cowpea is a very important crop to Brazilian Semi-Arid mainly small family-based farmers. Rhizobia inoculation is a practice, easy to use, and cheap technology that increases cowpea productivity. The aim of this study was to evaluate the efficiency of two new rhizobia isolates in greenhouse and field as well as classify them taxonomically. To bacterial identification the 16S rRNA gene of ESA 17 and ESA 18 isolates were sequenced. The greenhouse test was conducted with pots containing 3 L of soil and the bacterial isolates evaluated were ESA 17, ESA 18, BR 3267 or BR 3262 strains. A field experiment was implemented on a Vertisol in Juazeiro, Bahia State, to evaluate the cowpea growth and productivity. In this experiment, the peat-based inoculants with ESA 17, ESA 18, BR 3267 or UFLA 3-84 were used in 2 cowpea cultivars. Both bacteria were identified as Bradyrhizobium, but related to different species. ESA 17 was related to B. japonicum and ESA 18 was closer to B. pachyrhizi. At greenhouse, both isolates increased cowpea nitrogen content in the shoots due to the presence of very efficient nodules. In the field, the isolate ESA 18 inoculated at BRS Pujante cultivar induced higher production than observed for the absolute control, and for BR 17 Gurguéia cultivar, the ESA 17 and BR 3267 stood out both by inducing high production and grain protein content. The results indicate that both isolates can be evaluated in network experiments aiming at official recommendation for new bacteria to cowpea inoculant in Brazil.

O objetivo desse estudo foi avaliar as características fenotípicas dos isolados de nódulos de mulungu (Erythrina velutina Willd.) e determinar seu fingerprinting molecular por meio de Box-PCR. Todas as bactérias foram avaliadas pelas seguintes características fenotípicas: tempo de crescimento, alteração de pH, cor da colónia e produção de muco. As bactérias isoladas hábeis em renodular o hospedeiro original também foram avaliadas em relação à tolerância ao aumento da salinidade e da temperatura de incubação, capacidade de crescimento utilizando diferentes fontes de carbono, resistência intrínseca aos antibióticos e biossíntese de auxina “in vitro”. O fingerprint molecular foi estabelecido usando a técnica Box-PCR e os isolados foram agrupados pelos perfis obtidos. Dos 22 isolados obtidos, oito foram hábeis em renodular o hospedeiro original. Entre eles, alguns foram tolerantes a 1% de NaCl e a 39° C e todos metabolizaram as fontes maltose, frutose, glicose, sacarose e arabinose, resistiram à rifampicina e produziram auxina. As bactérias isoladas mostraram baixa similaridade genética entre elas e as estirpes de referência, o que indica a grande variabilidade genética dos isolados. Os resultados deste trabalho constituem o primeiro relato sobre bactérias capazes de nodular esta espécie endémica. O estudo mais aprofundado destas bactérias poderá revelar a existência de isolados tolerantes a stresses ambientais e adequados a serem utilizados como futuros inoculantes de mulungu.

This study aims to evaluate the phenotypical characteristics of bacterial isolates from mulungu (Erythrina velutina Willd.) nodules and determinate their Box-PCR fingerprinting. All bacteria were evaluated by the following phenotypic characteristics: growth rate, pH change, colony color and mucus production. The bacterial isolates able to re-nodulate the original host were also evaluated regarding its tolerance to increased salinity and different incubation temperatures, ability to growth using different carbon sources, intrinsic antibiotic resistance and “in vitro” auxin biosynthesis. The molecular fingerprints were set up using the Box-PCR technique and the isolates were clustered by its profiles. Among the 22 bacterial isolates obtained, eight were able to re-nodulate the original host. Among the nodule inducing isolates, some were tolerant to 1% of NaCl and 39° C and all of them metabolized the maltose, fructose, glucose, sucrose and arabinose, were resistant to rifampicin and produced auxin. The bacteria showed low genetic similarity among them and reference strains, which indicates the great genetic variability of the isolates. The results of this work are the first reports about the bacterial isolates able to nodulate this endemic specie. A more deep study of these bacteria may reveal the existence of isolates tolerant to environmental stresses and suitable as a future mulungu inoculant.