ALBERT

Notes: Field data on the sulphur and cation budget of growing Norway spruce canopies (Picea abies [L.] Karst.) are summarized. They are used to test a spruce decline model capable of quantifying effects of chronic SO2 pollution on spruce forests. At ambient SO2 concentrations, acute SO2 damage is rare, but exposure to polluted air produces reversible thinning of the canopy structure with a half-time of a few years. Canopy thinning in the spruce decline model is highest (i) at elevated SO2 pollution, (ii) in the mountains, (iii) at unfertilized sites with poor K+, Mg2+ or Zn2+ supply, (iv) at low spruce litter decomposition rates, and (v) acidic, shallow soils at high annual precipitation rates in the field and vice versa. Model application using field data from Würzburg (moderate SO2 pollution, alkaline soils, no spruce decline) and from the Erzgebirge (extreme SO2 pollution, acidic soils in the mountains, massive spruce decline) predicts canopy thinning by 2–11% in Würzburg and by 45–70% in the Erzgebirge. The model also predicts different SO2-tolerance limits for Norway spruce depending on the site elevation and on the nutritional status of the needles. If needle loss of more than 25% (damage class 2) is taken to indicate ‘real damage’ exceeding natural variances, then for optimum soil conditions SO2 tolerance limits range from (27.3 ± 7.4) μg m−3 to (62.6 ± 16.5) μg m−3. For shallow and acidic soils, SO2 tolerance limits range from (22.0 ± 5.5) μg m−3 to (37.4 ± 7.5) μ m−3. These tolerance limits, which are calculated on an ecophysiological data basis for Norway spruce are close to epidemiological SO2-toIerance limits as recommended by the IUFRO, UN-ECE and WHO. The observed statistical regression slope of the plot (damaged spruce trees vs. SO2-pollution) in west Germany is confirmed by modelling (6% error). Model application to other forest trees allows deduction of the observed sequence of SO2-sensitivity: Abies 〉 Picea 〉 Pinus 〉 Fagus 〉 Quercus. Thus, acute phytotoxicity of SO2 seems not to be involved in ‘forest decline’. Chronic SO2-pollution induces massive canopy thinning of Abies alba and Picea abies only at unfavourable sites, where natural stress factors and secondary effects of SO2pollution act together to produce tree decline.

Notes: Regarding time ranges of years, a rationale has been developed which is capable of explaining observed ‘spruce decline’ symptoms observed when spruce is exposed to air containing ambient levels of SO2. It integrates and interrelates (i) ecophysiological data (tree morphology, assimilate partitioning, canopy turnover, senescence physiology, stomatal conductance, canopy throughfall, sulphur metabolism, tonoplast symport), (ii) pedological data (soil leaching, cation recycling, litter decomposition, forest nutrition), and (iii) meteorological data (site elevation, length of the annual trunk growth period, SO2-pollution). Furthermore, it can explain field observations at numerous sites of spruce decline in central Europe where SO2 is implicated as a factor of forest decline: (i) thinning of the canopy structure; (ii) early needle senescence; (iii) cation deficiency; (iv) low SO2 tolerance at sites with depleted soils in the mountains; (v) synergism of SO2pollution and acidic precipitation; (vi) recovery after liming, fertilization and after decreasing SO2 pollution; and (vii) higher SO2 tolerances of deciduous angiosperms. Different SO2tolerance strategies are identified that are employed by more SO2-tolerant tree species. Ecophysiological SO2tolerance factors interact in a complex synergistic or antagonistic manner. It is concluded that chronic SO2 pollution at ambient concentrations predisposes mainly evergreen gymnosperms to suffer under synergistic environmental stresses (frost, drought, pathogens, etc.). Thinning of the crown structure is massive at extreme sites, where several stresses act simultaneously on the trees (depleted soils, high SO2 pollution, acidic rain, etc.). Mathematical formulations allow precise definitions of terms such as cooperativity, synergism, antagonism, vitality, predisposition, latency, etc. This universal rationale, which is applicable to all tree species, is exemplified here for Norway spruce (Picea abies [L.] Karst.). Integration of parameters yields an ordinary differential equation, which can be solved analytically. It predicts reversible dynamics of crown structures and gives an ecophysiological background to‘damage’.

Notes: Abstract Using experimental information obtained in earlier studies on the permeabilities of mesophyll and guard-cell membranes to abscisic acid (ABA), and on stress-induced pH shifts in the apoplasm and in symplasmic compartments (Hartung et al., 1988, Plant Physiol. 86, 908–913; Hartung et al. 1990, BPGRG Monogr. 215–235), a mathematical model is presented which will permit computer analysis of the stress-induced redistribution of ABA amongst different leaf cell types (mesophyll, epidermis, guard cells, phloem cells) and their compartments (cell wall, cytosol, chloroplast stroma, vacuole). Metabolism and conjugation of ABA and its transport in the xylem and the phloem are also taken into consideration. We ask whether the stressinduced redistribution of ABA is fast and intensive enough to induce stomatal closure within a few minutes. The model can be adapted to any other weak acid or base, e.g. to other phytohormones (auxins, gibberellins), which differ from ABA, e.g. by their membrane conductances, anion permeabilities and pKa values. Our wholeleaf model can predict the time course and the compartmentation of, for example, phytohormone concentrations as a function of changing source-sink patterns (e.g. by compartmental pH shifts in the leaf lamina). An analysis of the present knowledge of the ABA physiology of leaves and studies on stress effects are presented in subsequent publications. In this communication we describe the whole-leaf model and present and discuss all necessary morphological (volumes, surfaces etc.) and physiological (pH, membrane conductances etc.) parameters of an unstressed leaf of Valerianella locusta L. We draw fundamental conclusions by comparing determined and calculated ABA concentrations in the leaf-cell compartments. We found that the model predictions are close to measured data, and we conclude that in unstressed leaves ABA is close to flux equilibrium amongst the different tissues and compartments.

