Staphylococcus aureus is a bacterium that colonizes and infects both humans and animals. As little is known about the phenotypic and molecular characteristics of S. aureus from wild animals in sub-Saharan Africa, the objective of the study was to characterize S. aureus isolates from wildlife and to analyse if they differed from those found among humans. The resistance to penicillin was low in S. aureus isolates from non-human primates (2.9%). Phylogenetic analysis based on the concatenated sequences from multilocus sequence typing revealed two highly divergent groups of isolates. One group was predominated by S. aureus that belonged to known human-related STs (ST1, ST9 and ST601) and mainly derived from great apes. A second clade comprised isolates with novel STs. These isolates were different from classical human S. aureus strains and mainly derived from monkeys. Our findings provide the basis for future studies addressing the inter- and intra-species transmission of S. aureus in Africa.