In southeastern Australia, the overstory species Eucalyptus regnans F. Muell. commonly grows with either of the two leguminous understory trees, Acacia melanoxylon (R. Br. Ex Ait. f.) or Acacia dealbata (Link.). Our objective was to elucidate interactions between the dominant eucalypt and its companion acacias for nitrogen (N) sources. Use of stable N isotopes as tracers revealed that ammonium was the preferred soil N source for all species, nevertheless, total N uptake varied greatly among species. Studies with double-labeled ((13)C/(15)N) glutamine indicated the uptake of this form of organic N in small amounts by both E. regnans and the Acacia spp. These and other data imply that, in contrast to boreal forests, organic N is not a significant component of N nutrition in mountain ash forests. Field and laboratory studies provided evidence that N(2)-fixation capacity of acacias varies with stand development, with N-fixing species playing an important role in N nutrition during the early but not the mature stages of forest growth. An index of N-uptake efficiency - the amount of oxygen consumed per unit N taken up - was compared across four N sources and three species. Nitrate uptake was the least efficient form of N acquisition, especially compared with ammonium uptake which was up to 30-fold less costly. Efficiency of glutamine uptake was intermediate between that of ammonium and nitrate. Differences in uptake efficiency among N forms were most pronounced for the Acacia spp. and least for E. regnans. We conclude that an overlap in requirements among sympatric Acacia spp. and E. regnans for specific soil N sources can be bypassed because of changes in biochemical strategies of Acacia spp. triggered by increasing soil N concentrations during stand development. Further studies might elucidate whether this is a common feature of complex forest ecosystems, or a specialty of the interaction between eucalypts and acacias.