Two-spotted spider mites (Tetranychus urticae Koch, 1836) are pests of vegetables, ornamentals, and row crops around the world. Two-spotted spider mites have become an important long-season pests of cotton, causing injury to cotton from an early vegetative stage. In the past eight years, Arkansas cotton acreage treated for spider mites has more than doubled and most of the increase has been attributed to early season infestations. Yield losses of up to 30% have been observed in other studies where spider mite infestation started at third true leaf. Because of the apparent change in this pest's population dynamics, particularly at early stages of crop development, the objective of the present study was to understand the impact of two spotted spider mites on cotton growth and yield. This project focuses on the impact of the timing and duration of infestations. Cotton plots were artificially infested at fourth, sixth true leaf in 2012, and at cotyledon and fourth true leaf during 2013. Both years included three infestation durations (short, medium, and long) at each infestation time. Two-spotted spider mites remaining on cotton at damaging densities for two weeks or more regardless of infestation time, caused significant yield loss. However, spider mites did not cause significant yield loss when environmental conditions did not favor spider mite development for extended periods.