Differences in structural, physiological, and biochemical features between C 3 and C 4 species resulted in different wateruse efficiencies and different adaptations to climate. This paper aimed at investigating, at a late successional stage, the water-use efficiency of two forage species, Dichanthium ischaemum and Dasypyrum villosum, which exhibit different growth forms (perenial, annual) and photosynthetic mechanisms (C 4 and C 3 , respectively). The annual C 3 species Avena fatua, at an early successional stage, was included in our experiments to contrast its behaviour against D. villosum. The experiment was conducted during the growing season in low-elevation grasslands of North Greece. Midday leaf water potential, net photosynthetic rate, transpiration rate and stomatal conductance were measured. Instantaneous water-use efficiency (WUE) and intrinsic water-use efficiency (WUE i ) were calculated in D. ischaemum, D. villosum, and A. fatua. The results suggest that, under natural rainfall conditions, the annual C 3 grass species D. villosum exhibits a similar WUE with higher values of WUE i than the perennial C 4 species D. ischaemum at late stage of succession on the low elevation Mediterranean grasslands. Moreover, A. fatua at an early successional stage, exhibited different photosynthetic behaviour than D. villosum at a late successional stage. These findings indicate that the annual C 3 species D. villosum under drought and at a late successional stage seems to modify the WUE obtaining values similar to those of C 4 species. The extent to which the ecophysiological characteristics of D. villosum are environmentally or intrinsically determined remains to be answered.Additional key words: instantaneous water-use efficiency; intrinsic water-use efficiency; net photosynthetic rate; stomatal conductance; water stress.