Enhanced ozone strongly reduces carbon sink strength of adult beech (Fagus sylvatica)--resume from the free-air fumigation study at Kranzberg Forest.

Ground-level ozone (O(3)) has gained awareness as an agent of climate change. In this respect, key results are comprehended from a unique 8-year free-air O(3)-fumigation experiment, conducted on adult beech (Fagus sylvatica) at Kranzberg Forest (Germany). A novel canopy O(3) exposure methodology was employed that allowed whole-tree assessment in situ under twice-ambient O(3) levels. Elevated O(3) significantly weakened the C sink strength of the tree-soil system as evidenced by lowered photosynthesis and 44% reduction in whole-stem growth, but increased soil respiration. Associated effects in leaves and roots at the gene, cell and organ level varied from year to year, with drought being a crucial determinant of O(3) responsiveness. Regarding adult individuals of a late-successional tree species, empirical proof is provided first time in relation to recent modelling predictions that enhanced ground-level O(3) can substantially mitigate the C sequestration of forests in view of climate change.

Effect of 3 years' free-air exposure to elevated ozone on mature Norway spruce (Picea abies (L.) Karst.) needle epicuticular wax physicochemical characteristics.

We examined the effect of ozone (O(3)) on Norway spruce (Picea abies) needle epicuticular wax over three seasons at the Kranzberg Ozone Fumigation Experiment. Exposure to 2x ambient O(3) ranged from 64.5 to 74.2 microl O(3) l(-1) h AOT40, and 117.1 to 123.2 nl O(3) l(-1) 4th highest daily maximum 8-h average O(3) concentration. The proportion of current-year needle surface covered by wax tubes, tube aggregates, and plates decreased (P=0.011) under 2x O(3). Epistomatal chambers had increased deposits of amorphous wax. Proportion of secondary alcohols varied due to year (P=0.004) and O(3) treatment (P=0.029). Secondary alcohols were reduced by 9.1% under 2x O(3). Exposure to 2x O(3) increased (P=0.037) proportions of fatty acids by 29%. Opposing trends in secondary alcohols and fatty acids indicate a direct action of O(3) on wax biosynthesis. These results demonstrate O(3)-induced changes in biologically important needle surface characteristics of 50-year-old field-grown trees.

Photosynthesis, chloroplast pigments, and antioxidants in Pinus canariensis under free-air ozone fumigation.

High O3 levels, driving uptake and challenging defense, prevail on the Canary Islands, being associated with the hot and dry summers of the Mediterranean-type climate. Pinus canariensis is an endemic conifer species that forms forests across these islands. We investigated the effects of ozone on photosynthesis and biochemical parameters of P. canariensis seedlings exposed to free-air O3 fumigation at Kranzberg Forest, Germany, where ambient O3 levels were similar to those at forest sites in the Canary Islands. The twice-ambient O3 regime (2 x O3) neither caused visible injury-like chlorotic or necrotic spots in the needles nor significantly affected violaxanthin, antheraxanthin and zeaxanthin levels and the de-epoxidation state of the xanthophyll cycle. In parallel, stomatal conductance for water vapour, net photosynthesis, intercellular CO2 concentration, chlorophyll fluorescence parameters, as well as antioxidant levels were hardly affected. It is concluded that presently prevailing O3 levels do not impose severe stress on P. canariensis seedlings.

Free-air exposure systems to scale up ozone research to mature trees.

Because seedlings and mature trees do not necessarily respond similarly to O(3) stress, it is critically important that exposure systems be developed that allow exposure of seedlings through to mature trees. Here we describe three different O(3) Free-Air Exposure Systems that have been used successfully for exposure at all growth stages. These systems of spatially uniform O(3) release have been shown to provide reliable O(3) exposure with minimal, if any, impact on the microclimate. This methodology offers a welcome alternative to chamber studies which had severe space constraints precluding stand or community-level studies and substantial chamber effects on the microclimate and, hence physiological tree performance.

Evidence that branch cuvettes are reasonable surrogates for estimating O3 effects in entire tree crowns.

