We evaluated the production of indole-3-acetic acid (IAA) by Azospirillum brasilense strains in vitro (cell culture supernatants) and in vivo (stems and roots of maize seedlings) to clarify the role of this phytohormone as a signaling and effector molecule in the symbiotic interaction between maize and A. brasilense. The three strains all showed IAA production when cultured in NFb medium supplemented with 100 μg/ml L-tryptophan. The level of IAA production was 41.5 μg/ml for Yu62, 12.9 μg/ml for Az39, and 0.15 μg/ml for ipdC-. The release of IAA into culture medium by the bacteria appeared to be the main activator of the early growth promotion observed in the inoculated maize seedlings. The application of supernatants with different IAA contents caused significant differences in the seedling growth. This observation provides the basis for novel technological tools for effective quality control procedures on inoculants. The approach described can be incorporated into different inoculation methods, including line sowing, downspout, and foliar techniques, and increase the sustainability of symbiotic plant-bacteria systems.