Identification of miRNAs linked to peanut nodule functional processes

microRNAs (miRNAs) are non-coding small RNAs that regulate gene expression at post-transcriptional level.Thousands of miRNAs have been identified in legumes, but studies about miRNAs linked to peanut nodulefunctionality are scarce. In this work we analyzed transcriptional changes in peanut nodules to identifymiRNAs involved in functional processes of these organs. We found 32 miRNAs precursors differentiallyexpressed in nodules compared with roots, and predicted the potential targets of their corresponding maturemiRNAs. Among them, 20 belong to 14 conserved miRNAs families and 12 are Arachis hypogaea-specificmiRNAs. Expression levels of 3 miRNAs (ahy-miR399, ahy-miR159 and ahy-miR3508) were confirmedexperimentally by qPCR. We also demonstrated that the expression of these miRNAs was not affected byinoculation of a biocontrol bacterium or a fungal pathogen. The catalogue of differentially expressed miRNAprecursors and the expression of the corresponding mature miRNA potential targets in the nodules of A.hypogaea obtained in this work is a database of strong candidates, including A. hypogaea-specific miRNAs, forthe regulation of the nodule functionality. The analysis of their role in this process will certainly lead to thecharacterization of essential regulators in these particular aeschynomenoid nodules.

Effect of inoculation of Tarwi plants with Bradyrhizobium spp. strains isolated from a wild lupine, under greenhouse conditions / Efecto de la inoculación de plantas de Tarwi con cepas de Bradyrhizobium spp. aisladas de un lupino silvestre, en condiciones de invernadero

Abstract The "tarwi" or "chocho" (Lupinus mutabilis Sweet) is the unique specie of the genus Lupinus cultivated in America, appreciated for its high protein content in seeds, and the ability to fix nitrogen in symbiotic association with rhizobia. Its genetic variability is reinforced by approximately 84 wild species in Peru. The present investigation was carried out to show if the rhizobia of a wild lupino (Lupinus sp.) might be able to nodulate and promote the growth of tarwi. The root nodules were collected from an abandoned crop field of Huaraz (Ancash-Peru) at 3497 m of altitude, were isolated 8 slow-growing (6-7 days) rhizobial strains, which due to their microbiological and molecular characteristics correspond to the genus Bradyrhizobium. These strains were inoculated in tarwi seeds, and co-inoculated to the 15-day seedlings. It was applied a completely randomized design with 11 treatments (including the control strain LMRT28, N+ with nitrogen fertilization and N- without fertilization) and 5 repetitions. After 70 days of growth in greenhouse conditions, all treatments with native strains, with the exception of LSHZ-L1 and LSHZ-L2, showed reddish-colored root nodules, indicator of leghemoglobin activity. Six of the strains significantly increased the aerial length of the plants respect to the N- and the LMRT28 treatment; also, there were significant differences in relation of aerial dry weight being better in five treatments with native strains. The major foliar coverage was developed by LSHZ-L7; and the greatest number of secondary nodules was showed in LSHZ-L3, which is correlated with the dry weight of the root.

Resumen El "tarwi" o "chocho" (Lupinus mutabilis Sweet) es la única especie cultivada del género Lupinus en América, valorado por su alto contenido proteico y capacidad de fijar nitrógeno en asociación simbiótica con rizobios. Su variabilidad genética está reforzada por aproximadamente 84 especies silvestres en el Perú. La presente investigación se realizó para evidenciar si los rizobios de una especie silvestre de lupino (Lupinus sp.) fueran capaces de nodular y promover el crecimiento del tarwi. Los nódulos radiculares fueron colectados de un campo de cultivo en abandono de Huaraz (Ancash-Perú) a 3497 m de altitud; se aislaron 8 cepas rizobianas de crecimiento lento (6-7 días) que por sus características microbiológicas y moleculares corresponden al género Bradyrhizobium. Estas fueron inoculadas en semillas de tarwi y reinoculadas a plántulas de 15 días. Se aplicó diseño completamente aleatorizado con 11 tratamientos (incluyendo la cepa control LMRT28, N+ con fertilización nitrogenada, y N- sin fertilización) y 5 repeticiones. Después de 70 días en invernadero se observó que todas las cepas nativas, a excepción de LSHZ-L1 y LSHZ-L2, mostraron nódulos radiculares de coloración rojiza indicador de la actividad de la leghemoglobina. Seis de las cepas incrementaron significativamente la longitud aérea de las plantas respecto al N- y al control LMRT28; también hubo diferencias significativas en cuanto al peso seco aéreo destacando cinco cepas nativas; la mayor cobertura foliar fue desarrollada por LSHZ-L7; y la cepa LSHZ-L3 presentó significativamente mayor número de nódulos secundarios y estuvo correlacionada con el peso seco de la raíz.

