Rationale and Aim: Patient safety is of great interest in health care organisations, worldwide. In Sweden, a national patient safety initiative was launched in 2011. The aim of this study was to examine and compare patient safety culture change over time from health care staffs’ perspective. A further aim was to examine factors that have had an effect on patient safety culture. Methods: Patient safety culture was assessed in 2009, 2011 and 2013 using the Swedish version of the Hospital Survey of Patient Safety Culture. Respondents in this study were registered nurses (n = 2149), enrolled nurses (n = 959), physicians (n = 355) and managers (n = 159) working in three health care divisions in a Swedish county council. Results: Patient safety culture decreased significantly over time for all but two dimensions. The dimension “Information to Patients/Relatives” was the only dimension to increase significantly over time. Health care staffs’ profession and health care division belonging had significant main effects on 14 and 8 dimensions, respectively. Managers and enrolled nurses scored patient safety culture significantly higher than registered nurses and physicians. Health care staff working in a mixed medical-surgical health care division scored significantly higher than those working in medical or surgical divisions did. Conclusions: Despite efforts for patient safety work in the county council, health care staffs’ perceptions of patient safety culture decreased over time. To improve hospital patient safety culture, managers and staff groups must meet and communicate with each other in order to create a common view of patient safety.