Rooting differentiation between established trees and grasses has been well documented in savannas, but it remains unclear to what extent tree-grass rooting differences affect competition between newly established seedlings and grasses. To examine this question, a greenhouse experiment was conducted at the University of Missouri, USA. Twenty 3-mo-old seedlings each of two African savanna tree species (Acacia nigrescens and Colophospermum mopane) were grown for 8 mo with two crossed factors: grass competition and irrigation depth. Strong negative effects of grass competition on final seedling biomass and leaf photosynthetic and stomatal conductance occurred in both tree species, but no effects of irrigation depth were detected. There was a clear tree species by grass competition interaction, suggesting interspecific variation in competitive response. The results emphasize the importance of below-ground competition with grasses for physiological and morphological responses of tree seedlings, while minimizing the importance of tree-grass rooting depth differences as a factor in modulating the competitive response of trees to grasses at the seedling stage.