ABSTRACT Several lines of evidence indicate that the most primitive staphylococcal species, those of the Staphylococcus sciuri group, were involved in the first stages of evolution of the staphylococcal cassette chromosome mec (SCC mec ), the genetic element carrying the β-lactam resistance gene mecA . However, many steps are still missing from this evolutionary history. In particular, it is not known how mecA was incorporated into the mobile element SCC prior to dissemination among Staphylococcus aureus and other pathogenic staphylococcal species. To gain insights into the possible contribution of several species of the Staphylococcus sciuri group to the assembly of SCC mec , we sequenced the genomes of 106 isolates, comprising S. sciuri ( n = 76), Staphylococcus vitulinus ( n = 18), and Staphylococcus fleurettii ( n = 12) from animal and human sources, and characterized the native location of mecA and the SCC insertion site by using a variety of comparative genomic approaches. Moreover, we performed a single nucleotide polymorphism (SNP) analysis of the genomes in order to understand SCC mec evolution in relation to phylogeny. We found that each of three species of the S. sciuri group contributed to the evolution of SCC mec : S. vitulinus and S. fleurettii contributed to the assembly of the mec complex, and S. sciuri most likely provided the mobile element in which mecA was later incorporated. We hypothesize that an ancestral SCC mec III cassette (an element carried by one of the most epidemic methicillin-resistant S. aureus clones) originated in S. sciuri possibly by a recombination event in a human host or a human-created environment and later was transferred to S. aureus .