Within the scope of quantifying ozone (O(3)) effects on forest tree crowns it is still an open question whether cuvette branches of adult trees are reasonable surrogates for O(3) responses of entire tree crowns and whether twigs exhibit autonomy in defense metabolism in addition to carbon autonomy. Therefore, cuvette-enclosed branches of mature beech (Fagus sylvatica) trees were compared with branches exposed to the same and different ozone regimes by a free-air fumigation system under natural stand conditions by means of a VICE VERSA experiment. For this purpose, cuvettes receiving 1 x O(3) air were mounted in trees exposed to 2 x O(3) and cuvettes receiving 2 x O(3) air were mounted in trees exposed to 1 x O (3) in the upper sun crown. At the end of the fumigation period in September 2004, leaves were examined for differences in gas exchange parameters, pigments, antioxidants, carbohydrates, and stable isotope ratios. No significant differences in foliar gas exchange, total carbohydrates, stable isotope ratios, pigment, and antioxidant contents were found as a consequence of cuvette enclosure (cuvette versus free-air branches) of the same O(3) concentrations besides increase of glucose inside the cuvettes and reduction of the de-epoxidation state of the xanthophyll cycle pigments. No significant ozone effect was found for the investigated gas exchange and most biochemical parameters. The total and oxidized glutathione level of the leaves was increased by the 2 x O(3) treatment in the cuvette and the free-air branches, but this effect was significant only for the free-air branches. From these results we conclude that cuvette branches are useful surrogates for examining the response of entire tree crowns to elevated O(3) and that the defence metabolism of twigs seems to be at least partially autonomous.

Competitive strategies in adult beech and spruce: space-related foliar carbon investment versus carbon gain.

In Central Europe, Fagus sylvatica and Picea abies represent contrasting extremes in foliage type, crown structure and length of growing season. In order to examine the competitive strategies of these two co-occurring species, we tested the following hypotheses: (1) the space occupied by the foliage of sun branches is characterized by greater foliar mass investment compared to shade branches, (2) the carbon (C) gain per unit of occupied space is greater in sun than in shade branches, and (3) annual C and water costs of the foliage for sustaining the occupied space are low, wherever C gain per unit of occupied space is low. These were investigated in a mature forest in Southern Germany. The examination was based on the annual assessment of space-related resource investments and gains of the foliage. The foliated space around branches was regarded as the relevant volume with respect to aboveground resource availability. Occupied crown space per standing foliage mass was higher in shade compared to sun branches of beech, whereas no difference existed in crown volume per foliage mass between sun and shade branches of spruce (hypothesis 1 accepted for beech but rejected for spruce). However, beech occupied more space per foliage mass than spruce. The C gain per occupied crown volume was greater in sun than in shade branches (hypothesis 2 accepted) but did not differ between species. The amount of occupied space per respiratory and transpiratory costs did not differ between species or between sun and shade branches. In beech and spruce, the proportion of foliage investment in the annual C balance of sun and shade branches remained rather stable, whereas respiratory costs distinctly increased in shade foliage. Hence, shade branches were costly structures to occupy space, achieving only low and even negative C balances (rejection of hypothesis 3), which conflicts with the claimed C autonomy of branches. Our findings suggest that competitiveness is determined by the standing foliage mass and the annual branch volume increment rather than annual investments in foliage. Expressing competitiveness in terms of space-related resource investments versus returns, as demonstrated here, has the potential of promoting mechanistic understanding of plant-plant interactions.

Comparison of ozone uptake and sensitivity between a phytotron study with young beech and a field experiment with adult beech (Fagus sylvatica).

Chamber experiments on juvenile trees have resulted in severe injury and accelerated loss of leaves along with reduced biomass production under chronically enhanced O3 levels. In contrast, the few studies conducted on adult forest trees in the field have reported low O3 sensitivity. In the present study, young beech in phytotrons was more sensitive to O3 than adult beech in the field, although employed O3 regimes were similar. The hypotheses tested were that: (1) differences in O3 uptake were caused by the ontogenetically higher stomatal conductance of young compared to adult trees, (2) the experimental settings in the phytotrons enhanced O3 uptake compared to field conditions, and (3) a low detoxification capacity contributes to the higher O3 sensitivity of the young trees. The higher O3 sensitivity of juvenile beech in the phytotrons is demonstrated to relate to both the experimental conditions and the physiological responsiveness inherent to tree age.

Tree age dependence and within-canopy variation of leaf gas exchange and antioxidative defence in Fagus sylvatica under experimental free-air ozone exposure.

