The water use of desert vegetation is a critical component of ecohydrological processes in arid regions, with groundwater and soil water serving as the primary moisture sources. To better understand the effects of ecological water conveyance on the water sources of desert vegetation, this study employed hydrogen and oxygen stable isotope techniques combined with the Bayesian mixing (MixSIAR) model to analyze changes in water uptake sources of Populus euphratica (of different ages) under varying groundwater depths during ecological water conveyance and non-conveyance periods. The results indicate: (1) The delta 18O and delta 2H values of soil water decreased during the water conveyance period compared to the non-conveyance period, gradually increased with distance from the riverbank, and decreased with soil depth. The delta 18O and delta 2H values of xylem water increased during the water conveyance period, with the highest values observed in young trees, followed by mature trees, and the lowest in intermediate-aged trees. The delta 18O and delta 2H values of groundwater decreased during the water conveyance period and declined with increasing distance from the riverbank. (2) During the water conveyance period (groundwater depth: 1.95-4.73 m), the primary water sources for P. euphratica of different ages were groundwater, river water, and deep soil water, with the maximum utilization proportion of groundwater reaching 23.1%. During the non-conveyance period (groundwater depth: 4.13-6.26 m), the main water sources shifted to groundwater and deep soil water, with the maximum groundwater utilization proportion dropping to 19.9%. As groundwater depth increased, mature trees exhibited the highest reliance on groundwater, followed by intermediate-aged trees, while young trees showed the lowest dependency. (3) The utilization proportion of soil water by P. euphratica increased with higher soil moisture and clay content but decreased with greater groundwater depth. Elevated soil salinity significantly inhibited water absorption by P. euphratica. This study elucidates the adaptive water use strategies of P. euphratica under fluctuating groundwater conditions during ecological water conveyance and non-conveyance periods, providing theoretical support for the ecological restoration of desert riparian forests in the lower Tarim River Basin.