Although fatigue has been implicated in anterior cruciate ligament (ACL) injury, few researchers have examined the timing of injury across a competitive sport season or within a competitive session to gain insight into the potential effects of fatigue on the incidence of ACL injury. To identify the time segments across a competitive season or within an individual competition associated with the greatest ACL injury incidence. Descriptive epidemiology study. Data from the National Collegiate Athletic Association Injury Surveillance Program for 2004-2005 through 2015-2016 for basketball, lacrosse, and soccer were analyzed. Incidence rate ratios (IRRs) compared ACL injury rates by sport, sex, season segment, and competition period. Poisson regression was used to examine the associations between each of these categories and the incidence of ACL injury as well as interaction effects. During the early regular season, the incidence rate was elevated relative to the preseason (IRR = 1.86; 95% confidence interval [CI] = 1.27, 2.74), middle regular season (IRR = 1.48; 95% CI = 1.01, 2.15), late regular season (IRR = 1.56; 95% CI = 1.08, 2.27), and postseason (IRR = 2.20; 95% CI = 1.06, 4.56). A sport-by-season interaction indicated this effect was largely attributable to a higher incidence in the early season among lacrosse athletes. An interaction between season segment and competition period (P = .02) revealed a greater injury incidence before halftime in the early regular season (IRR = 0.38, 95% CI = 0.19, 0.76), but a greater incidence after halftime in the late regular season (IRR = 2.40, 95% CI = 1.15, 5.02). Fewer noncontact injuries occurred in soccer than in basketball or lacrosse. The ACL injury incidence was higher in the early part of the regular season, particularly among lacrosse athletes. Although the injury incidences before and after halftime were similar, further analyses of player time and time of injury within each half are necessary. Coaches and practitioners should be cognizant of the elevated injury incidence during the early season among lacrosse athletes. Future authors should consider more specific analyses to further elucidate the potential role of fatigue development in exacerbating the incidence of ACL injury in collegiate athletes both within games and across the season.