Defining groundwater dependence and water-use requirements of terrestrial vegetation represents a significant challenge to water-resources managers. Terrestrial vegetation may exhibit complex spatial and temporal patterns of groundwater dependence. In this study we have assessed the sources of water used by dominant tree species in remnant vegetation of Pioneer Valley, Mackay, in northern Queensland. Water use by tree species was determined by sapflow techniques and the sources of water were investigated by using a combination of isotopic and water-potential measurements. Within the remnant vegetation communities of the Pioneer Valley there were complex patterns of water use and water-resource partitioning. However, all communities within the study showed some degree of groundwater use. Riparian communities that were reliant on groundwater discharge for maintenance of river baseflow exhibited high species diversity and complex forest structure and different species within these communities accessed a range of water sources including shallow soil water, river water and groundwater. In contrast, the woodlands and open forest were principally reliant on soil water. Although, species such as Corymbia clarksoniana appeared to be reliant on groundwater for their dry-season water-use requirements. This study demonstrated use of groundwater by remnant vegetation communities in the Pioneer Valley but determination of groundwater dependence requires a better understanding of the temporal patterns of water use and sources of water used by each species.