BACKGROUND: The outbreak of COVID-19 has laid unprecedented psychological stress on health workers (HWs). We aimed to assess the immediate psychological impact on HWs at Tongji Hospital in Wuhan, China. METHODS: We conducted a single-center, cross-sectional survey of HWs via online questionnaires between February 8th and 10th, 2020. We evaluated stress, depression and anxiety by Impact of Event Scale-Revised (IES-R), Patient Health Questionnaire-9 (PHQ-9), and Generalized Anxiety Disorder 7-item (GAD-7), respectively. We also designed a questionnaire to assess the effect of psychological protective measures taken by Tongji Hospital. Multivariate logistic regression was used to identify predictors of acute stress, depression, and anxiety. RESULTS: We received 5062 completed questionnaires (response rate, 77.1 percent). 1509 (29.8 percent), 681 (13.5 percent) and 1218 (24.1 percent) HWs reported stress, depression and anxiety symptoms. Women (hazard ratio[HR], 1.31; P=0.032), years of working> 10 years (HR, 2.02; P<0.001), concomitant chronic diseases (HR, 1.51; P<0.001), history of mental disorders (HR, 3.27; P<0.001), and family members or relatives confirmed or suspected (HR, 1.23; P=0.030) were risk factors for stress, whereas care provided by hospital and department administrators(odds ratio [OR], 0.76; P=0.024) and full coverage of all departments with protective measures (OR, 0.69; P=0.004) were protective factors. CONCLUSIONS: Women and those who have more than 10 years of working, concomitant chronic diseases, history of mental disorders, and family members or relatives confirmed or suspected are susceptible to stress, depression and anxiety among HWs during the COVID-19 pandemic. Psychological protective measures implemented by the hospital could be helpful.