Very small proteins with less than 100 amino acids (SP100) of Staphylococcus aureuse are underrepresented in proteomic analyses so far. However, in the last few years a variety of such small proteins with regulatory and virulence associated functions have been detected in several bacteria. The introduction of a new open source command line tool (Pepper) that provides a fully automated proteogenomic workflow enabled us to identify proteins encoded by non-annotated open reading frames based on identified peptides. Altogether, 185 soluble proteins with up to 100 amino acids have been detected of which 69 were not covered by the used gene annotation. Of these, 83 % were identified by at least two methods.