Fitness is determined by the ability of an organism to both survive and reproduce; however, the mechanisms that lead to increased survival may not have the same effect on reproductive success. We used nineteen natural Drosophila melanogaster genotypes from the Drosophila Genetic Reference Panel to determine if adaptive plasticity following short-term acclimation through rapid cold-hardening (RCH) affects mating behavior and mating success. We confirmed that exposure to the acclimation temperature is beneficial to survival following cold stress; however, we found that this same acclimation temperature exposure led to less efficient male courtship and a significant decrease in the likelihood of mating. Cold tolerance and the capacity to respond plastically to cold stress were not correlated with mating behavior following acclimation, suggesting that the genetic control of the physiological effects of the cold temperature exposure likely differ between survival and behavioral responses. We also tested whether the exposure of males to the acclimation temperature influenced courtship song. This exposure again significantly increased courtship duration; however, courtship song was unchanged. These results illustrate costs of short-term acclimation on survival and reproductive components of fitness and demonstrate the pronounced effect that short-term thermal environment shifts can have on reproductive success.