Objectives The aim of the present study was to explore nurse perceptions of safety climate in acute Australian hospitals. Methods Participants included 420 nurses who have worked on 24 acute wards from six Australian hospitals. The Safety Attitudes Questionnaire (SAQ) Short Form was used to quantify nurse perceptions of safety climate and benchmarked against international data. Generalised linear mixed models were used to explore factors that may influence safety climate. Results On average, 53.5% of nurses held positive attitudes towards job satisfaction followed by teamwork climate (50.5%). There was variability in SAQ domain scores across hospitals. The safety climate and perceptions of hospital management domains also varied across wards within a hospital. Nurses who had worked longer at a hospital were more likely to have poorer perceptions of hospital management (β=-5.2; P=0.014). Overall, nurse perceptions of safety climate appeared higher than international data. Conclusions The perceptions of nurses working in acute Victorian and New South Wales hospitals varied between hospitals as well as across wards within each hospital. This highlights the importance of surveying all hospital wards and examining the results at the ward level when implementing strategies to improve patient safety and the culture of safety in organisations. What is known about the topic? Prior studies in American nursing samples have shown that hospitals with higher levels of safety climate have a lower relative incidence of preventable patient complications and adverse events. Developing a culture of safety in hospitals may be useful in targeting efforts to improve patient safety. What does this paper add? This paper has shown that the perceptions of safety climate among nurses working in acute Australian hospitals varied between hospitals and across wards within a hospital. Only half the nurses also reported positive attitudes towards job satisfaction and teamwork climate. What are the implications for practitioners? Programs or strategies that aim to enhance teamwork performance and skills may be beneficial to improving the culture of safety in hospitals. Wards may also have their own safety 'subculture' that is distinct from the overall hospital safety culture. This highlights the importance of tailoring and targeting quality improvement initiatives at the ward level.