Today’s flight operations work on a pressurised 24/7 timetable as a result of the unrelenting escalation in international long-haul, short-haul, regional and overnight flights within commercial aviation. Air traffic around the world has doubled every 10 years for the last 30 years. Whilst there have been considerable advancements in aviation technology and operational demands, the human operators need for sleep remains. Commercial airline pilots are presently highly suspectible to sleep loss and fatigue due to these demanding, round-the- clock requirements. Whilst flight and duty time limitation regulations are in place to prevent pilot fatigue, they are not based on sound scientific evidence regarding their ability to do so. Furthermore, various European-based investigations have reported very high levels of sleep disruption and fatigue in European cockpits. Therefore, this study aimed to examine the influence of sleep deprivation and associated fatigue on incidents in flight and mental health and to investigate its effects on performance in commercial airline pilots. Firstly, this research found a critical pathway from duty hours through to self-reported incidents in flight with sleep disruption and feelings of fatigue in the cockpit found to be key factors contributing to this pathway (Study 1). This research also found very high incidences of self-reported sleep disturbance, feelings of fatigue in the cockpit, and consequential errors and incidents in flight as a result of fatigue. Further to this, self-reported sleep disruption and feelings of fatigue were also found to significantly influence pilots’ self-reported perceived depression or anxiety with those who reported higher incidences of sleep disturbance and fatigue being more likely to report feeling depressed or anxious (Study 2). Additionally, 24 hours’ sleep deprivation and subsequent fatigue was found to significantly impair mood and airline pilot core competencies, specifically cognitive flexibility, hand-eye coordination, multi-tasking ability, sustained attention, problem-solving, situation awareness and perceived workload with significant impairments becoming evident following 20 hours of continuous wakefulness (Study 3 & 4). Flying performance was not significantly impaired. Sleep disruption and fatigue is a highly serious and prevalent problem in European cockpits. It negatively impacts flight safety and pilot mental health and well-being. Further investigation in to the current flight and duty time limitation regulations as well as in to potential measures which could act as early detection and warning indicators of declining performance, as a result of sleep loss and fatigue, would enhance flight safety and promote pilot mental health and well-being.