Filogenia e diversidade molecular de sabiá (Mimosa caesalpiniifolia Benth.) e de bactérias diazotróficas

Abstract

Leguminosae is the third largest angiosperm family, with around 700 genera, of which the Mimosoideae subfamily comprehends 78 genera, with emphasis to Acacia, Mimosa and Inga. Mimosa caesalpiniifolia Benth., known as “sabiá” or “sansão do campo” is considered to be one of the most important native tree species of the Brazilian semiarid due to its multiuse capability, and high potential for degraded area recovery, since it fixes nitrogen in symbiosis with diazotrophic bacteria. Diazotrophic bacteria taxonomy has been changing due to joint use of phenotypic, physiologic and molecular tools. This work aimed to evaluate “sabiá” and its symbiotic bacteria diversity in five Northeastern municipalities. It was conducted from April, 2010 to March, 2011, at Pernambuco Federal Agricultural University and the Genome Laboratory of the Pernambuco Agronomic Institute. For “sabiá” phylogenetic studies, leaves from native or naturalized plants were collected at Crato, Gravatá, Itambé, Mossoró and Serra Talhada, with their genomic DNA extracted with a commercial kit. The intergenomic atpB-rcbL region of chloroplastic DNA was amplified and used to construct Bayesian Inference phylogenesis of the accesses. Soil samples were collected at the same time of plant collection, and “sabiá” plants were used as bait for rhizobial nodules using Leonard jars, at a greenhouse. Nodule bacteria were isolated and purified in YMA media with Congo Red, and morpho-physiologically characterized on YMA media with Bromothymol Blue. The isolates were later grown in liquid TY media for DNA extraction with a commercial kit. Amplifications were conducted with REP, ERIC and BOX primers, and 16S rDNA was amplified and sequenced. Intergenic atpB-rbcL chloroplast DNA sequences did not match any NCBI entry. CRATO 4 and SERRA TALHADA 20 accesses formed an external group indicated they may be genetically closer to a Mimosa ancestor. The high segregation of the species affected diversity within and among the different areas, and plant biogeography was not confirmed. Genomic fingerprinting of the 47 isolates had different patterns for REP, ERIC and BOX elements, but compilation of the results created the dendrogram with the most groups. The 16S rDNA sequences were blasted in GenBank, and the isolates had 68 to 99% similarity with Burkholderia strains. The phylogenetic tree constructed with the 16S rDNA, combined with sequences from strains recommended for legume inoculation, show these isolates to be diverse from the currently recommended. Isolates PE-MO01, PE-MO02 and PE-MO04 were the most different among the isolates. The lack of molecular phylogeny data for “sabiá” shows the need of using other taxonomic markers to evaluate this species diversity, and the 16S rDNA sequences confirm the Mimosa symbiosis preference for Burkholderia strains.