IMPACT OF CRUDE OIL ON FUNCTIONAL GROUPS OF CULTURABLE BACTERIA AND COLONIZATION OF SYMBIOTIC MICROORGANISMS IN THE Clitoria-Brachiaria RHIZOSPHERE GROWN IN MESOCOSMS / Impacto del petróleo crudo sobre grupos funcionales de bacterias cultivables y colonización de microorganismos simbióticos en la rizosfera de Clitoria-Brachiaria crecidas en mesocosmos

ABSTRACT This research evaluated the changes on populations of culturable N-fixing free bacteria (NFFB) and P-solubilizing bacteria (PSB), as well as on the root nodulation by native rhizobia, the root colonization and spore number of arbuscular mycorrhizal fungi (AMF), in the rhizosphere of Clitoria ternatea and Brachiaria brizantha grown in mesocosms contaminated with crude oil (0, 3000, 6000, 9000, and 12000 mg kg-1), for 240 days. After 24 h of soil contamination, the highest populations of NFFB and PSB (5.5 and 4.9 LogUFC, respectively) were found in control, and the lowest populations were obtained at 12000 mg kg-1 (5.1 and 4.2 LogUFC, respectively). In contrast, at 60 and 240 days, the control showed lower populations of NFFB and PSB (5.4 and 4.8 LogUFC, respectively) than contaminated treatments. The highest number of root nodules in C. ternatea was quantified in control at 60 and 240 days (25 and 27 nodules, respectively) in comparison to those observed at the treatment with 12000 mg kg-1 (7 and 1 nodule, respectively). At 60 days, AMF colonization in both plant species, and the number of spores significantly decreased as the crude oil concentration increased; however, at 240 days, the highest number of AMF spores was recorded at treatments with 6000 and 12000 mg kg-1. The dry weight of both plant species significantly decreased as crude oil concentrations increased. Although C. ternatea was more susceptible to the toxic effects of crude oil, this plant species showed greater content of total chlorophyll than B. brizantha.

RESUMEN Esta investigación evaluó los cambios en la población cultivable de bacterias de vida libre fijadoras de nitrógeno (BVLFN) y de bacterias solubilizadoras de fósforo (BSP), así como en la nodulación de raíces por rizobios nativos, y en la colonización y número de esporas de hongos micorrízicos arbusculares (HMA) en la rizósfera de Clitoria ternatea y Brachiaria brizantha cultivadas en mesocosmos contaminados con petróleo crudo (0, 3000, 6000, 9000 y 12000 mg kg-1), durante 240 días. A las 24 h de la contaminación del suelo, las poblaciones más altas de BVLFN y BSP (5,5 y 4,9 LogUFC, respectivamente) se encontraron en el control, mientras que las poblaciones más bajas se obtuvieron a 12000 mg kg-1 (5,1 y 4,2 LogUFC, respectivamente). En contraste, a los 60 y 240 días, el control mostró bajas poblaciones de BVLFN y BSP (5,4 y 4,8 LogUFC, respectivamente) que los tratamientos contaminados. El mayor número de nódulos en raíz de C. ternatea se cuantificó en el control a los 60 y 240 días (25 y 27 nódulos, respectivamente) en comparación con el tratamiento con 12000 mg kg-1 (7 y 1 nódulos, respectivamente). A los 60 días, la colonización de HMA en ambas especies vegetales y el número de esporas disminuyeron significativamente al aumentar la concentración de petróleo crudo; sin embargo, a los 240 días, se registró el mayor número de esporas de HMA en los tratamientos con 6000 y 12000 mg kg-1. El peso seco vegetal disminuyó significativamente al aumentar las concentraciones de petróleo crudo. Clitoria ternatea fue más susceptible a la toxicidad del petróleo, aunque esta especie vegetal mostró mayor contenido de clorofila total que B. brizantha.

