The IBPPM Collection maintains unique pool of bacterial strains related to Azospirillum genus, notable for its biotechnological potential. Progressively developing systematics of this genus requires checking and clarifying the taxonomic position for long-term stored isolates in the collection. Consequently, the species membership was confirmed or defined in the tested Azospirillum strains using a comparative analysis of their full-length 16S rRNA genes combined with morphological, cultural, physiological and biochemical properties, as well as DNA-DNA hybridization data.