Evaluation of a tetracycline-inducible promoter in Staphylococcus aureus in vitro and in vivo and its application in demonstrating the role of sigB in microcolony formation

An inducible promoter system provides a powerful tool for studying the gene tic basis for virulence. A variety of inducible systems have been used in o ther organisms, including pXyl-xylR-inducible promoter, the pSpac-lacI syst em, and the arabinose-inducible P-BAD, promoter, but each of these systems has limitations in its application to Staphylococcus aureus. In this study, we demonstrated the efficacy of a tetracycline-inducible promoter system i n inducing gene expression in S. aureus in vitro and inside epithelial cell s as well as in an animal model of infection. Using the xyl/tetO promoter:: gfp(uvr) fusion carried on a shuttle plasmid, we demonstrated that dose-dep endant tetracycline induction, as measured by bacterial fluorescence, occur red in each of the above environments while basal activation under noninduc ed conditions remained low. To ascertain how the system can be used to eluc idate the genetic basis of a pathogenic phenotype, we cloned the sigB gene downstream of the inducible promoter. Induction of SigB expression led to d ose-dependent attachment of the tested strain to polystyrene microliter wel ls. Additionally, bacterial microcolony formation, an event preceding matur e biofilm formation, also increased with tetracycline induction of SigB.