Distribution of adeG, adeB, adeE, adeY, abeM, and adeJ Efflux Pump Genes in Clinical Isolates of Acinetobacter Species from Korea

BACKGROUND: The aim of the present study was to determine the frequency of six efflux pump genes in Acinetobacter clinical isolates collected from South Korean hospitals. METHODS: In this study, we used a total of 339 Acinetobacter strains, comprising 279 Acinetobacter calcoaceticus–Acinetobacter baumannii (ACB) complex and 60 non-ACB complex strains. We performed specific PCR assays to detect adeG, adeB, adeE, adeY, abeM, and adeJ, transporter genes of the multidrug efflux pumps AdeFGH, AdeABC, AdeDE, AdeXYZ, AbeM, and AdeIJK, respectively. RESULTS: Frequencies of six efflux pump genes varied according to the species of Acinetobacter. Frequencies of adeE, abeM, and adeJ between A. baumannii group and A. nosocomialis group were found to be significantly different. Significant differences were found in the frequencies of adeB, adeE, adeY, and adeJ among the susceptible A. baumannii (SAB), multidrug-resistant A. baumannii (MDRAB), and extensively drug-resistant A. baumannii (XDRAB) groups within the 154 strains of A. baumannii. The frequencies of efflux pump genes in imipenem-susceptible and imipenem-nonsusceptible groups were significantly different for adeB, adeY, and adeJ. The frequencies of efflux pump genes in ciprofloxacin-susceptible and ciprofloxacin-nonsusceptible groups were significantly different for adeB and adeY. No significant difference was found in the frequency of efflux pump genes among groups sampled from different regions of Korea, across 86 strains of A. baumannii collected in 2012. CONCLUSIONS: The frequencies of six efflux pump genes obtained in this study demonstrate the fundamental epidemiological feature of efflux pump genes in Korean Acinetobacter clinical isolates.

Draft genome sequence of Bradyrhizobium manausense strain BR 3351T, an effective symbiont isolated from Amazon rainforest

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.

Genetic and phenotypic diversity of Rhizobium isolates from Southern Ecuador / Diversidade genética e fenotípica de isolados de Rhizobium do sul do Equador

ABSTRACT Rhizobium-legume symbioses play relevant roles in agriculture but have not been well studied in Ecuador. The aim of this study was to characterize the genetic and phenotypic diversity of Rhizobium isolates associated with Phaseolus vulgaris from southern Ecuador. Morpho-cultural characterization, biochemical tests and physiological analyses were conducted to authenticate and determine the diversity of bacteria Rhizobium-like isolates. The genetic diversity of the isolates was determined by molecular techniques, which consisted of bacteria DNA extraction and amplification and sequencing of the 16S rRNA gene. The nodulation parameters and nitrogen fixation for P. vulgaris under greenhouse conditions were also assessed to determine the phenotypic diversity among isolates. Furthermore, bacteria indole-acetic-acid production was evaluated by the colorimetric method. Morpho-cultural and biochemical characteristic assessments demonstrated that Rhizobium-like bacteria was associated with the P. vulgaris nodules. The diversity among the isolates, as determined by physiological analyses, revealed the potential of several isolates to grow at different pH values, salinity conditions and temperatures. Partial sequencing of the 16S rRNA gene identified the Rhizobium genus in every sampling site. From a total of 20 aligned sequences, nine species of Rhizobium were identified. Nodule formation and biomass, as well as nitrogen fixation, showed an increase in plant phenotypic parameters, which could be influenced by IAA production, especially for the strains R. mesoamericanum NAM1 and R. leguminosarum bv. viciae COL6. These results demonstrated the efficiency of native symbiotic diazotrophic strains inoculants for legume production. This work can serve as the basis for additional studies of native Rhizobium strains and to help spread the use of biofertilizers in Ecuadorian fields.

