Daytime leaf water use efficiency does not explain the relationship between plant N status and biomass water-use efficiency of tobacco under non-limiting water supply

The increasing probability of seasonal droughts and freshwater scarcity emphasizes the importance of crop traits such as water-use efficiency (WUE) and its relation to nutrient management. In an earlier study using soil substrate in a pot experiment, we reported significant positive effects of N supply on biomass WUE of tobacco. However, there was a debate that the latter may be due to indirect effects of N supply (hidden drought), most likely because plants supplied with adequate N generally have greater biomass and hence faster depletion of soil water in the root zone. In pot and in field situation, therefore, it is difficult to relate any variation in leaf or biomass WUE to the direct effect of N supply. In this context, the aim of the current study was to re-examine to what extent N fertilization directly affects biomass WUE and related parameters under non-limiting water supply in hydroponics. About 2 weeks after the transfer of tobacco seedlings into nutrient solution containing 2 mM N, plants were treated with high-N or low-N in the form of NH4NO3. A marked decrease in CO2 assimilation with low-N supply compared to high-N supply was measured already 5 d after onset of treatments (DAO). In contrast, three different experiments clearly showed that stomatal conductance (g(s)) remained almost constant until 5 DAO resulting in significantly lower leaf WUE under low-N compared to high-N. Leaf WUE decreased gradually (up to 42% lower leaf WUE) at later stages. Surprisingly, biomass WUE and whole-plant C-13 values were not affected by N supply at any harvest date, which is in contrast to our earlier report where we observed clearly positive effects of N supply on biomass WUE of the same tobacco variety in a pot experiment with soil substrate. Night-time respiration and transpiration rates (measured by gas exchange and thermal imaging) were significantly higher with high-N supply than with low-N supply. The data show that 48.6% and 9.8% of the beneficiary effect of N on daytime leaf WUE were lost when nocturnal stomatal conductance and night-time respiration of the same leaves were taken into consideration. Thus, we conclude that earlier reports showing positive effects of N supply on biomass WUE and C-13 values in soil or field experiments may be due to indirect effect of N supply (e.g., hidden/mild drought).