The aim of our study was to assess university students’ sleep quality and fatigue before and during the academic exam period and identify potential associated factors. A Web-based survey was completed by 940 students of 20 different Tertiary Institutions including demographics, sleep habits, exercise, caffeine, tobacco, alcohol use, subjective sleep quality (Pittsburgh Sleep Quality Index—PSQI), and fatigue (Fatigue severity scale—FSS) at the beginning of the semester and during the examination period. During the exam period, PSQI (8.9 vs. 6.1, p < 0.001) and FSS scores (36.9 vs. 32.7, p < 0.001) were significantly elevated compared to the pre-exam period. An increase in the PSQI score was associated with age (β = 0.111, p = 0.011), presence of chronic disease (β = 0.914, p = 0.006), and depressive symptoms (β = 0.459, p = 0.001). An increase in the FSS score was associated with female gender (β = 1.658, p < 0.001), age, (β = 0.198, p = 0.010), increase in smoking (β = 1.7, p = 0.029), coffee/energy drinks consumption (β = 1.988, p < 0.001), decreased levels of physical exercise (β = 1.660, p < 0.001), and depressive symptoms (β = 2.526, p < 0.001). In conclusion, our findings indicate that exam periods have a negative impact on the sleep quality and fatigue levels of university students. Potential factors were identified that could contribute to the formulation of strategies for improved sleep quality and wellness.