Objective:The purpose of this study is to investigate the mental health status of public health nurses (PHNs) in public health centers (PHCs) and to clarify its related factors during the coronavirus disease 2019 (COVID-19) pandemic in Japan. Methods: Participants were 1,320 full-time PHNs in PHCs, and self-administered structured questionnaire regarding individual factors, work environment factors, and mental health status measured using the 12-item General Health Questionnaire (GHQ-12) was surveyed. The survey period was from June 10 to July 15, 2021. We asked the respondents for a response regarding the situation during the fourth wave of COVID-19 infectious spread. For analysis, participants were divided into two groups according to a cut-off point of the GHQ-12 score, and the χ 2 test was conducted. Afterward, stepwise logistic regression analysis was conducted. Results: The data of 640 participants were analyzed (valid response rate: 48.5%). Of the 640 participants, 64.2% worked in the infectious diseases section. Among the participants, 63.6% reported working overtime every day, and 38.3% reported working every weekend. Regarding the mental health status, the mean GHQ-12 score was 6.3±3.4, and 75.5% of the participants had a higher level of psychological distress. As shown in the result of the logistic regression analysis, the items "belonging to the infectious disease section" (OR = 1.73), "frequency of working overtime" (OR = 2.23), "frequency of working on the weekends" (OR = 2.11) and "workplace social support" (OR = 0.31) affected the PHNs' mental health. Conclusions: This study revealed that the mental health status of PHNs was at a critical level during the COVID-19 pandemic, and the needs for immediate mental health support for PHNs was suggested.