, 2004). S. aureus and P. aeruginosa are often found together in the airways of

cystic fibrosis patients (Hoffman et al., 2006) and both opportunistic human pathogens readily form biofilms on diverse surfaces. Hence, both biofilm control and interspecies interactions are important in the course of disease. The current results demonstrated that P. aeruginosa PAO1 supernatant can inhibit and disperse S. aureus biofilm via the protease activity from P. aeruginosa, which is independent of a bactericidal effect (Fig. 1). Pseudomonas aeruginosa apparently produced various determinants to control S. aureus biofilm, including staphylolytic protease secretion (Kessler et al., 1993) and 4-hydroxy-2-heptylquinoline-N-oxide (HQNO) BYL719 concentration production (Mitchell et al., 2010). While protease dispersed S. aureus biofilm (Fig. 1), HQNO stimulated S. aureus to form a biofilm and small-colony variants (Hoffman et al., 2006; Mitchell et al., 2010). Analysis of gene expression showed that HQNO induced sigma factor B (sigB) and repressed the quorum-sensing regulator (agrA) and the α-hemolysin

(hla) in S. aureus (Mitchell et al., 2010). In contrast, P. aeruginosa protease induced S. aureus protease genes (aur, clp, scpA, splA, and sspA), two regulatory genes (agrA and sigB), and hemolysin gene (hla) in S. aureus (Fig. 3). These results imply that the interaction between two species in vivo is dependent on the amount and types of exoproducts, such as HQNO and proteases influenced by temporal and spatial, environmental conditions. Although speculative, Ceramide glucosyltransferase S. aureus may have the ability to control its biofilm (up-regulation mTOR inhibitor by HQNO and down-regulation by protease) by interacting exoproducts from P. aeruginosa. The action mechanism of protease-involved biofilm control in S. aureus has been partially elucidated that agr-mediated dispersal requires the

activity of protease, but in an ica-independent manner (Boles & Horswill, 2008). The expression of protease is positively regulated by agr (Novick, 2003) and negatively by sarA (Cheung et al., 2004) and sigB (Gertz et al., 2000; Martí et al., 2010). In accord with previous studies, this study has demonstrated that the protease activity was accompanied by the induction of agr and the repression of sarA (Fig. 3). Moreover, the addition of exogenous protease induced the gene expression of all five proteases (aur, clp, scpA, splA, and sspA), which led to the rapid dispersal of S. aureus biofilms (Fig. 1c and f). The protease activity assay (Fig. 2) and biofilm assay (Fig. 4.) of protease-deficient P. aeruginosa mutants partially revealed that LasB elastase is a main protease decreasing the biofilm formation of the tested S. aureus strain. Because other factors such as poly-N-acetylglucosamine, protein adhesins and extracellular DNA also play an important role in the biofilm formation of S. aureus (Izano et al., 2008; Mann et al.