Isolation, characterization and selection of diazotrophic bacteria in Paspalum genotypes

Autor: Amaral, Mayan Blanc
Přispěvatelé: Baldani, Vera L?cia Divan, Baldani, Jos? Ivo, Ara?jo, Adelson Paulo de, Goi, Silvia Regina
Jazyk: portugalština
Rok vydání: 2017
Předmět:
Zdroj: Biblioteca Digital de Teses e Dissertações da UFRRJ
Universidade Federal Rural do Rio de Janeiro (UFRRJ)
instacron:UFRRJ
Popis: Submitted by Jorge Silva (jorgelmsilva@ufrrj.br) on 2022-07-18T19:32:36Z No. of bitstreams: 1 2013 - Mariane Del Carmen da Costa Diaz.pdf Made available in DSpace on 2022-07-18T19:32:36Z (GMT). No. of bitstreams: 1 2013 - Mariane Del Carmen da Costa Diaz.pdf Previous issue date: 2017-02-17 CNPq - Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico Nitrogen is considered scarce in tropical soils and the use of N-fertilizer can lead to water resources contamination, raising the energy cost of production and contributing to the generation of greenhouse gases (GHG) that affects the generation of global warming. To achieve sustainability of crops becomes necessary to use alternative sources of N as the Biological Nitrogen Fixation. This work aimed to isolate, characterize and select efficient strains in the plant growth promotion into 10 genotypes of Paspalum. The isolation was performed in roots and rizosphere soil of 10 genotypes of Paspalum sp. from the Active Germplasm Bank of Embrapa Pecu?ria Sudeste, S?o Carlos, SP. The bacteria were isolated in nitrogen-free semi-solid media, NFB and LGI and LG solid medium. A total of 161 isolates were obtained, 127 obtained in LG medium, 17 in NFB medium and 17 in LGI medium. The isolates were morphologically characterized and grouped with this features. From this groups were selected 80 representative isolates to genetic characterization. This isolates had their DNA extracted using the kit Wizard? Genomic DNA Purification Kit (Promega?), according to manufacturer's specification. The DNA of 80 isolates was analyzed by BOX-PCR. For the construction of the dendrogram, the similarity matrices were calculated by Jaccard coefficient and the grouping of similarity matrices using the UPGMA algorithm (Unweighted Pair Group Method with Arithmetic Mean). Isolates were grouped and representatives of each group were selected for taxonomic identification by 16S rDNA gene sequencing and for physiological characterization by phosphates solubilization, indole compounds production and siderophores production. After this, the most promising isolates of BRA 19186, BRA 9610 and BRA 23540 genotypes, were tested in 3 greenhouse expirements. The BOX-PCR analysis showed high genotypic diversity, forming 55 isolates grouped with 70% similarity, being obtained 11 groups in the LG medium, none similar group in LGI medium and only 2 similar isolates in NFB medium. Of the 55 isolates tested, 73% of isolates produced siderophores, only 25% solubilized phosphates and 9% produced indolic compounds. We sequenced 30 isolates, 16 isolates belonging to the genus Bacillus, 6 isolates to the genus Rhizobium, 2 to the genus Burkholderia, 2 to Pseudomonas genus, 1 isolate to Dyadobacter, 1 to the genus Acinetobacter, 1 isolated to the genus Mycobacterium and 1 isolated to the Azospirillum genus. We obtained on the greenhouse experiments, gains on the biometrics variables and on the shoot dry matter up to 56%. There are difference in the inoculation answer in relation to mowing regimes, genotype and isolates tested on the experiments. O nitrog?nio ? considerado escasso em solos tropicais e o uso do N-fertilizante pode ocasionar a contamina??o dos recursos h?dricos, elevar o custo energ?tico da produ??o, al?m de contribuir para a gera??o de gases do efeito estufa que afetam o aquecimento global. Para alcan?ar a sustentabilidade das culturas torna-se necess?rio o uso de fontes alternativas de N, como a Fixa??o Biol?gica de Nitrog?nio. Objetivou-se nesse trabalho isolar, caracterizar e selecionar estirpes eficientes na promo??o de crescimento vegetal em gen?tipos de Paspalum. O isolamento foi realizado em solo rizosf?rico e ra?zes de 10 gen?tipos de Paspalum spp. oriundos do Banco Ativo de Germoplasma da Embrapa Pecu?ria Sudeste, S?o Carlos, SP. As bact?rias foram isoladas nos meios semiss?lidos livres de Nitrog?nio, NFB e LGI e no meio s?lido LG. Foi obtido um total de 161 isolados, 127 obtidos em meio LG; 17 em meio NFB e 17 em meio LGI. Os isolados foram caracterizados morfologicamente e agrupados de acordo com essas caracter?sticas. Desses grupamentos foram selecionados 80 isolados representativos para serem caracterizados geneticamente. Esses isolados tiveram o seu DNA extra?do com o uso do Kit Wizard? Genomic DNA purification Kit (Promega?), conforme especifica??o do fabricante. O DNA dos 80 isolados foi analisado pela t?cnica de BOX-PCR. Para a constru??o do dendrograma, as matrizes de similaridade foram calculadas pelo coeficiente de Jaccard e o agrupamento de matrizes de similaridade utilizando o algoritmo UPGMA. Os isolados foram agrupados e representantes foram selecionados para a identifica??o taxon?mica atrav?s do seq?enciamento do gene 16S rRNA e para a caracteriza??o fisiol?gica quanto ? solubiliza??o de fosfatos, produ??o de compostos ind?licos e produ??o de sider?foros. Ap?s isto, os isolados dos gen?tipos BRA 19186, BRA 9610 e BRA 23540 mais promissores na caracteriza??o fisiol?gica foram testados em 3 experimentos em casa-de-vegeta??o. As an?lises de BOX-PCR revelaram uma alta diversidade genot?pica, formando 55 isolados agrupados com 70 % de similaridade, sendo obtidos no meio LG, 11 grupos, no meio LGI nenhum grupo similar e no meio NFB apenas 2 isolados similares. Do total de 55 isolados, 73% isolados produziram sider?foros, 25% solubilizaram fosfatos e 9% produziram compostos ind?licos. Foram seq?enciados 30 isolados, sendo 16 isolados pertencentes ao g?nero Bacillus, 6 isolados ao g?nero Rhizobium, 2 ao g?nero Burkholderia, 2 ao g?nero Pseudomonas, 1 isolado ao g?nero Dyadobacter, 1 isolado ao g?nero Acinetobacter, 1 isolado ao g?nero Microbacterium e 1 isolado ao g?nero Azospirillum. Os experimentos mostraram ganhos nas vari?veis biom?tricas e na massa seca da parte a?rea de at? 56%. Houve varia??o na resposta a inocula??o em rela??o ?s coletas, gen?tipos e isolados testados nos 3 experimentos.
Databáze: OpenAIRE
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