RESUMO A simbiose Rhizobium-leguminosa desempenha um relevante papel na agricultura, entretanto não tem recebido suficiente atenção de estudos científicos no Equador. O objetivo deste artigo foi caracterizar a diversidade genética e fenotípica de isolados de Rhizobium associados com Phaseolus vulgaris do sul do Equador. A caracterização morfo-cultural, testes bioquímicos e análises fisiológicas foram realizados para autenticar e determinar a diversidade de isolados de bactérias Rhizobium. A diversidade genética foi determinada por por técnicas moleculares consistindo na extração de DNA genômico bacteriano, amplificação e sequenciamento parcial do gene 16SrRNA; e parâmetros de nodulação e fixação de nitrogênio de P. vulgaris sobre condições de estufa foram testados para determinar a diversidade fenotípica entre os isolados. Além disso, a produção de ácido indolacético foi avaliada por um método colorimétrico. A análise fisiológica da diversidade entre os isolados revelou o potencial de crescimento de diversos isolados em diferentes níveis de pH, salinidade e temperatura. O sequenciamento parcial do gene 16S rRNA mostrou o gênero Rhizobium em todas os locais de amostragem. De um total de 20 sequências alinhadas, 9 espécies de Rhizobium foram identificadas. A formação de nódulos e biomassa, bem como a fixação de nitrogênio mostraram um aumento nos parâmetros fenotípicos das plantas, os quais devem ser influenciados pela produção de IAA, especialmente pelas cepas R. mesoamericanum NAM1 e R. leguminosarum bv. viciae COL6. Estes resultados demonstram a eficiência de cepas diazotróficas simbióticas nativas para produção de inoculantes para leguminosas e fornece informações valiosas e uteis para a agricultura sustentável equatoriana. Neste sentido, este trabalho deve ser um elemento essencial para a realização de futuras pesquisas aplicadas relacionadas a cepas Rhizobium nativas e espalhar o uso de biofertizantes em campos equatorianos.

Soil management and application of agricultural gypsum in a Planosol for soybean cultivation / Manejo do solo e aplicação de gesso agrícola em Planossolo para o cultivo de soja

ABSTRACT: This study investigated the effects of soil management systems, tillage, and application of gypsum agricultural to soil, on soybean development in lowland areas. The experiment was carried out on an Alfisol in a randomized complete block design in a factorial arrangement. The two soil tillage practices were without deep tillage and with deep tillage. Gypsum treatments were no gypsum application, 500kg of gypsum ha-1, 1000kg of gypsum ha-1, and 1500kg of gypsum ha-1. Deep tillage resulted in less soil resistance to root penetration during ryegrass cultivation during the soybean offseason, 11 months after applying the management treatments, resulting in higher dry mass of ryegrass in the offseason and higher soybean yield in the following year.

RESUMO: O trabalho visou identificar o efeito de dois sistemas de manejo do solo, com e sem escarificação, associado à aplicação de gesso agrícola no desenvolvimento de plantas de soja em áreas de terras baixas. O trabalho foi realizado a campo, em Planossolo Háplico Eutrófico arênico, em esquema bifatorial (2x4) no delineamento experimental de blocos ao acaso composto por dois sistemas de manejo do solo: sistema sem escarificação do solo e sistema com escarificação do solo. Os tratamentos com gesso agrícola foram: testemunha sem aplicação de gesso agrícola, 500kg ha-1 de gesso agrícola, 1000kg ha-1 de gesso agrícola e 1500kg ha-1 de gesso agrícola. A escarificação do solo proporciona maior rendimento de grãos de soja em áreas de terras baixas com presença de camada compactada, independente do uso de gesso agrícola. Os efeitos da escarificação do solo resultam na menor resistência à penetração de raízes no final do cultivo do azevém na entressafra da soja, 11 meses após o manejo, resultando em maior massa seca do azevém em sucessão e produtividade da soja no ano seguinte.

Genome sequence of Bradyrhizobium embrapense strain CNPSo 2833T, isolated from a root nodule of Desmodium heterocarpon

Abstract Bradyrhizobium embrapense CNPSo 2833T is a nitrogen-fixing symbiont of the legume pasture Desmodium. Its draft genome contains 8,267,832 bp and 7876 CDSs. The symbiotic island includes nodulation and nitrogen fixation genes resembling the operon organization of B. japonicum. Several CDSs related to secretion proteins and stress tolerance were also identified.

Nitrogen-fixing bacteria and arbuscular mycorrhizal fungi in Piptadenia gonoacantha ( Mart. ) Macbr

Abstract The family Leguminosae comprises approximately 20,000 species that mostly form symbioses with arbuscular mycorrhizal fungi (AMF) and nitrogen-fixing bacteria (NFB). This study is aimed at investigating and confirming the dependence on nodulation and biological nitrogen fixation in the specie Piptadenia gonoacantha (Mart.) Macbr., which belongs to the Piptadenia group. Two consecutive experiments were performed in a greenhouse. The experiments were fully randomized with six replicates and a factorial scheme. For the treatments, the two AMF species and three NFB strains were combined to nodulate P. gonoacantha in addition to the control treatments. The results indicate this species’ capacity for nodulation without the AMF; however, the AMF + NFB combinations yielded a considerable gain in P. gonoacantha shoot weight compared with the treatments that only included inoculating with bacteria or AMF. The results also confirm that the treatment effects among the AMF + NFB combinations produced different shoot dry weight/root dry weight ratios. We conclude that AMF is not necessary for nodulation and that this dependence improves species development because plant growth increases upon co-inoculation.

