As a framework for sequential decision-making, Reinforcement Learning (RL) has been regarded as an essential component leading to Artificial General Intelligence (AGI). However, RL is often criticized for having the same training environment as the test one, which also hinders its application in the real world. To mitigate this problem, Distributionally Robust RL (DRRL) is proposed to improve the worst performance in a set of environments that may contain the unknown test environment. Due to the nonlinearity of the robustness goal, most of the previous work resort to the model-based approach, learning with either an empirical distribution learned from the data or a simulator that can be sampled infinitely, which limits their applications in simple dynamics environments. In contrast, we attempt to design a DRRL algorithm that can be trained along a single trajectory, i.e., no repeated sampling from a state. Based on the standard Q-learning, we propose distributionally robust Q-learning with the single trajectory (DRQ) and its average-reward variant named differential DRQ. We provide asymptotic convergence guarantees and experiments for both settings, demonstrating their superiority in the perturbed environments against the non-robust ones.