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Objective: To investigate the impact of coronavirus disease 2019 (COVID-19) on psychosocial and behavioral responses of the non-health care workforce and to evaluate transmission prevention behavior implementation in the workplace., Participants and Methods: We deployed the baseline questionnaire of a prospective online survey from November 20, 2020, through February 8, 2021 to US-based employees. The survey included questions on psychosocial and behavioral responses in addition to transmission prevention behaviors (e.g., mask wearing). Select questions asked employees to report perceptions and behaviors before and during the COVID-19 pandemic. Data were analyzed descriptively and stratified by work from home (WFH) percentage., Results: In total, 3607 employees from 8 companies completed the survey. Most participants (70.0%) averaged 90% or more of their time WFH during the pandemic. Employees reported increases in stress (54.0%), anxiety (57.4%), fatigue (51.6%), feeling unsafe (50.4%), lack of companionship (60.5%), and feeling isolated from others (69.3%) from before to during the pandemic. Productivity was perceived to decrease for 42.9% of employees and non-work-related screen time and alcohol consumption to increase for 50.7% and 25.1% of employees, respectively, from before to during the pandemic. Adverse changes were worse among those with lower WFH percentages. Most employees reported wearing a mask (98.2%), washing hands regularly (95.7%), and physically distancing (93.6%) in the workplace., Conclusion: These results suggest worsened psychosocial and behavioral outcomes from before to during the COVID-19 pandemic and higher transmission prevention behavior implementation among non-health care employees. These observations provide novel insight into how the COVID-19 pandemic has impacted non-health care employees., (© 2021 THE AUTHORS.)