Pathogenic bacterial colonisation in Cystic Fibrosis patients is associated with a poor prognosis; thus, protective measures need to be taken to prevent their transmission. We studied the extent of contamination in the environment of hospitalised children with cystic fibrosis (CF) associated with specific activities. We assessed the levels of bacterial contamination in 432 air and surface samples collected from various locations in our CF centre over a three-month period: the bedrooms, corridor, communal showers, school, leisure centre and the respiratory functional explorations (RFE) unit. Staphylococcus aureus and Pseudomonas aeruginosa strains found in bedrooms and the RFE were compared with those found in patient expectorations using pulsed field gel electrophoresis. In all sampling locations, there were high levels of airborne contamination just after the presence of patients or nursing staff. In the bedrooms, the amount of S. aureus or P. aeruginosa in the air, at wake-up and after physiotherapy, were significantly higher than that after the bedroom had been cleaned. For P. aeruginosa, 33% of isolates were multiresistant to antibiotics; 50% of the colonised patients had the same P. aeruginosa strain in their sputum as in air taken from their bedroom. P. aeruginosa was detected in 23% of samples taken from the surfaces in the showers after patient washing. Very low levels of pathogenic bacteria were found in samples from the other locations. Overall, activities with the highest risk of contamination in the CF ward are physiotherapy and washing in the communal shower room. We therefore recommend to open windows after physiotherapy and to implement a strong decontamination after showers.