To confirm the roles of agr in biofilm-associated events we found in Se 1457 genetic mutants above, here we treated Se 1457 wt strain GDC-0449 mouse with or without human hemoglobin (40 or 200 μg/mL). The results indicated that hemoglobin significantly reduced RNAIII transcripts (~40%-70% of inhibition) while increased atlE (~2.5-5.5 folds) but almost not affecting icaA (Figure 7). Functional assays further confirmed that hemoglobin increased biofilm formation, initial attachment, extracellular DNA release and cell autolysis

in a dose-dependent manner (Figure 7), while which does not affect bacterial growth (data not shown). Figure 7 Chemical inhibition of agr exhibit increased biofilm formation, extracellular DNA release and cell autolysis through upregulation of atlE . S. epidermidis 1457 was treated with or without hemoglobin (40 or 200 μg/mL), then (a) Biofilm-associated gene transcripts were measured by using qRT-PCR; (b) Biofilm biomass was quantified using

a crystal violet assay; (c-e) Initial attachment, extracellular DNA release and cell autolysis were determined as described above, respectively. Error bars represent the S.E.M. for three independent experiments. Discussion Se biofilm formation on implanted medical devices may result in recurrent or refractory infection unless the devices are removed, and removal and replacement BMN 673 in vivo of these devices incurs significant cost and risk for the patient. Flow-chamber systems simulate blood or other body-fluid flow in the vasculature of patients [18]. Using this and other complimentary approaches, we found that clinical Cediranib (AZD2171) Se isolates from patients with implanted

catheter infections display greater microcolony densities, spontaneous cell death, and self-renewal capacity during biofilm development relative to reference strains. Bacteria in biofilms are 100 ~ 1000 times more resistant to antibiotics than Selleck AZD1080 planktonic cells [21–23], although our study does not directly address antibiotic sensitivity for our clinical isolates. Staphylococcal biofilm dispersal is associated with severe infection, including endocarditis, pneumonia and sepsis [24–26]. In addition, dispersal cells help bacteria establish new biofilms in more suitable niches, resulting in infection within multiple tissues [27]. Of interest, we collected the detached and “flow-out” cells in the flow-chamber systems for our clinical isolates and found living cells capable of forming new biofilms as quickly as their parent cells (Qin et al., unpublished data). Interestingly, expression of RNAIII, a gene for the effector molecule of the agr system, was significantly reduced in all 4 Se clinical isolates, suggesting that the functions of agr quorum-sensing system were impaired in these isolates. Besides its regulatory function, RNAIII also encodes a δ-toxin, which effectively reduces cell attachment and subsequent biofilm formation of a Se agr mutant [13]. Our work does not address how RNAIII transcripts might be downregulated in our clinical isolates.