Time-Kill Assay

The time-kill kinetics of blestriacin was determined for four S. aureus strains as previously reported (Guo et al., 2016). Briefly, the antimicrobial agent was added to a logarithmic-phase broth culture of approximately 10⁶ CFU/mL to yield concentrations of 1× MIC, 2× MIC, and 4× MIC. The same volume of DMSO was added to the control tube. The culture was incubated at 37°C with shaking for 24 h. One hundred microliter aliquots were removed from each test tube after 0, 0.5, 1, 3, 6, and 24 h of incubation, and serially diluted (1:10) with cold saline. The appropriate dilutions (100 μL) were then spotted on MH agar plates (5 μL per spot), and viable bacteria were quantitated after 24 h of incubation at 37°C. Results were obtained from three independent experiments presented as the average ± standard deviation (SD).