Proteomic analysis of Rhizobium freirei PRF 81 responses to low pH.

Rhizobium freirei PRF 81 is employed in common bean commercial inoculants in Brazil, due to its outstanding efficiency in fixing nitrogen, competitiveness and tolerance to abiotic stresses. Among the environmental conditions faced by rhizobia in soils, acidity is perhaps the encountered most, especially in Brazil. So, we used proteomics based approaches to study the responses of PRF 81 to a low pH condition. R. freirei PRF 81 was grown in TY medium until exponential phase in two treatments: pH 6,8 and pH 4,8. Whole-cell proteins were extracted and separated by two-dimensional gel electrophoresis, using IPG-strips with pH range 4-7 and 12% polyacrilamide gels. The experiment was performed in triplicate. Protein spots were detected in the high-resolution digitized gel images and analyzed by Image Master 2D Platinum v 5.0 software. Relative volumes (%vol) of compared between the two conditions tested and were statistically evaluated (p ≤ 0.05). Even knowing that R. freirei PRF 81 can still grow in more acid conditions, pH 4.8 was chosen because didn´t affect significantly the bacterial growth kinetics, a factor that could compromise the analysis. Using a narrow pH range, the gel profiles displayed a better resolution and reprodutibility than using broader pH range. Spots were mostly concentrated between pH 5-7 and molecular masses between 17-95 kDa. From the six hundred well-defined spots analyzed, one hundred and sixty-three spots presented a significant change in % vol, indicating that the pH led to expressive changes in the proteome of R. freirei PRF 81. Of these, sixty-one were up-regulated and one hundred two was downregulated in pH 4.8 condition. Also, fourteen spots were only identified in the acid condition, while seven spots was exclusively detected in pH 6.8. Ninety-five differentially expressed spots and two exclusively detected in pH 4,8 were selected for Maldi-Tof identification. Together with the genome sequencing and the proteome analysis of heat stress, we will search for molecular determinants of PRF 81 related to capacity to adapt to stressful tropical conditions.

Agronomic efficiency of Bradyrhizobium in peanut under different environments in Brazilian Northeast.

Several legumes have natural ability to associate with nitrogen - fixing bacteria known as rhizobia. The efficiency of this association depends on the plant and bacterial genotype and the edaphoclimatic conditions. Peanut is a tropical legume able to associate with a wide range of rhizobia and the selection of efficient bacteria is important to increase the nitrogen fixation in this crop. In order to investigate the agronomic efficiency of two Bradyrhizobiumstrains, two peanut genotypes were used in field trails carried out in three environments located at Brazilian Northeast. The genotypes (BR1 and L7 Bege) were submitted to rhizobial inoculation (SEMIA 6144 or ESA 123, both Bradyrhizobium strains, and chemical nitrogen fertilization in randomized block design experiments. The following traits were analyzed: flowering (F), main axis height (MAH), number of nodules/plant (NN), number of pods/plant (NP) and weight of pods (WP). Differential responses were found in all to treatments to NN, NP and WP, in the three environments studied. Overall, ESA 123 showed good agronomic performance inducing higher pod production. The results supportthe evaluation of the Bradyrhizobium in further experiments aiming at its recommendation to commercial inoculants in Brazilian Northeast region.