C 4 plants show higher photosynthetic capacity and resource use efficiency than C 3 plants. However, the genetic variations of these traits and their regulatory factors in C 4 plants still remain to be resolved. We investigated physiological, biochemical, and structural traits involved in photosynthesis and photosynthetic water and nitrogen use efficiencies (PWUE and PNUE) in 22 maize lines and four teosinte lines from various regions of the world. Net photosynthetic rate (P N) ranged from 32.1 to 46.5 μmol m −2 s −1. P N was positively correlated with stomatal conductance, transpiration rate, and chlorophyll, nitrogen and soluble protein contents of leaves, but not with specific leaf weight. P N was positively correlated with the activities of ribulose-1,5-bisphosphate carboxylase/oxygenase and the C 4-acid decarboxylases, NADP-malic enzyme and phosphoenolpyruvate carboxykinase, but not with the activity of phosphoenolpyruvate carboxylase. Leaf structural traits (stomatal parameters, leaf thickness, and interveinal distance) were not correlated with P N. These data suggest that physiological and biochemical traits are involved in the genetic variation of P N , but structural traits are not directly involved. PWUE is in the lower class of values reported for C 4 plants, whereas PNUE is in the highest class of values reported for C 4 plants. PNUE was negatively correlated with leaf nitrogen content but not significantly correlated with P N. PWUE was not correlated with δ 13 C values of leaves, indicating difficulty in using δ 13 C values as an indicator of PWUE of maize. In general, teosinte lines showed lower P N but higher PWUE than maize lines.