Culture-independent molecular approaches reveal a mostly unknown high diversity of active nitrogen-fixing bacteria associated with Pennisetum purpureum—a bioenergy crop

AIMS: Previous studies have shown that elephant grass is colonized by nitrogen-fixing bacterial species; however, these results were based on culture-dependent methods, an approach that introduces bias due to an incomplete assessment of the microbial community. In this study, we used culture-indepen...

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Bibliographic Details
Published in:Plant and soil 2013-12, Vol.373 (1-2), p.737-754
Main Authors: Videira, Sandy Sampaio, Pereira e Silva, Michele de Cássia, de Souza Galisa, Péricles, Dias, Armando Cavalcante Franco, Nissinen, Riitta, Divan, Vera Lúcia Baldani, van Elsas, Jan Dirk, Baldani, José Ivo, Salles, Joana Falcão
Format: Article
Language:English
Subjects:
Agricultural research
Agricultural soils
Agronomy. Soil science and plant productions
Animal, plant and microbial ecology
Bacteria
bacterial communities
Biological and medical sciences
Biological diversity
Biomedical and Life Sciences
Bradyrhizobium
Burkholderia
clones
complementary DNA
denaturing gradient gel electrophoresis
Ecology
Elephants
endophytes
energy crops
Environmental aspects
Fundamental and applied biological sciences. Psychology
General agronomy. Plant production
genes
genotype
Genotype & phenotype
Genotypes
Grasses
in vitro culture
Leptothrix (bacteria)
Libraries
Life Sciences
nitrogen fixation
nitrogen-fixing bacteria
Nitrogen-fixing microorganisms
nucleotide sequences
Pennisetum purpureum
phylogeny
Plant Physiology
Plant roots
Plant Sciences
plant tissues
Plants
Polymerase chain reaction
Regular Article
rhizoplane
ribosomal RNA
roots
Soil microorganisms
Soil Science & Conservation
Soil-plant relationships. Soil fertility
Soil-plant relationships. Soil fertility. Fertilization. Amendments
species diversity
stems
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Summary:AIMS: Previous studies have shown that elephant grass is colonized by nitrogen-fixing bacterial species; however, these results were based on culture-dependent methods, an approach that introduces bias due to an incomplete assessment of the microbial community. In this study, we used culture-independent methods to survey the diversity of endophytes and plant-associated bacterial communities in five elephant grass genotypes used in bioenergy production. METHODS: The plants of five genotypes of elephant grass were harvested from the experimental area of Embrapa Agrobiologia and divided into stem and root tissues. Total DNA and RNA were extracted from plant tissues and the bacterial communities were analyzed by DGGE and clone library of the 16S rRNA and nifH genes at both the cDNA and DNA levels. RESULTS: Overall, the patterns based on DNA- and RNA-derived DGGE-profiles differed, especially within tissue samples. DNA-based DGGE indicated that both total bacterial and diazotrophic communities associated with roots (rhizoplane + endophytes) differed clearly from those obtained from stems (endophytes). These results were confirmed by the phylogenetic analyses of RNA-derived sequences of 16S rRNA (total bacteria; 586 sequences), but not for nifH (186). In fact, rarefaction analyses showed a higher diversity of diazotrophic organisms associated with stems than roots. Based on 16S rRNA sequences, the clone libraries were dominated by sequences affiliated to members of Leptotrix (12.8 %) followed by Burkholderia (9 %) and Bradyrhizobium (6.5 %), while most of the nifH clones were closely related to the genus Bradyrhizobium (26 %). CONCLUSIONS: Our results revealed an unexpectedly large diversity of metabolically active bacteria, providing new insights into the bacterial species predominantly found in association with elephant grass. Furthermore, these results can be very useful for the development of new strategies for selection of potential bacteria that effectively contribute to biological nitrogen fixation and enhance the sustainable production of elephant grass as bioenergy crop.
ISSN:0032-079X
1573-5036
DOI:10.1007/s11104-013-1828-4