Draft genome sequence of Bradyrhizobium sp. strain BR 3262, an effective microsymbiont recommended for cowpea inoculation in Brazil

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.

Draft genome sequence of Bradyrhizobium sp. strain BR 3267, an elite strain recommended for cowpea inoculation in Brazil

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.

Symbiotic potential and survival of native rhizobia kept on different carriers

Native rhizobia are ideal for use as commercial legume inoculants. The characteristics of the carrier used to store the inoculants are important for the survival and symbiotic potential of the rhizobia. The objective of this study was to investigate the effects of peat (PEAT), perlite sugarcane bagasse (PSB), carboxymethyl cellulose plus starch (CMCS), and yeast extract mannitol supplemented with mannitol (YEMM) on the survival, nodulation potential and N₂ fixation capacity of the native strains Sinorhizobium mexicanum ITTG R7^T and Rhizobium calliandrae LBP2-1^T and of the reference strain Rhizobium etli CFN42^T. A factorial design (4 × 3) with four repetitions was used to determine the symbiotic potential of the rhizobial strains. The survival of the strains was higher for PEAT (46% for strain LBP2-1^T, 167% for strain CFN42^T and 219% for strain ITTG R7^T) than for the other carriers after 240 days, except for CFN42^T kept on CMCS (225%). All the strains kept on the different carriers effectively nodulated common bean, with the lowest number of nodules found (5 nodules) when CFN42^T was kept on CMCS and with the highest number of nodules found (28 nodules) when ITTG R7^T was kept on PSB. The nitrogenase activity was the highest for ITTG R7^T kept on PEAT (4911 μmol C₂H₄ per fresh weight nodule h⁻¹); however, no activity was found when the strains were kept on YEMM. Thus, the survival and symbiotic potential of the rhizobia depended on the carrier used to store them.

Interrelationships between Bacillus sp. CHEP5 and Bradyrhizobium sp. SEMIA6144 in the induced systemic resistance against Sclerotium rolfsii and symbiosis on peanut plants.

Plant-growth-promoting bacteria are often used to enhance crop yield and for biological control of phytopathogens. Bacillus sp. CHEP5 is a biocontrol agent that induces systemic resistance (ISR) in Arachis hypogaea L. (peanut) against Sclerotium rolfsii, the causal agent of root and stem wilt. In this work, the effect of the co-inoculation of Bacillus sp. CHEP5 and the peanut nodulating strain Bradyrhizobium sp. SEMIA 6144 was studied on induction of both systemic resistance and nodulation processes. Bradyrhizobium sp. SEMIA 6144 did not affect the ability of Bacillus sp. CHEP5 to protect peanut plants from S. rolfsii by ISR and the priming in challenged-plants, as evidenced by an increment in phenylalanine ammonia-lyase enzyme activity. Additionally, the capacity of Bradyrhizobium sp. SEMIA 6144 to induce nodule formation in pathogen-challenged plants was improved by the presence of Bacillus sp. CHEP5.

Role of arbuscular mycorrhiza fungi on tolerance to salinity of the tree legume Albizia lebbeck (L.) inoculated by Rhizobium.

Myrica esculenta (Myricaceae) and Syzygium cumini(Myrtaceae), the Indian traditional fruits The effect of different level of salinity on growth, nodulation and Nitrogen fixation of single and dual inoculated tree legumes which help arbascular mycorrhizal fungi (AMF) Acaulospora laevis Gerd. and Trappe with Rhizobium bacteria in the presence of different levels of salinity (concentration of NaCl) conducted in earthen pots in a completely randomized block design with three replications resulted in effective plant growth, shoot and root biomasses, nodulation and N2 fixation. The effect of different levels of salinity (concentration of NaCl) on growth, nodulation and N2 fixation of single (Rhizobium or AMF alone) and dual inoculated (Rhizobium + AMF ) tree legume has a pot culture experiment using The parameter growth , nodulation, nitrogen fixation and % AMF colonization of roots were considerable influenced with the increase in salt concentration from 0.5 % to 4.0 % NaCl. It was observed that the mycorrhizal tree legume (inoculated with the most preferred AMF Acaulospora laevis) performed better in the increasing levels of salinity in comparison to non-mycorrhozal ones. These investigations suggested a protective role play by AMF in providing resistance to the tree legume against injurious effects of salinity. Inoculation of efficient strain of AMF (Acaulospora laevis) during the course of study, prevented the injurious effects of salinity in the test plants due to enhanced water and sustainable nutrient uptake thereby promoting growth, nodulation and biogeochemical N2 cycle (fixation of nitrogen) of the tree legume under investigation.