Notes: Abstract Using a computer model written for whole leaves (Slovik et al. 1992, Planta 187, 14–25) we present in this paper calculations of abscisic acid (ABA) redistribution among different leaf tissues and their compartments in relation to stomatal regulation under drought stress. The model calculations are based on experimental data and biophysical laws. They yield the following results and postulates: (i) Under stress, compartmental pH-shifts come about as a consequence of the inhibition of the pH component of proton-motive forces at the plasmalemma. There is a decrease of net proton fluxes by about 8.6 nmol · s−1 · m−2. (ii) Using stress-induced pH-shifts we demonstrate how ‘stress intensities’ can be quantified on a molecular basis. (iii) As the weak acid ABA is the only phytohormone which behaves in vivo and in vitro ideally according to the Henderson-Hasselbalch equation, pH-shifts induce a complicated redistribution amongst compartments in the model leaf. (iv) The final accumulation of ABA in guard-cell walls is intensive: up to 16.1-fold compared with only up to 3.4-fold in the guard-cell cytosol. We propose that the binding site of the guard-cell ABA receptor faces the apoplasm. (v) A twoto three-fold ABA accumulation in guard-cell walls is sufficient to induce closure of stomata. (vi) The minimum time lag until stomata start to close is 1–5 min; it depends on the stress intensity and on the guard-cell sensitivity to ABA: the more moderate the stress is, the later stomata start to close or they do not close at all. (vii) In the short term, there is almost no influence of the velocity of pH-shifts on the velocity of the ABA redistribution, (viii) Six hours after the termination of stress there is still an ABA concentration 1.4-fold the initial level in the guard-cell cytosol (delay of ABA relaxation, ‘aftereffect’), (ix) The observed ‘induction’ of net ABA synthesis after onset of stress may be explained by a decrease in cytosolic ABA degradation. About 1 h after onset of stress the model leaf would start to synthesise ABA (and its conjugates) automatically, (x) This ABA net synthesis serves to ‘inform roots’ via an increased ABA concentration in the phloem sap. The stress-induced ABA redistribution is per se not sufficient to feed the ploem sap with ABA. (xi) The primary target membrane of ‘stress’ is the plasmalemma, not thylakoids. (xii) The effective ‘stress sensor’, which induces the proposed signal chain finally leading to stomatal closure, is located in epidermal cells. Mesophyll cells are not capable of creating a significant ABA signal to guard cells if the epidermal plasmalemma conductance to undissociated molecular species of ABA (HABA) is indeed higher than the plasmalemma conductance of the mesophyll (plasmodesmata open), (xiii) All model conclusions which can be compared with independent experimental data quantitatively fit to them. We conclude that the basic experimental data of the model are consistent. A stress-induced ABA redistribution in the leaf lamina elicits stomatal closure.

