aureus controls its biofilm so as to discover novel compounds capable of find more inhibiting or dispersing biofilms without allowing bacteria to develop drug resistance. The diverse mechanisms that have been reported for biofilm control in S. aureus include quorum sensing, protease, DNase, cis-2-decenoic acid, d-amino acids, phenol-soluble polypeptides, several

surface proteins, and pH change (Boles & Horswill, 2011). Particularly, the activation of agr quorum-sensing and protease treatment in S. aureus inhibited its own biofilm formation and dispersed the established biofilms (Vuong et al., 2000; Boles & Horswill, 2008). A serine Esp protease in Staphylococcus epidermidis inhibited S. aureus biofilm formation and nasal colonization (Iwase et al., 2010). However, the target of these agr controlled protease and the specific

target of Esp protease is not known (Boles & Horswill, 2008; Iwase et al., 2010). Recently, we have shown that various Actinomycetes strains produce a large I-BET-762 cost amount of protease that rapidly dispersed S. aureus biofilm (Park et al., 2012). In the present study, more diverse bacteria are used to screen for S. aureus biofilm reduction. We find that two Pseudomonas aeruginosa supernatants dispersed S. aureus biofilm and contained high protease activities. Another study goal is to identify a main antibiofilm component and a possible mechanism of protease-involved biofilm dispersal. Transcriptional analysis and phenotypic assays are conducted to confirm that S. aureus Ribonuclease T1 triggers its biofilm dispersal through the accelerated effect of protease activity. All experiments were conducted at 37 °C, and Luria-Bertani (LB) medium was used for culturing all strains (Table 1). Two S. aureus strains (ATCC 25923 and ATCC 6538) were obtained from the Korean Agricultural Culture Collection and used to reinforce our findings in two different strains. To identify a main antibiofilm protease, thirteen P. aeruginosa PAO1 transposon

mutants (Jacobs et al., 2003) were obtained from the University of Washington Genome Center (Supporting information, Table S1). To obtain culture supernatants, a fresh single colony of bacteria was inoculated and cultured in LB at 250 r.p.m. for 24 h. All bacterial supernatants were filtered with a 0.45 μm filter to completely remove any bacteria before further use. Fresh culture supernatants were used in every experiment. Crystal violet, casamino acid, succinic acid, calcium carbonate, sodium phosphate, ethanol, soluble starch, and potassium phosphate were of analytical grade. DNase I (Cat. 79254) was purchased from Qiagen (Valencia, CA), and RNase A (Cat. 12091021) was purchased from Life Technologies (Grand Island, NY). For the cell growth measurements, the optical density was measured at 600 nm using a spectrophotometer (UV-160, Shimadzu, Japan). Each experiment was performed with at least two independent cultures.