Staphylococcus aureus is the leading cause of wound and hospitalacquired infections worldwide. The emergence of S. aureus strains with resistance to multiple antibiotics requires the identification of bacterial virulence genes and the development of novel therapeutic strategies. Herein, bursa aurealis, a mariner-based transposon, was used for random mutagenesis and for the isolation of 10,325 S. aureus variants with defined insertion sites. By screening for loss-of-function mutants in a Caenorhabditis elegans killing assay, 71 S. aureus virulence genes were identified. Some of these genes are also required for S. aureus abscess formation in a murine infection model. transposon ͉ Caenorhabditis elegans