We evaluated the effect of Azospirillum brasilense strain HM053 inoculation on maize seeds, a spontaneous mutant that excrete ammonium and fix nitrogen constitutively. Maize was grown with different nitrogen (urea) concentration and inoculated with A. brasilense Ab-V5 (Brazilian commercial strain) or HM053 strain in four field experiments, in three regions of Parana State, Southern Brazil. We evaluated yield components, nutrient content on leaves and grains and productivity during the crop cycle. Inoculation with A. brasilense strain Ab-V5 and HM053 associated with base fertilization (30 kg ha−1 N) improved crop yield in all trials. Ab-V5 increased production between 2.2 to 10.4%, or 178.0 to 759.9 kg ha−1, respectively. HM053, by itself, increased production between 4.7 to 29%, or 460.5 to 1769.3 kg ha−1, respectively. The new strain HM053 showed to be a great biofertilizer for maize seeds and a new alternative for a more sustainable agriculture.