Background: Catheter-related blood stream infection (CRBSI) is considered a major challenge for health care providers working in intensive care units.Aim: This study aimed to evaluate and compare the knowledge of evidence-based guidelines for preventing catheter-related blood Stream infections among physicians and nurses working in critical care units in Jordan.Design: A cross-sectional survey design was utilized. Methods:The study was conducted in five intensive care units; 181 nurses and 68 physicians were recruited from four governmental hospitals and one universityaffiliated hospital. Results:The mean total knowledge score for physicians' knowledge was low at 2.6, SD 1.5 (out of 10). While the mean total score for nurses' knowledge was low at 3.3, SD 1.8 (out of 10). The independent t test shows that nurses had a significantly higher mean total knowledge score than physicians (P < .05). Conclusions:The current study demonstrates that Jordanian nurses and physicians working in intensive care units have a low level of knowledge of the guidelines for preventing CRBSI. This problem should be addressed through routine assessment and evaluation and adoption and policies that make nurses and physicians comply with the evidence-based guidelines. Finally, conducting regular educational programmes on CRBSI prevention, and evaluation of compliance with the guidelines in daily clinical practice, are strongly recommended.Relevance to clinical practice: The results of the current study suggest the need for regular assessment of physicians' and nurses' knowledge and skills with regard to central venous catheter insertion and maintenance care. Based on this assessment and using the best available evidence, training programmes on the prevention of CRBSI should be developed. Courses might be online, face to face, or using high fidelity simulation. Finally, regular observation and evaluation of compliance with the CRBSI guidelines in daily clinical practice is needed.