A novel streptomycete strain, designated XY25T, was isolated from the rhizosphere soil in an alfalfa field in Jingyang, Shanxi, China. The isolate showed optimal growth at 37 °C, and was capable of growing at pH 6-10 and in the presence of 0-6 % (w/v) NaCl. Mycelia of strain XY25T appeared spiral and developed into white spore chains with long-rod spores and a smooth surface. The 16S rRNA gene sequence of XY25T was determined and was found to be highly similar to those of species of the genus Streptomyces including Streptomyces silaceus DSM 41861T (99.11 % 16S rRNA gene sequence similarity), Streptomyces flavofungini DSM 40366T (98.49 %) and Streptomyces intermedius DSM 40372T (98.43 %), all of which were used for further characterization. Each of the four streptomycetes showed distinctive patterns of carbon usage and fatty acids composition. Analysis of cellular components of strain XY25T revealed ll-diaminopimelic acid as diagnostic diamino acid and xylose as the major sugar, whereas polar lipids were determined as phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylinositol, an unknown phospholipid, two unknown phosphatidylinositol mannosides and several unknown lipids. Menaquinones were dominated by MK-9(H6) and MK-9(H8), and the main fatty acids were anteiso-C15 : 0, iso-C16 : 0 and anteiso-C17 : 0. DNA-DNA hybridization studies indicated that strain XY25T showed relatedness values of 35.2-40.42 % with the closest related species. Based on these results, strain XY25T represents a novel species of the genus Streptomyces, for which the name Streptomyces alfalfae sp. nov. is proposed. The type strain is XY25T ( = KCTC 39571T = CCTCC AA2015019T).