Study objective: Assess the changes in anxiety, depression, and stress levels over time and identify risk factors among healthcare workers in French emergency departments (EDs) during the first COVID-19 outbreak. Method: A prospective, multicenter study was conducted in 4 EDs and an emergency medical service (SAMU). During 3 months, participants completed fortnightly questionnaires to assess anxiety, depression, and stress using the Hospital Anxiety and Depression and the Chamoux-Simard scale. The changes in anxiety, depression, and stress levels over time were modelled by a linear mixed model including a period effect and a continuous time effect within periods. Results: A total of 211 respondents (43.5 %) completed the survey at inclusion. There was a decrease in mean anxiety (from 7.33 to 5.05, p < 0.001), mean depression (from 4.16 to 3.05, p = 0.009), mean stress at work (from 41.2 to 30.2, p = 0.008), and mean stress at home (from 33.0 to 26.0, p = 0.031) at the beginning of each period. The mean anxiety level was higher for administrative staff (+0.53) and lower for paramedics (-0.61, p = 0.047) compared to physicians. The anxiety level increased with the number of day and night shifts (0.13/day, p < 0.001, 0.12/night, p = 0.025) as did stress at work (1.6/day, p < 0.001, 1.1/night, p = 0.007). Reassigned healthcare workers were at higher risk of stress particularly compared to SAMU workers (stress at work: p = 0.015, at home: p = 0.021, in life in general: p = 0.018). Conclusion: Although anxiety, depression, and stress decreased over time, anxiety was higher among physicians and administrative staff. Reassignment and working hours were identified as potential risk factors for mental health distress in EDs. Trial registration: ClinicalTrials.gov NCT04383886.