Community analysis of diazotrophic bacteria associated with elephant grass

Autor: Videira, Sandy Sampaio
Přispěvatelé: Baldani, Vera L?cia Divan, Caballero, Segundo Sacramento Urquiaga, Ara?jo, Jean Luiz Sim?es de, Santos, Leandro Azevedo, Alves, Bruno Jos? Rodrigues, Zilli, Jerri Edson, Goi, Silvia Regina, Xavier, Deise Ferreira
Jazyk: portugalština
Rok vydání: 2012
Předmět:
Zdroj: Biblioteca Digital de Teses e Dissertações da UFRRJ
Universidade Federal Rural do Rio de Janeiro (UFRRJ)
instacron:UFRRJ
Popis: Submitted by Sandra Pereira (srpereira@ufrrj.br) on 2020-06-18T22:19:09Z No. of bitstreams: 1 2012 - Sandy Sampaio Videira.pdf: 6638879 bytes, checksum: f8948f450c10e31be903f25ddc06684f (MD5) Made available in DSpace on 2020-06-18T22:19:09Z (GMT). No. of bitstreams: 1 2012 - Sandy Sampaio Videira.pdf: 6638879 bytes, checksum: f8948f450c10e31be903f25ddc06684f (MD5) Previous issue date: 2012-02-15 Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior, CAPES, Brasil. In this study, a fraction of the diazotrophs community associated with roots and shoots of different elephant grass genotypes (Pennisetum purpureum) was assessed by combined use of culture-dependent and -independent approaches. A total of 204 bacterial strains were isolated from Cameroon and CNPGL91F06-3 genotypes and genomically fingerprinted by BOX-PCR. From the cluster analysis generated from BOX-PCR profiles, 47 strains were selected for identification by partial 16S rRNA sequencing. Similarity analysis of the 16S rRNA gene fragments revealed that 36% of the sequences belonged to the Gluconacetobacter genus, 31% to Azospirillum spp and 21% to Enterobacter spp. These results were confirmed by nifH gene analysis, although bacteria identified as Gluconacetobacter showed sequences homologous to nifH gene from Enterobacter spp. Additionally, 204 strains were investigated for their ability to fix nitrogen, produce phytohormones and phosphate solubilization. Of the total, 75.5% had nitrogenase activity, 97% produced indole compounds, 22% solubilized phosphate and 15% showed all three characteristics together. To evaluate, under vessel experiment conditions, the response of Cameroon and Roxo genotypes to inoculation with Azospirillum, Klebsiella, Enterobacter and Gluconacetobacter strains previously characterized. The biomass yield, nutrient accumulation in the shoot as well as the protein content in elephant grass inoculated with diazotrophs were not significantly improved by inoculation, although increases in root dry weight and N and P have been detected. As the benefits to the host plant showed variation from strain to strain, and the analyzes were carried out punctually at 60 days after planting, it was not possible to determine if the inoculated strains were not able to penetrate into the tissues and which factors influenced the plant-bacterium interaction and the final production of the plants. The structural and functional diversity was assessed by PCR-DGGE from 16S rRNA and nifH sequences directly-obtained from root and stem of five elephant grass genotypes. The bacterial and diazotrophic community structure was more influenced by plant tissue than the genotypes, the root being considered a more complex niche in relation to the stem. The metabolically active bacterial population was identified in three of the five tested genotypes, and the groups most frequently detected were Proteobacteria, consisting of - (Leptothrix spp and Burkholderia spp), - (Bradyrhizobium spp, Methylobacterium spp and Rhizobium spp), - (Steroidobacter spp) and Actinobacteria (Actinoplanes spp, Conexibacter spp, Solirubrobacter spp and Amycolatopsis spp). In nifH-cDNA libraries, 26.4% of the fragments were related to different Bradyrhizobium spp. strains sequences. Less abundant sequences belonging to the genera Azospirillum, Burkholderia, Klebsiella and Enterobacter spp were also detected, but their distribution among the samples was at random. Neste estudo, uma fra??o da comunidade de bact?rias diazotr?ficas associada a ra?zes e parte a?rea de diferentes gen?tipos de capim-elefante (Pennisetum purpureum) foi analisada por m?todos dependentes e independentes de cultivo. Duzentas e quatro estirpes bacterianas foram isoladas dos gen?tipos Cameroon e CNPGL91F06-3 e caracterizadas genotipicamente atrav?s de BOX-PCR. Das an?lises de agrupamento geradas a partir dos perfis de BOX-PCR, um total de 47 estirpes foram selecionadas para identifica??o taxon?mica atrav?s do sequenciamento parcial do gene 16S rRNA. A an?lise de similaridade de fragmentos do gene 16S rRNA revelaram que 36% das sequ?ncias pertenciam ao g?nero Gluconacetobacter, 31% a Azospirillum spp, 21% a Enterobacter spp. Estes resultados foram confirmados pela an?lise do gene nifH, embora, bact?rias identificadas como Gluconacetobacter apresentaram sequ?ncias de nifH hom?logas a Enterobacter spp. Adicionalmente, as 204 estirpes foram investigadas quanto ? capacidade de fixar nitrog?nio, produzir fitoreguladores e solubilizar fosfatos. Do total, 75.5% apresentaram atividade da nitrogenase, 97% produziram compostos ind?licos, 22% solubilizaram fosfato e 15% apresentaram as tr?s caracter?sticas. Para avaliar, sob condi??es de vaso, a resposta dos gen?tipos Cameroon e Roxo ? inocula??o com estirpes dos g?neros Azospirillum, Klebsiella, Enterobacter e Gluconacetobacter previamente caracterizadas. O rendimento de biomassa, o ac?mulo de nutrientes na parte a?rea, bem como o teor de prote?na nos tecidos, n?o foram beneficiados significativamente pela inocula??o com bact?rias diazotr?ficas; embora incrementos na massa seca de raiz e ac?mulo de N e P tenham sido detectados. Como os benef?cios proporcionados ? planta hospedeira apresentaram varia??o de estirpe para estirpe, e as an?lises foram realizadas pontualmente aos 60 dias ap?s plantio, n?o foi poss?vel determinar se as estirpes inoculadas apresentaram compet?ncia no estabelecimento e quais fatores influenciaram a intera??o planta-bact?ria.A diversidade, estrutural e funcional, foi acessada pela t?cnica de PCR-DGGE a partir de sequ?ncias de 16S rRNA e nifH obtidas diretamente de amostras de raiz e colmo de 5 gen?tipos de capimelefante. A estrutura das comunidades, bacteriana e diazotr?fica, foi mais influenciada pelo tecido vegetal do que pelos gen?tipos, sendo a raiz considerada um ambiente mais complexo em rela??o ao colmo. A popula??o bacteriana metabolicamente ativa foi identificada em 3 dos 5 gen?tipos testados; e os grupos detectados com maior frequ?ncia foram Proteobacteria, constituido de - (Leptothrix spp e Burkholderia spp), - (Bradyrhizobium spp, Methylobacterium spp e Rhizobium spp), - (Steroidobacter spp) e Actinobacteria (Actinoplanes spp, Conexibacter spp, Solirubrobacter spp e Amycolatopsis spp). Nas bibliotecas constru?das de nifH-cDNA, 26,4% do total de fragmentos foram relacionados com sequ?ncias de diferentes estirpes de Bradyrhizobium spp. Sequ?ncias menos abundantes pertencentes aos g?neros Azospirillum, Burkholderia, Klebsiella e Enterobacter spp tamb?m foram detectadas, mas sua distribui??o entre as amostras foi aleat?ria.
Databáze: OpenAIRE
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