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Background Burnout is a widely recognised occupational phenomenon, with emergency department (ED) healthcare workers globally facing a heightened risk. Despite this, data on the burden of burnout among ED healthcare workers, especially in low-resource settings, are limited. Our study sought to establish the prevalence and factors associated with burnout and workplace stressors among ED healthcare workers in Uganda.Methods An online-based survey was conducted among ED professionals across four private and public hospitals. Burnout was assessed using the Maslach Burnout Inventory—Human Services Survey tool. Data were analysed using univariable and multivariable logistic regression analyses.Results Data from 82 participants were analysed (response rate of 88%), with approximately equal numbers of males and females, just over half providing care to both adult and paediatric patients and 61% working in public facilities. Overall, 9.7% of participants met the criteria for burnout. In total, 48.7% (n=40) reported high emotional exhaustion, 21.9% (n=18) reported high depersonalisation and 35.4% (n=29) reported low personal accomplishment. At least 80% of participants identified key ED stressors including work-related fatigue, patients’ financial problems, work overload, equipment shortages and challenges in balancing professional and personal responsibilities. After adjusting for covariates such as overload of health professional literature needed to be read, patients’ financial problems, educational issues, lack of sufficient clinical skills and ED violence, burnout was positively associated with poor communication with colleagues in the ED (adjusted OR (AOR) 9.90; 95% CI 1.60 to 61.17; p=0.014) and with caring for the old and terminally ill patients (AOR 7.54; 95% CI 1.38 to 41.29; p=0.020). These associations were consistent with the high depersonalisation domain of burnout.Conclusion Burnout, particularly in the emotional exhaustion domain, is notably prevalent among ED healthcare workers in Uganda. There is a pressing need for context-specific interventions aimed at improving early recognition of burnout and addressing persistent ED stressors. Such measures are essential to enhance ED healthcare workers’ well-being, and ultimately improve in-hospital emergency care in Uganda.