Many of the socio-cultural lifestyle and dietary changes that take place during Ramadan may affect the risk of injury in athletes, but little evidence is available. The aim of the present study was to examine the effects over two consecutive years of the holy month of Ramadan on injury rates in 42 professional players of a Tunisian top-level professional soccer team. Players were retrospectively organized into fasting and non-fasting groups and monitored for 3 months: 4 weeks before Ramadan, during the month of Ramadan (4 weeks), and 4 weeks after Ramadan each year. During Ramadan, training started at 22.00 h. The circumstances (training/match) and mechanism of injury (traumatic/overuse) were recorded. No significant differences between the three periods were observed for weekly mean training load, training strain, training duration, and Hooper's Index (quality of sleep, and quantities of stress, delayed-onset muscle soreness, and fatigue). Compared with non-fasting players, fasters had a lower (P < 0.05) Hooper's Index and stress during and after Ramadan. No significant difference in injury rates was observed between fasting and non-fasting players. Nevertheless, the rates of non-contact (6.8 vs. 0.6 and 1.1) and training overuse (5.6 vs. 0.6 and 0.5) injuries were significantly higher in fasting players during the month of Ramadan than before or after Ramadan. In conclusion, Ramadan, along with the corresponding changes in nutritional habits, sleeping schedule, and socio-cultural and religious events, significantly increased overuse and non-contact injuries in fasting players despite the fact that the training load, strain, and duration were maintained.