Transmission of Staphylococcus aureus colonization in community-based populations is not well understood. We sought to describe the molecular epidemiology of S. aureus colonization in the Old Order Amish. The study was a prospective, observational study of healthy adults and their same-sex siblings who were cultured from the anterior nares twice. S. aureus isolates were characterized using spa typing. Overall, 40% (159/398) of the study population was colonized with S. aureus. There were 84 spa types with the most abundant spa types being t012 (13%) and t021 (7%). There was no clustering of spa types within sibling groups; however, there was clustering within households. There were 111 S. aureus-colonized participant pairs living within the same household. Of these, 47% had concordant spa types. The diversity of spa types across a relatively isolated, genetically homogenous population with a similar lifestyle is striking. Taken together this suggests that S. aureus transmission is a local phenomenon limited to very close contact.