We characterized leaf gas exchange and antioxidative defence of two-year-old seedlings and 60-year-old trees of Fagus sylvatica exposed to ambient (1 x O3) or two-fold ambient (2 x O3) O3 concentrations (maximum of 150 ppb) in a free-air canopy exposure system throughout the growing season. Decline in photosynthesis from sun-exposed to shaded conditions was more pronounced in adult than juvenile trees. Seedling leaves and leaves in the sun-exposed canopy had higher stomatal conductance and higher internal CO2 concentrations relative to leaves of adult trees and leaves in shaded conditions. There was a weak overall depression of photosynthesis in the 2 x O3 variants across age classes and canopy positions. Pigment and tocopherol concentrations of leaves were significantly affected by canopy position and tree age, whereas differences between 1 x O3 and 2 x O3 regimes were not observed. Glutathione concentrations were significantly increased under 2 x O3 across both age classes and canopy levels. Seedlings differed from adult trees in relevant physiological and biochemical traits in ozone response. The water-soluble antioxidative systems responded most sensitively to 2 x O3 without regard of tree age or canopy position.

Growth of adult Norway spruce (Picea abies [L.] Karst.) and European beech (Fagus sylvatica L.) under free-air ozone fumigation.

This study attempted to detect the impact of ozone on adult trees of Norway spruce ( Picea abies [L.] Karst.) and European beech ( Fagus sylvatica L.) in an experimental mixed stand in Southern Bavaria, Germany. The aim was to examine whether there is a decrease in growth when trees are exposed to higher than atmospheric concentrations of ozone. This exposure was put into effect using a free-air fumigation system at tree crown level. Growth analysis was carried out on a group of 47 spruce and 36 beech trees, where radial stem increment at breast height - a sensitive index for stress - was measured. The ozone monitoring system allowed values to be obtained for the accumulated ozone exposure (SUM00) of each individual tree, so that their radial increment over three years could be correlated with the corresponding ozone exposure for the same time period. Correlation and regression analysis were then carried out to test the influence of ozone on diameter increment. In both spruce and beech, the initial stem diameter was the most influential factor on radial increment in the following year. A linear model was applied, including the diameter of the preceding year and the ozone exposure of the current year as predicting factors. For spruce trees, a significant negative influence of ozone exposure was found. In contrast, no significant ozone effect on diameter increment of beech was detected. The effect of ozone stress on a large spruce tree can lead to a decrease in potential radial increment of 22 %. The results are discussed in relation to other stress factors such as drought and lack of light.

Beech leaf colonization by the endophyte Apiognomonia errabunda dramatically depends on light exposure and climatic conditions.

Ozone and light effects on endophytic colonization by Apiognomonia errabunda of adult beech trees (Fagus sylvatica) and their putative mediation by internal defence compounds were studied at the Kranzberg Forest free-air ozone fumigation site. A. errabunda colonization was quantified by "real-time PCR" (QPCR). A. errabunda-specific primers allowed detection without interference by DNA from European beech and several species of common genera of plant pathogenic fungi, such as Mycosphaerella, Alternaria, Botrytis, and Fusarium. Colonization levels of sun and shade leaves of European beech trees exposed either to ambient or twice ambient ozone regimes were determined. Colonization was significantly higher in shade compared to sun leaves. Ozone exhibited a marginally inhibitory effect on fungal colonization only in young leaves in 2002. The hot and dry summer of 2003 reduced fungal colonization dramatically, being more pronounced than ozone treatment or sun exposure. Levels of soluble and cell wall-bound phenolic compounds were approximately twice as high in sun than in shade leaves. Acylated flavonol 3- O-glycosides with putatively high UV-B shielding effect were very low in shade canopy leaves. Ozone had only a minor influence on secondary metabolites in sun leaves. It slightly increased kaempferol 3- O-glucoside levels exclusively in shade leaves. The frequently prominent hydroxycinnamic acid derivative, chlorogenic acid, was tested for its growth inhibiting activity against Apiognomonia and showed an IC50 of approximately 8 mM. Appearance of Apiognomonia-related necroses strongly correlated with the occurrence of the stress metabolite, 3,3',4,4'-tetramethoxybiphenyl. Infection success of Apiognomonia was highly dependent on light exposure, presumably affected by the endogenous levels of constitutive phenolic compounds. Ozone exerted only minor modulating effects, whereas climatic factors, such as pronounced heat periods and drought, were dramatically overriding.