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Fluorescence transient (FT) analysis (fast kinetics) was carried out on sun and shade leaves of beech seedlings in an experimental field at Curno (North Italy), during the 2008 growing season (June-August). The aim of the research was to determine: (1) morphology in sun leaves as long term acclimation to high light; (2) short term responses to high light, i.e. during the course of the day; (3) evolution of responses during the growing season and the ageing process. Sun leaves differ from shade leaves by their lower trapping capacity (expressed by the ratio FV/FM) and higher capacity to reduce end acceptors beyond PSI (expressed by phase I-P of the FT). These features were assessed at pre-dawn, when the entire plant is fully dark-adapted. Short term responses (differences between midday and pre-dawn assessments) occur especially in sun leaves, indicating that the photosynthetic machinery reacts to the high excitation pressure by increasing photochemical and non-photochemical de-excitation processes. Other responses concern the inactivation of the oxygen evolving system and a stability loss of the tripartite system reaction centre (RC) - harvesting light complex - core antenna, expressed by the onset of K and L-bands at midday. Sun leaves are well acclimated to high light from a structural and functional point of view, however, the irradiance at midday provokes conditions of instability in the photosynthetic machinery, possibly through reactive oxygen species (ROS) production at both PSII and PSI sides. The temporal patterns across the growing season indicate the progression of senescence processes, that are more pronounced in sun leaves which present a permanent (chronic) condition of photoinhibition. Differences between sun and shade leaves decrease over time. [ABSTRACT FROM AUTHOR]

Fluorescence transient (FT) analysis (fast kinetics) was carried out on sun and shade leaves of beech seedlings in an experimental field at Curno (North Italy), during the 2008 growing season (June–August). The aim of the research was to determine: (1) morphology in sun leaves as long term acclimation to high light; (2) short term responses to high light, i.e. during the course of the day; (3) evolution of responses during the growing season and the ageing process. Sun leaves differ from shade leaves by their lower trapping capacity (expressed by the ratio FV/FM) and higher capacity to reduce end acceptors beyond PSI (expressed by phase I–P of the FT). These features were assessed at pre-dawn, when the entire plant is fully dark-adapted. Short term responses (differences between midday and pre-dawn assessments) occur especially in sun leaves, indicating that the photosynthetic machinery reacts to the high excitation pressure by increasing photochemical and non-photochemical de-excitation processes. Other responses concern the inactivation of the oxygen evolving system and a stability loss of the tripartite system reaction centre (RC) – harvesting light complex – core antenna, expressed by the onset of K and L-bands at midday. Sun leaves are well acclimated to high light from a structural and functional point of view, however, the irradiance at midday provokes conditions of instability in the photosynthetic machinery, possibly through reactive oxygen species (ROS) production at both PSII and PSI sides. The temporal patterns across the growing season indicate the progression of senescence processes, that are more pronounced in sun leaves which present a permanent (chronic) condition of photoinhibition. Differences between sun and shade leaves decrease over time. [ABSTRACT FROM AUTHOR]

An experiment in open-top chambers was carried out in summer 2008 at Curno (Northern Italy) in order to study the effects of ozone and mild water stress on poplar cuttings (Oxford clone). In this experiment direct fluorescence parameters (JIP-test) were measured in leaves from different sections of the crown (L: lower; M: medium; U: upper parts of the crown). The parameters considered were calculated at the different steps of the fluorescence transient, and include maximum quantum yield efficiency in the dark-adapted state (F(v)/F(M)); the L-band, at 100 ∝ s, that expresses the stability of the tripartite system reaction centre-harvesting light complex-core antenna; the K-band, at 300 ∝ s, that expresses the efficiency of the oxygen-evolving complex; the J-phase, at 2 ms, that expresses the efficiency with which a trapped exciton can move an electron into the electron transport chain from Q(A)(-) to the intersystem electron acceptors; the IP-phase, which expresses the efficiency of electron transport around the photosystem 1 (PSI) to reduce the final acceptors of the electron transport chain, i.e., ferredoxin and NADP; and finally the performance index total (PItot) for energy conservation from photons absorbed by PSII to the reduction flux of PSI end acceptors. The main results are: (i) different dynamics were observed between leaves in the lower section, whose PItot decreased over time, and those in the upper sections in which it increased, with a dynamic connected to the leaf age; (ii) ozone depressed all the considered fluorescence parameters in basal leaves of well-watered plants, while it had little or no damaging effect on medium-level or upper-section leaves; (iii) PItot and IP-phase increased in upper leaves of plants subjected to ozone stress, as well as the net photosynthesis; (iv) water stress increased PItot of leaves in all levels of the crown. The results suggest that ozone-damaged poplar plants compensate, at least partially, for the loss of photosynthesis with higher photosynthetic rates in young leaves (in the upper section of the crown), more efficient to fix carbon.

