The ability to sequester iron is a primary defense mechanism against bacterial infection. Iron chelation therapy has been considered as a possible treatment for various infectious diseases. S. aureus isolated from neonatal septicemia were used to study the effect of deferoxamine and sub-minimal inhibitory concentration of gentamicine on some virulence factors of this isolates. Also an experimental sepsis was inducted in mice and treated with gentamicin and deferoxamine. The expression of virulence factor (alpha-hemolysin, beta-hemolysin, delta-hemolysin, coagulase, and DNase) by Staphylococcus aureus isolates was significantly decreased (p < 0.05) after exposure to DFO and/or gentamicin. The data of the present study showed that using of DFO led to significant decrease (p < 0.05) in the mortality rate of mice infected with S. aureus. In a murine model of S. aureus sepsis, deferoxamine treatment had an additional effect on survival and bacterial eradication from the organs of septicemic mice. In vitro exposure of S. aureus isolated to gentamicin and deferoxamine led to a decrease in the production of some virulence factors by these isolates.