Effect of resistance nodulation division efflux pumps on reduced susceptibility to tigecycline in carbapenem-resistant Acinetobacter baumanii / 中华临床感染病杂志

Objective To investigate the effect of resistance nodulation division (RND) efflux pumps on reduced susceptibility to tigecycline in carbapenem-resistant Acinetobacter baumanii.Methods Totally 631 isolates of Acinetobacter baumanii were collected from 16 hospitals in 7 provinces in 2010.Genes oxa-51 and oxa-23 were detected by PCR method,and the ST profiles were determined by multilocus sequence typing (MLST).The disk susceptibility assay was used to determine the inhibition zone diameters of β-lactams,aminoglycosides,macrolides,tetracyclines,carbapenems,tigecycline and polymyxin.The minimum inhibitory concentration (MIC) of tigecycline was determined by E-test in strains with inhibition zone diameters ≤ 12 mm on tigecycline.The expression of operon genes adeB,adeG and adeJ was determined with efflux pump inhibitor NMP (N-methyl-2-pyrrolidone) for detection of efflux pump inhibitor phenotype.The isolates of Acinetobacter baumanii which were resistant both to tigecycline and carbapenems and with the inhibited phenotype of efflux pump inhibitor were collected as the experiment group,the isolates which were susceptible to tigecycline but resistant to carbapenems were collected as the control group,and ATCC 19606 was used as the reference strain.The expressions of adeABC,adeFGH and adeIJK were quantified by q-PCR at the transcriptional level.Genes adeR,adeS and adeL were amplified and sequenced using PCR method to find polymorphic locus and insertion sequences.Results There were 32 isolates of Acinetobacter baumanii with reduced susceptibility to tigecycline and carbapenem-resistant.Eight isolates were with the inhibited phenotype by efflux pump inhibitor.And 4 strains which were susceptible to tigecycline but resistant to carbapenems were selected as the control.The expressions of adeABC in A518,Z1219 and A527 of experiment group were 13-fold,5-fold and 7-fold higher than reference strain ATCC19606,respectively.The expressions of adeFGH and adeIJK were up-regulated slightly in some isolates.Transcript of adeABC was not found in control group strains A207 and A1731,and the expressions of adeABC,adeFGH and adeIJK were not up-regulated in other isolates.The single nucleotide polymorphism (SNP) was detected in adeR (E220K) and adeS (A130D),respectively.ISAab1 insertion sequence was identified in adeS of adeABC-over expressed isolates.No mutation was found in adeL.Conclusion High expression of adeABC pump may play an important role in tigecycline resistance in carbapenem-resistant Acinetobacter baumannii,mainly due to the insertion of ISAba1 sequence in its regulator gene adeS,but other mechanism of tigecycline resistance may not be excluded.

Resposta de cultivares de feijoeiro-comum à inoculação das sementes com duas estirpes de rizóbio / Common bean cultivars response to seed inoculation with two rhizobium strains

Fixação simbiótica de nitrogênio pode ser uma alternativa para reduzir, ou mesmo substituir, a adubação nitrogenada do feijoeiro-comum, cultura na qual ainda há descrédito quanto aos benefícios da inoculação e que se caracteriza por aplicações de altas doses de N mineral. Com o objetivo de verificar a resposta de cultivares de feijoeiro-comum à inoculação das sementes com duas estirpes de rizóbio, foram conduzidos dois experimentos de campo na safra de inverno 2010 (Patos de Minas e Uberaba, MG). Utilizou-se o delineamento em blocos casualizados, com três repetições e esquema fatorial 8 x 3, envolvendo oito cultivares (União, Madrepérola, Supremo, Ouro Negro, Ouro Vermelho e Majestoso) e três tipos de inoculação (sementes não inoculadas e estirpes CIAT 899T e UFLA 04-173). Na floração, avaliaram-se o número e massa seca de nódulos, massa seca de parte aérea e teor e acúmulo de nitrogênio (N) na parte aérea e, na colheita, o rendimento de grãos com seus componentes primários, teor e acúmulo de N nos grãos. Constatou-se que as populações nativas promovem nodulação equivalente à das estirpes inoculadas. As estirpes CIAT 899T e UFLA 04-173 favorecem o crescimento vegetativo do feijoeiro. A estirpe CIAT 899T proporciona maior número de vagens e maior massa de grãos que a estirpe UFLA 04-173, e em Uberaba também proporciona maior rendimento de grãos. Quando a inoculação é realizada com a estirpe UFLA 04-173, as cultivares diferem quanto ao teor foliar de N. As cultivares Madrepérola, Ouro Negro e Ouro Vermelho apresentam mais nódulos e maior massa seca de parte aérea, além de maior produtividade em Patos de Minas. Ouro Negro situa-se entre as mais produtivas em Uberaba. Em Uberaba há maior acúmulo de matéria seca de nódulos e de parte aérea, mas o ambiente Patos de Minas resulta em maiores valores do rendimento de grãos e componentes.

