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Riparian vegetation in arid regions is subject to frequent water environmental fluctuations and plays an important role in maintaining the riparian ecosystem. In recent decades, Populus euphratica (P. euphratica) has been threatened by decreasing groundwater levels, and large areas of P. euphratica seedlings are withered. To better understand the adaptive mechanism underlying hydrological process alterations, we designed an experiment to identify the water sources of P. euphratica seedlings using δ 18 O values (plant stem xylem, soil water and groundwater) under different groundwater scenarios by simulating the wild habitats conditions in the Tarim River bank of Central Asia. Seedlings were grown in lysimeters (2.0 m tall × 0.4 m inner diameter) under varying hydrological conditions. P. euphratica seedlings generally took up water from the shallow soil layer (0–80 cm). Under the T. ramosissima disturbance, the water source depths of the P. euphratica seedlings moved down as the groundwater depth increased. With increasing groundwater depth, the proportions of groundwater used by P. euphratica seedling monocultures (only P. euphratica seedlings) and P. euphratica seedling mixtures ( P. euphratica and T. ramosissima seedlings) were reduced. Under shallow groundwater treatment (W 1 : 25 cm), the plant height growth of the P. euphratica seedlings slowed, and their biomass accumulation decreased. The aboveground and total biomasses of the non-coexistence seedlings under W 2 (75 cm) treatment were maximized, while the maximum value for coexistence seedlings occurred under W 3 (125 cm) treatment. Thus, we suggest that shallow groundwater depth is not beneficial to P. euphratica seedling growth, and appropriately decreasing the groundwater depth may promote their growth. Such information will be valuable to provide the configuration of water resources for a typical river basin in an arid region of Central Asia based on the ecological water requirements of desert riparian vegetation. [ABSTRACT FROM AUTHOR]