Responses of beech forests to N fertilization: C to N ratio, C and N isotope composition, structural and biochemical traits along the canopy vertical profile

Nitrogen (N) deposition induced by human activities can heavily afect the biogeochemical cycles of forest ecosystems. In this respect, long-term fertilization experiments are important tools to explore the responses of forests to altered N availability. In this vision, in two beech (Fagus sylvatica L.) forests of LTER and ICP-Forests Networks, we have simulated increased N deposition using a mineral fertilizer. The two experimental sites are located in Italy, Collelongo (Abruzzo) and Cansiglio (Veneto). In both forests, in suficiently structural homogeneous areas, nine to twelve 30 × 30 m plots were established and randomly assigned to treatments increasing the natural N deposition by 0 kg N ha-1 (control), 30 kg N ha-1 and 60 kg N ha-1. The fertilizer was distributed to the soil at the beginning, in the middle and at the end of the 2015 and 2016 growing seasons. For only the Cansiglio site, a fourth treatment was tested, distributing the lower dose (30 kg N ha-1) above the canopy. Leaves were sampled at the end of the 2016 growing season. A vertical light gradient along the canopy was identified in both forests and the efects of N fertilization on leaf C to N ratio (C:N), C and N isotope compositions (? 13C, ?15N) and structural and biochemical traits (LMA, pigments) were examined. Significant relationships were found between ?13C, LMA and N content per unit of leaf area in both sites, with higher values in Collelongo for both N and ?13C. A progressive 15N-enrichment was observed at all the light exposition levels with the increasing of fertilizer dose. In the Collelongo site, the highest N dose enhanced the N content per unit of leaf dry mass and decreased the C:N in the shade canopy leaves compared to control. Conversely, in the Cansiglio site the sun canopy leaves of fertilized plots showed a higher N content per unit area and a 13C-enrichment compared to control. Moreover, the fertilization above the canopy induced an increase of the C:N ratio. The correlations of these results with biochemical and structural data and possible morpho-physiological implications are under analysis. In conclusion, our study highlights a gradient of leaf physiological traits along the vertical canopy profile in response to diferent N supplies, depending on the site characteristics, fertilizer dose and distribution approach, causing a change in the N allocation strategy within the canopy and afecting important leaf ecophysiological traits at the forest ecosystem level.