Background Meticillin-resistant Staphylococcus aureus (MRSA) has spread throughout the world and has become highly endemic worldwide. The need for implementing MRSA control strategies is no longer a matter of debate. Aim To determine the temporal association between various infection control practices, the use of antibiotics and the incidence of MRSA in a 1200-bed French university hospital. Methods A multi-variate time-series analysis, based on monthly data from a nine-year period (January 2000-December 2008), was performed in a 1200-bed French university hospital to determine the temporal association between different variables and the incidence of MRSA. MRSA colonization pressure, infection control practices and use of antibiotics were considered in the analysis. Findings Time-series analysis showed a positive significant relationship between the incidence of hospital-acquired MRSA (HA-MRSA) and MRSA colonization pressure, the use of antibiotics (fluoroquinolones, macrolides and aminoglycosides) and the use of gloves. Conversely, a global negative correlation was observed between the incidence of HA-MRSA and the use of alcohol-based hand rub. Overall, the model explained 40.5% of the variance in the monthly incidence of MRSA. Conclusion This study showed that admission of patients with MRSA, the use of antibiotics and infection control practices contributed to the incidence of HA-MRSA. This suggests that efforts should be focused on high compliance with hand disinfection. These results also raise concerns about the use of gloves when caring for patients with MRSA.