Non-adherent bacteria at multiplicity of infec- tion (MOI) of 25 or 50 retrieved from the supernatant of S9 cells after 1 hour of infection (nonad25-1h and nonad50-1h)
Cell culture infection experiments performed with the human bronchial epithelial cell line S9 and the human monocyte cell line THP-1. Staphylococci internalized by eukaryotic host cells were isolated from samples taken 2/ 2.5 hours (intTHP1-2h and intS9-2h) and 6/ 6.5 hours (intTHP1-6h and intS9-6h) post infection.
Bacteria after 2.5 hours of anaerobic incubation in pMEM medium at 37°C (anaer-2h)
S. aureus was grown in the presence of the cationic antimicrobial peptide colistin (colist). Colistin was added to TSB medium at 50 μg/mL and cells were grown until an OD 600 nm of 1.
S. aureus was grown with shaking at 37°C in rich medium (TSB), minimal medium (CDM), cell culture media (RPMI, pMEM), and in human plasma (plasma). Samples were taken during exponential growth phase (exp) as well as 2 hours (t2) and 4 hours (t4) after entry into stationary phase.
Bacteria grown in RPMI medium with fetal calf serum (RPMI/FCS), the pre-culture condition for the THP-1 infection experiments.
Bacteria after 1, 2.5, and 6.5 hours of incubation in cell culture dishes in S9 infection medium at 37°C and 5% CO2 without agitation (CO2 -1h, CO2 -2h and CO2 -6h)
S. aureus was grown in RPMI supplemented with subinhibitory concentrations of flucloxacillin (fluc) and vancomycin (vanc). Samples were taken during exponential growth phase (exp) and 4 hours (t4) after entry into stationary phase.
S. aureus was grown in the presence of subinhibitory concentrations of antibiotics: flucloxacillin (fluc), vancomycin (vanc), ciprofloxacin (cipro), clindamycin (clind), erythromycin (ery), linezolid (line), and trimethoprim-sulfamethoxazole (T/Smx) using the following concentrations: flucloxacillin 0.02 μg/mL, vancomycin 0.63 μg/mL, ciprofloxacin 0.10 μg/mL, clindamycin 0.01 μg/mL, erythromycin 0.05 μg/mL, linezolid 0.10 μg/mL, and trimethoprim-sulfamethoxazole 0.75 μg/mL. S. aureus was pre-cultured in TSB till early-exponential phase (OD 600 of 0.5), after which the culture was diluted 10-fold using TSB supplemented with the respective antibiotics. Samples were taken during exponential growth phase (exp) and 4 hours (t4) after entry into stationary phase.
S. aureus strain HG001 wild-type and rho-mutant cells were grown aerobically at 37 °C in TSB and RPMI medium. Samples were taken during exponential phase and 4 hours after entry into stationary phase from independent cultures (duplicates). For each of the four growth conditions, the wild-type profile is represented in black and the mutant in color (green: RPMI expo.; blue: RPMI t4; red: TSB expo.; yellow: TSB t4).