Many developing countries face various challenges in managing medical waste properly, and most of these challenges lie within the healthcare facilities. This study was conducted to investigate the challenges of effective management of medical waste in Vhembe District from the perspective of healthcare workers. The convergent parallel approach of mixed-method design was adopted. In-depth interviews were conducted with managers, infection prevention and control coordinators and environmental health practitioners of 15 public healthcare facilities in the district to gain insight into the challenges they identify in their respective healthcare facilities. A self-administered questionnaire was employed to obtain data from medical waste generators and handlers. Qualitative data were thematically analysed, and the Statistical Package for the Social Sciences version 25 was used to analyse the quantitative data. The challenges identified by healthcare workers include poor understanding of medical waste management guidelines and poor compliance; lack of regular training; poor attitudes of medical waste generators; insufficiency of waste management equipment and sub-standard central storage rooms. Most of the challenges were found to be linked to inadequate training of healthcare workers. For effective management of medical waste, efforts should be intensified towards adequate training of healthcare workers and provision of necessary equipment.