The standard process of central sterilization is crucial for the optimal functioning of the operating room. The outcome of this process is closely linked to the steps preceding the steps prior to the sterilization step itself. These steps include pre-disinfection carried out in the operating rooms and other stages, namely washing, drying and packaging, which must be performed in the central sterilization unit. In this context, this study aimed to describe the knowledge of the staff in the operating rooms and the central sterilization unit at Sahloul University Hospital in Sousse (Tunisia) in 2022, regarding the steps prior to the sterilization of reusable thermoresistant medical devices and to describe their practices in terms of compliance with these steps. A descriptive study was conducted from January 2022to June 2022with the aforementioned staff, using a self-administered questionnaire to assess their knowledge of the pre-sterilization steps and a direct observation audit of their practices with regard to these steps. Both measurement instruments were pre-tested. Out of 102self-administered questionnaires (knowledge assessment) distributed to the staff concerned, only 80were returned and correctly filled out, giving a response rate of 78.4%. Participants' responses regarding the order of steps prior to the sterilization were incorrect in 64% of cases. With regard to the evaluation of professional practices, 224observations were made in the study area (practice audit). In 82% of these observations, the pre-disinfection step was confused with the washing step. The use of Betadine brushes and scrubbing pads for device washing was noted in 89.3%, along with the absence of swabbing of the canals and hollow parts in 9.4% of cases and the absence of drying of the canals with compressed air. Mastery of the steps prior to sterilisation of reusable thermoresistant medical devices was insufficient in our institution, suggesting the importance of reinforcing the implementation of the procedure through a continuous training program followed by action plans.