This paper aims to investigate early responses to ozone in leaves of Fagus sylvatica (beech) and Populus maximowiczii x Populus berolinensis (poplar). The experimental setup consisted of four open-air (OA) plots, four charcoal-filtered (CF) open-top chambers (OTCs), and four nonfiltered (NF) OTCs. Qualitative and quantitative analyses were carried out on nonsymptomatic (CF) and symptomatic (NF and OA) leaves of both species. Qualitative analyses were performed applying microscopic techniques: Evans blue staining for detection of cell viability, CeCl3 staining of transmission electron microscope (TEM) samples to detect the accumulation of H2O2, and multispectral fluorescence microimaging and microspectrofluorometry to investigate the accumulation of fluorescent phenolic compounds in the walls of the damaged cells. Quantitative analyses consisted of the analysis of the chlorophyll a fluorescence transients (fast kinetics). The early responses to ozone were demonstrated by the Evans blue and CeCl3 staining techniques that provided evidence of plant responses in both species 1 month before foliar symptoms became visible. The fluorescence transients analysis, too, demonstrated the breakdown of the oxygen evolving system and the inactivation of the end receptors of electrons at a very early stage, both in poplar and in beech. The accumulation of phenolic compounds in the cell walls, on the other hand, was a species-specific response detected in poplar, but not in beech. Evans blue and CeCl3 staining, as well as the multispectral fluorescence microimaging and microspectrofluorometry, can be used to support the field diagnosis of ozone injury, whereas the fast kinetics of chlorophyll fluorescence provides evidence of early physiological responses.

Between 2004 and 2005 a combined open plot and open-top chamber (OTC) experiment was carried out at Curno (Northern Italy) with cuttings of the poplar clone Oxford (Populus maximowiczii Henry x Populus berolinensis Dippel) grown in open plots (OPs, ambient air), charcoal-filtered OTCs (CF, ozone concentration reduced to 50% of ambient) or non-filtered OTCs (NF, ozone concentration reduced to 95% of ambient). Plants in half of the chambers were kept well-watered (WET), and plants in the remaining chambers were not watered (DRY). The onset and development of visible foliar injury and the stomatal conductance to water vapor (g(w)) were assessed during each growing season. A stomatal conductance model was parameterized by the Jarvis approach, allowing the calculation of ozone stomatal fluxes of plants in each treatment. The pattern of visible symptoms was analyzed in relation to ozone exposure (AOT40, accumulated ozone over a threshold of 40 ppb) and accumulated ozone stomatal fluxes (AF(ST)). Symptoms became visible at an AOT40 between 9584 and 13,110 ppb h and an AF(ST) between 27.85 and 30.40 mmol O(3) m(-2). The development of symptoms was more widespread and faster in plants in WET plots than in DRY plots. A slightly higher dose of ozone was required to cause visible symptoms in plants in DRY plots than in WET plots. By the end of each growing season, plants in the CF OTCs had absorbed a high dose of ozone (31.60 mmol O(3) m(-2) in 2004 and 32.83 mmol O(3) m(-2) in 2005, for WET plots), without developing any visible symptoms. A reliable dose-response relationship was defined by a sigmoidal curve model. The shape of this curve expresses the change in leaf sensitivity and physiologic state over a prolonged ozone exposure. After the appearance of the first symptoms, foliar injury increased more rapidly than the increases in ozone exposure and ozone absorbed dose; however, when the injury incidence reached 75%, the plant response declined.

This paper compares the responses to ozone in five woody species: Fagus sylvatica (FS), Acer pseudoplatanus (AP), Fraxinus excelsior (FE), Viburnum lantana (VL) and Ailanthus altissima (AA). The hypothesis being tested was that the strategies that plants adopt to resist oxidative pressure are species-specific. The study was carried out on field grown plants in an area in Northern Italy characterized by elevated levels of ozone pollution. The observations were made both at ultrastructural (using light and electronic microscopy) and physiological (using chlorophyll a transient fluorescence and microspectral fluorometry) level. Common responses were: the hypersensitive response (i.e. the death of palisade mesophyll cells) and the formation of callose layers separating injured from healthy cells. FS and AP were capable of thickening the palisade mesophyll cell walls. This thickening process involved changes in cell wall chemical structure, evidenced by the accumulation of yellow autofluorescence compounds. Species-specific behaviours were observed with the fluorescence analysis, with special reference to the photochemical de-excitation constant (Kp). This value increased in FE and AP, and decreased in AA. The observed responses are interpreted as adaptative strategies against the ozone stress. The increase of Kp indicates that the reaction centres were working as more effective quenchers.

Potted plants of Ailanthus altissima, produced by root suckers coming from a single symptomatic mother tree, were placed in two sites in the vicinity of Florence (central Italy), with different levels of ozone pollution. These plants were kept in well watered conditions during the period May-September 1999. In the high pollution site (Settignano-SET) the level of ozone exposure (AOT40) reached at the end of the season a value of 31 ppm h, whereas in the "low pollution" site (Cascine-CAS) the exposure to ozone was 11 ppm h. A. altissima showed foliar symptoms in early July at SET and in the second half of July at CAS when exposure values reached 5 ppm h at both sites. However, at the end of August the conditions of the plantlets were rather similar in both sites. Microscopic and ultrastructural analysis were performed at the first onset of symptoms at SET (the CAS leaflets were asymptomatic). Observing the upper leaf surface where the brown stipples were visible, it was found that the cells of the palisade mesophyll displayed loss of chlorophyll and the organelles in the cytoplasm were damaged. Swelling of thylacoids was observed in the CAS leaflets, thus indicating the possible onset of a pre-visual damage. The injured cells were separated from the healthy ones by a layer of callose. We conclude that the sensitivity to ozone of A. altissima leaves is related to its leaf structure, with low leaf density and large intercellular spaces. Cell walls, as well as acting as mechanical barriers against the spread of ozone within the cell, also provide important detoxifying processes.