Eight coagulase-negative, oxidase-negative and novobiocin-susceptible staphylococcal strains were isolated from the gastrointestinal tracts of South American squirrel monkeys (Saimiri sciureus L.). These strains were differentiated from known staphylococcal species on the basis of 16S rRNA gene and hsp60 gene sequencing, and from the most closely related species by using DNA-DNA hybridization, ribotyping, whole-cell protein profiles and biotyping. Phylogenetic analysis based on 16S rRNA gene sequences confirmed that these strains are members of the Staphylococcus aureus species group (99% similarity) but are biochemically similar to Staphylococcus piscifermentans, from which they can be phenotypically distinguished by resistance to polymyxin B, acid production from D-mannitol, the inability to hydrolyse aesculin and DNA and the absence of alpha-glucosidase. On the basis of these analyses, a novel species of the genus Staphylococcus is described, for which the name Staphylococcus simiae sp. nov. is proposed, with CCM 7213(T) (=LMG 22723(T)) as the type strain.