Background: Patient safety is an essential component in the quality of patient care.  It remains a challenge for developing countries. Assessment of patient safety culture in healthcare organizations is becoming an integral component to improve patients' safety. Aim: To assess the perceptions of patient safety culture attitude among health-care workers in Beni-Suef University hospital.  Participants andMethods:A cross sectional study was conducted from January till December 2015 using the Hospital Survey on Patient Safety Culture (HSOPSC) self-administered questionnaire which includes 12 dimensions on 500  university hospital staff members in Beni-Suef University hospital with a response rate of 84.6% (423/500). Results:The overall mean positive response rate for patient safety in Beni-Suef University Hospital was 39.3%. Healthcare workers had lower perceptions about patient safety culture. Only two dimensions showed positivity above 50%. The highest positive mean score dimension was “Teamwork within units" (57.4%) while the lowest positive mean score dimension was “Frequency of events reporting" (23.2%). Patient safety grade in relation to the staff position showed significant difference between the studied groups (p=0.009). Conclusion and Recommendations:Healthcare workers appear to have a lower perception of patient safety culture. Training of healthcare workers on patient safety and broad based research are highly recommended in order to improve patient safety culture. Patient safety needs to be incorporated into the education of health professionals across the spectrum of healthcare. There should be a blame-free system for identifying threats to patient safety, sharing information and learning from events.