Resumo

The Azospirillum genus is known by its plant growth promotion ability, and is used to produce commercial inoculants for different agriculture crops. The faculty of aggregation and floculation observed in Azospirillum has great importance for its survival and dispersion in the soil, and for the maintenance of quality of commercial inoculants. In this sense, the exopolysaccharides (EPS) produced by Azospirillum shows a strong correlation with the cell aggregation, and its presence can be used as an additional parameter to evaluate the quality of bacterial cultures targeted to be used as plant inoculants. Aiming to develop a high-quality Azospirillum inoculant, we evaluated the EPS production and cell agregation of A. brasilense strain Ab-V5 under different liquid broths formulated with simple and low-cost components. Based on a previous study, we selected three formulations (named FORM2, FORM4 and FORM15) to perform this work. Bacterial cultures of 100 mL of the formulations tested were added with a cell concentration of 1 x 10⁴ cells mL of A. brasilense Ab-V5. Following inoculation with A. brasilense, the formulations were incubated in orbital shaker at 180 rpm and 30 ºC for up to 120 hours in triplicate. Samples were then collected after 72, 96 and 120 hours of growth to microscopic and analytical determination of EPS production. For each sampling time, 10 mL of cultures were centrifuged (9,960 g, 4oC 15 min) and the EPS in the cell free extract (CFE) was precipitated by adding of 30 mL of ethanol. The EPS was dialyzed for 3 consecutive days, and then the material was lyophilized and weighed. The cellular aggregation 30 μμL of samples were stained with 30 μμL DAPI, incubated for 5 min in the dark at room temperature, and observed using an epifluorescence microscope. The results showed that all the formulations induced A. brasilense Ab-V5 to produce EPS and the maximum EPS content was observed after 120 hours of growth, although the amount of EPS produced varied amongst the tested formulations. When grown in the formulation FORM2, A. brasilense Ab-V5 produced the higher amount of EPS, reaching up to 1.56 g EPS/L culture, followed by FORM15 and FORM4 with up to 0.71 and 0.67 g EPS/L culture, respectively. The microscopic evaluations showed microaggregates of bacterial cells on the three formulations. The effect of EPS content over the attachment of A. brasilense to maize seeds and bacterial survival in soil and under storage of inoculants is under investigation.