Polyphasic characterization of nitrogen-fixing and co-resident bacteria in nodules of Phaseolus lunatus inoculated with soils from Piaui State, Northeast Brazil

摘要

Phaseolus lunatus (Lima bean) is an important legume for the poor population of the Brazilian northeast region. The legume is able to take advantage of the nitrogen fixation process, but the diversity of indigenous microsymbionts is poorly known. In this study, 29 bacteria isolated from root nodules of P. lunatus inoculated under greenhouse conditions with soils from Piaui State, in the northeast semi-arid region of Brazil, were obtained and characterized. Classical morphological and biochemical essays revealed high phenotypic diversity, splitting the bacteria into four clusters. Genetic fingerprinting by BOX-PCR indicated outstanding diversity, with the 29 strains positioned in 19 different clusters with 30% of final similarity. Nine genera were confirmed in the analysis of the 16S rRNA gene, with two typical nodulating N-2-fixing clades, Bradyrhizobium and Agrobacterium/Rhizobium, accounting for 38% and 21% of the isolates, respectively; the results were confirmed with the housekeeping gyrB gene, that also indicated putative new species. Bradyrhizobium was confirmed as the main symbiont, being present in nodules of all plants. All rhizobia except for those with higher resemblance to agrobacteria carry nifH genes. Six other genera were isolated as nodules endophytes, Bacillus, Burkholderia, Enterobacter, Franconibacter, Pseudomonas and Williamsia. Several of these endophytes exhibited one or more important biochemical properties, such as the synthesis of catalase, gelatinase and the ability to solubilize phosphate, that might confer ecological advantages to the rhizobia associated with them in the harsh environment of the Brazilian semi-arid, explaining the high rate of co-infection detected in the nodules.