A new actinobacterium strain, designated 2AT, was isolated from a soil sample collected from Guru Nanak Dev University, Punjab (India) and characterized using a polyphasic taxonomic approach. It showed antimicrobial activity against various Gram-positive and Gram-negative bacteria including drug resistant bacteria and fungi. The strain had chemotaxononomic and morphological properties typical of the genus Streptomyces. The 16S rRNA gene sequence of the strain showed 99.9, 99.5 and 99.5 % similarity with Streptomyces flavotricini DSM 40152T, Streptomyces toxytricini DSM 40178T and Streptomyces globosus DSM 40815T, respectively. This strain formed a coherent cluster with them and shared DNA–DNA homology of 37.6 ± 0.6, 34.4 ± 0.5 and 33.1 ± 0.4 % with type strains, S. flavotricini DSM 40152T, S. globosus DSM 40815T and S. toxytricini DSM 40178T, respectively. Further, the strain was readily distinguished from the phylogenetic close relatives in a variety of morphological, physiological and biochemical properties. Based on the genotypic and phenotypic characteristics, it is proposed that strain 2AT represents a novel species in the genus Streptomyces, for which the name Streptomyces amritsarensis sp. nov. is proposed, with the type strain 2AT (=MTCC 11845T=JCM 19660T).