The symbiotic nitrogen fixation can be an alternative to reduce, or even replace, the nitrogen (N) fertilization of the common bean crop, characterized by application of high doses of this nutrient. This work aimed to verify the response of common bean cultivars to the seed inoculation with two Rhizobium strains, conducting two field experiments in the winter-spring crop of 2010, at Patos de Minas and Uberaba, Minas Gerais State, Brazil. The experiment had a randomized block design, with three replications and a 8 x 3 factorial squemme, involving eight cultivars and three levels of inoculation. At flowering, it was evaluated the number of nodules, nodules dry mass, shoots dry weight, N content and accumulation in shoots and, at harvest, the grain yield and its primary components (100 grains weight, number of pods per plant, number of grains per pod) and the N content and accumulation in the grains. The Madrepérola, Ouro Negro and Ouro Vermelho cultivars present the highest number of nodules and higher dry mass; in Patos de Minas, the same cultivars show higher productivity, but in Uberaba, only Ouro Negro cultivar is among the most productive. In Uberaba there is the greater nodules dry weight and higher shoots dry matter, but the most favorable conditions in Patos de Minas results in higher values for grain yield and its components. In Uberaba, the CIAT 899T strain overcomes the UFLA 04-173 strain in relation to grain yield. In both environments, the CIAT 899T strain also shows superiority in number of pods per plant and weight of one hundred grains.

Dinâmica de rizóbios em solo do cerrado de Roraima durante o período de estiagem / Dynamic of rhizobia in the soil during the dry season in cerrado of Roraima

The biological nitrogen fixation in legumes is performed by a group of bacteria known as rhizobia. The survival of these bacteria in soils is affected by several factors, such as temperature, drought and soil fertility. This study was performed to evaluate the dynamics of rhizobia in the soil after soybean cultivation and during a dry season in the cerrado of Roraima. Three areas were sampled: i) native cerrado as reference; ii) an area previously cultivated with soybean for one season; and iii) another one cultivated for two seasons also with soybean. The soil was sampled at a depth of 0-10 cm in five times (0, 45, 90, 135 and 180 days) during the dry season (September 2006 to March 2007). The rhizobial density in the soil was evaluated by the most probable number method with infection of soybean and cowpea plants. It was observed very low number of soybean nodulating bacteria in the reference area, but a high density, of up to several hundred rhizobia capable to nodulate cowpea was measured in this same area. Cropping of soybean with inoculated seeds increased rhizobial density evaluated by both trapping hosts. In cropped areas, an intense reduction of rhizobium density was observed just after soybean harvest, and this reduction continued until the end of the period of evaluation. It was concluded that soybean cultivation increases the density of rhizobial in the cerrado soil; however, this density is drastically reduced, during the dry season, by 99% at the end of the dry period.

