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Work-related stress is a growing concern among healthcare professionals, with potential adverse effects on psychological well-being and quality of care. Midwifery practice, characterised by high emotional demands and frequent exposure to critical events, may increase vulnerability to occupational stress and psychological distress. This study aimed to evaluate perceived stress and psychological well-being among italian midwives and to compare hospital-based and community-based settings. We conducted a cross-sectional multicentric observational study on 175 midwives (140 hospital-based; 35 community-based). Perceived stress was assessed using the 10-item Perceived Stress Scale (PSS-10), and psychological well-being was evaluated with the Psychological General Well-Being Index (PGWBI). Hospital-based midwives reported significantly higher perceived stress than community-based midwives. A significant negative correlation between perceived stress and psychological well-being was found in both groups, with a stronger association observed in the hospital setting. The work setting significantly influences midwives' mental health, with hospital practice associated with higher stress and lower psychological well-being. These findings emphasize the urgent need for tailored organizational support, structural interventions, and policy initiatives to safeguard midwives' psychological health and enhance workforce retention.