Two Gram-positive, coccoid, non-spore-forming bacteria (strains 3-M5-R-4(T) and 3-M5-R-7), cells of which formed diploid, tetrad and cluster arrangements, were isolated from air of the Regenerative Enclosed Life Support Module Simulator system. On the basis of 16S rRNA gene sequence similarity, these strains were shown to belong to the family Intrasporangiaceae and were related to members of the genus Tetrasphaera, with similarities to the seven known species of the genus Tetrasphaera of 96.71-97.76 %. The fatty acid profile supported affiliation of these novel isolates to the genus Tetrasphaera, although larger amounts of octadecanoic acid (C(18 : 0)) and cis-9-octadecenoic acid (C(18 : 1)) were observed in the isolates, thus enabling them to be differentiated from other Tetrasphaera species. In addition, DNA-DNA hybridization studies indicated that these strains belonged to a novel species that could be readily distinguished from its nearest neighbour, Tetrasphaera japonica DSM 13192(T), which had less than 20 % DNA-DNA relatedness. Physiological and biochemical tests showed few phenotypic differences, but genotypic analysis enabled these gelatin-liquefying strains to be differentiated from the seven Tetrasphaera species. The strains described in this study therefore represent a novel species, for which the name Tetrasphaera remsis sp. nov. is proposed; the type strain is 3-M5-R-4(T) (=ATCC BAA-1496(T) =CIP 109413(T)).