A fixação biológica de nitrogênio que ocorre em leguminosas é realizada por um grupo de bactérias conhecidas como rizóbios. A sobrevivência destas bactérias no solo é influenciada por diversos fatores como a temperatura, umidade e fertilidade do solo. O objetivo deste trabalho foi avaliar a dinâmica da população de rizóbios em solo após o cultivo de soja, durante o período de estiagem no cerrado de Roraima. Foram amostradas três áreas: i) cerrado nativo como referência; ii) área cultivada uma vez com soja inoculada com rizóbio; iii) e área cultivada duas vezes com soja inoculada com rizóbio em anos consecutivos. O solo foi coletado na profundidade de 0-10 cm em cinco períodos a partir do inicio da estiagem no mês de setembro de 2006 coincidindo com a época de colheita da soja e prolongando-se até março de 2007 (0, 45, 90, 135 e 180 dias). A população de rizóbios no solo foi avaliada pela técnica do número mais provável (NMP) utilizando plantas de soja e de feijão-caupi como espécies isca. Foi observado que na área nativa praticamente não existiam bactérias nodulantes de soja, mas havia uma população capaz de nodular o feijão-caupi de até algumas centenas de rizóbios por gramas de solo. O cultivo da soja utilizando sementes inoculadas elevou a população de rizóbios no solo que foi constatada por ambas às espécies de plantas isca. Nas áreas cultivadas, constatou-se uma intensa redução da população de rizóbios no solo, em especial logo após a colheita da soja, continuando o decréscimo até o último período de avaliação. Conclui-se que o cultivo da soja inoculada com rizóbio eleva a densidade de rizóbios em solo do cerrado, mas durante a estiagem ocorre uma drástica redução dessa população, que pode chegar a mais de 99% considerando o início e final do período.

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.

Desempenho de cultivares transgênicas de soja em sucessão a culturas de inverno em semeadura direta / Performance of transgenic soybean cultivars in succession to winter cover crops under no tillage system

Objetivou-se com esse trabalho avaliar o desempenho de cultivares de soja transgênica em sucessão a culturas de inverno na implantação do sistema de semeadura direta. O delineamento experimental utilizado foi o de blocos casualizados, em esquema de parcelas subdivididas, com quatro repetições. As parcelas foram representadas por cinco espécies cultivadas no inverno, aveia branca, nabo forrageiro, cevada, trigo e ervilha forrageira e área de pousio. As subparcelas foram constituídas por seis cultivares de soja (BRS 243 RR, BRS 244 RR, BRS 245 RR, BRS 247 RR, BRS 255 RR e BRS 256 RR). Para as características agronômicas e os componentes de produção da cultura da soja indicou que a interação culturas de inverno x cultivares de soja foi significativa para a população final de plantas, número de nódulos por planta, massa de matéria seca de nódulos por planta, número de vagens por planta, número de grãos por vagem e produtividade de grãos. O tratamento de pousio proporcionou os menores valores de nodulação da cultura da soja. Para produtividade de grãos, houve diferenças entre as culturas de inverno antecessoras para o cultivar BRS 243 RR, sendo que, a aveia proporcionou maiores valores.

The objective of this research was to evaluate crop yield and some characteristics and yield components of transgenic soybean cultivars sown after different winter cover crops in the first year under no tillage system. The experimental design was the completely randomized block with split plots and four replications. The main plots consisted of five winter cover crops, white oat (Avena sativa L.), forage turnip (Raphanus sativus L.), barley (Hordeum vulgare L.), wheat (Triticum aestivum L.) and ground pea (Pisum sativum L.) and an area under fallow (spontaneous vegetation). The subplots consisted of six soybean cultivars (BRS 243 RR, BRS 245 RR, BRS 247 RR, BRS 255 RR, BRS 256 RR and BRS 244 RR). Variance analysis for agronomic characteristics showed that soybean yield components were influenced by the interaction between winter crop and soybean cultivar. Thus, final population, number of nodes and pods per plant, nodes dry matter per plant, number of grains per pod and grain yield were affected significantly. When soybean nodulation was evaluated, the treatment with the area under fallow showed lower values. There was difference among winter crops for BRS 243 RR grain yield, white oat showed the highest values.

Nodulation and N2 fixation effectiveness of Bradyrhizobium strains in symbiosis with Adzuki Bean, Vigna angularis

In pot experiment, one isolate Knj from a Serbian soil, four strains of Bradyrhizobium japonicum and three strains of Bradyrhizobium spp. were examined for the effect on adzuki bean nodulation and effectiveness in symbiotic N2 fixation. All the tested strains produced root nodules in adzuki bean. Strains of B. japonicum showed high potential of N2 fixation, particularly 525 and 542. B. japonicum strains resulted 65-71 percent shoot dry weight and 99-138 percent total N content of uninoculated control with full N content (100 percent). No significant difference was found between the plants inoculated with Bradyrhizobium spp. strains and uninoculated control plants without N (40-42 and 42 percent shoot dry weight, respectively), which indicated symbiotic N2 fixation inactivity of the Bradyrhizobium spp. strains. Knj strain had the middle position (56 percent shoot dry weight). These data showed that B. japonicum 525 and 542 strains could be used in further investigations in order to apply them as inoculants in microbiological N fertilizers.