Notes: Abstract A computer model written for whole leaves and described in the preceding publication (Slovik et al. 1992, this volume) has been developed for calculating the distribution and fluxes of weak acids or bases amongst different leaf tissues and their compartments, considering membrane transport, transpiration-driven mass transport, symplasmic and apoplasmic diffusion, and metabolic turnover rates in specified compartments. The model is used to analyse flux equilibria and the transport behaviour of the phytohormone abscisic acid (ABA) in unstressed and stressed leaves. We compare experimental data of unstressed Valerianella locusta L. leaves and expectations based on the detailed analysis of the data. (i) The mean daily influx of ABA into the leaf lamina via the xylem sap is about 10 nmol · m−2 · day−1. It is balanced by the sum of an export of ABA via the phloem sap (0.7%), possibly also by a basipetal ABA transport in the petiole parenchyma of young leaves (up to 18%), by an irreversible conjugation of ABA (0.4–4%) and by net degradation of ABA in the leaf lamina (80–95%). (ii) The estimated kinetic parameters of this net degradation are for the mesophyll apoplasm: apparent K m = 3.7 nM and V max = 12.9 nmol · m−3 · s−1, or for the mesophyll cytosol: apparent K m = 8.1 nM and V max = 32.3 nmol · m−3 · s−1. (iii) The dynamic ABA concentration in the phloem sap of Valerianella is 2.8 nM. This is only 5.5% of the static ABA equilibrium concentration in excised leaves or 70% of the ABA concentration in the mesophyll apoplasm, and it equilibrates within a few hours after source concentrations in the mesophyll apoplasm are changed under stress. Thus, the phloem sap is a flexible medium for transporting ‘new phytohormone information’ from the lamina to the shoot and roots, (iv) Measured compartmental ABA concentrations are close to calculated equilibrium concentrations in unstressed leaves. We conclude that model calculations are close to reality, (v) pH gradients within the apoplasm influence the apoplasmic distribution of ABA. Its concentration is maximally about twofold higher in guard-cell walls relative to the mesophyll apoplasm. (vi) Unexpectedly, all compartmental equilibrium concentrations of ABA in the leaf lamina depend on plasmalemma conductances for undissociated ABA and on the transport properties of the plasmodesmata. This is a consequence of the cyclic diffusion pathway: mesophyll cytosol — mesophyll plasmalemma — mesophyll apoplasm — epidermal apoplasm — epidermal plasmalemma — epidermal cytosol — plasmodesmata — mesophyll cytosol (in this direction), if there are different apoplasmic or cytosolic pH values in both tissues. The cyclisation rate is 42 fmol · s−1 · m−2 leaf area, which corresponds to a turnover time = 11.0 h for the total ABA content within the leaf lamina. A decrease of the epidermal plasmalemma conductance by 90% yields a threefold ABA concentration in the guard-cell free space, (vii) Compartmental relaxation-time coefficients are estimated and summarised for all leaf tissues and its major compartments. They range from 1.5 min for chloroplasts up to 3.3 d for mesophyll vacuoles, (viii) The highest ABA concentration, which can be expected in any leaf compartment, is 7 mM in the guard-cell cytoplasm of certain plant species, (ix) We employed circadian changes (equal day + night, 12 h each = equinoctium) of the stromal pH ± 0.3 in C3 plants, and for Crassulacean acid metabolism (CAM) plants, additionally, vacuolar pH ± 2.5 changes, and calculated the consequences for ABA redistribution within the lamina. In plants of both photosynthesis types, the ABA concentration in guard-cell walls is only 1.5 times higher in the night relative to the day. We conclude that stomata may not be regulated by ABA in a night-day regime. The influence of the extreme vacuolar pH changes on ABA distribution is small in CAM plants for two reasons: the ABA content in CAM mesophyll vacuoles is low (maximum 2.7% of the total ABA mass per unit leaf area) and there is only a 6.5-fold increase of the mole fraction of undissociated ABA when the the vacuolar pH is lowered from 5.5 to 3.0 (importance of the absolute pKa = 4.75 of ABA).

Notes: Abstract Contents of organic sulfur, sulfate and the inorganic cations K+, Ca2+, Mg2+, Mn2+ and Na+ were compared in needles of three conifer species differing in tolerance to chronic SO2 immissions. Sulfate and organic sulfur compounds were also measured in bark and wood. Field material was collected from Norway Spruce (Picea abies (L.) Karst.), Colorado Spruce (Picea pungens Engelm.) and Scots Pine (Pinus sylvestris L.) at sites where the SO2 concentration in air was high, and at another site where it was low. In general, sulfate contents were higher but cation contents lower at the sites where SO2 concentrations were high than where they were low. Up to 114mmol · (kg DW)−1 sulfate was measured in fouryear-old needles of Norway Spruce from the Erzgebirge (annual mean of SO2 in air 32 nl · 1−1). Sulfate accumulation in this SO2-sensitive conifer increased with SO2 concentration in ambient air and with needle age, indicating that the main part of the sulfate resulted from the oxidative detoxification of SO2. Loss of inorganic cations from ageing needles was reduced, or cation levels even increased, with increasing needle age, while sulfate accumulated. Apparently, cations served as counter-ions for sulfate, which is sequestered in the vacuoles. Individual trees differed in regard to the nature of cations which accumulated with sulfate. Calcium, potassium and magnesium were the dominating cations. Sodium levels were very low. Needles of the SO2-tolerant conifers Colorado Spruce and Scots Pine growing next to Norway Spruce in the Erzgebirge did not accumulate, or accumulated less, sulfate with increasing needle age as compared to needles of Norway Spruce. However, somewhat more sulfate was found in the bark of the SO2-tolerant species than in the bark of Norway Spruce. Scots Pine contained distinctly more sulfate in the wood than the other conifers. Since accumulation of organic sulfur compounds could not be observed with increasing needle age, or in bark and wood, reduction does not appear to play a major role in the detoxification of SO2 by the investigated species. Physiological mechanisms permitting Colorado Spruce and Scots Pine to avoid the sulfate accumulation in the needles and the accompanying sequestration of cations that are observed in neighbouring Norway Spruce are discussed on the basis of the obtained data.

Notes: Abstract The emission of reduced volatile sulfur compounds from twigs of Norway spruce (Picea abies (L.) Karst.) was measured in the field by cryosampling and gaschromatographic analysis. Trees were growing in the Erzgebirge (E-Germany) at Oberbärenburg and at the Kahleberg and at a third stand in NW-Bavaria (S-Germany). Emission rates were also measured for Scotch pine (Pinus sylvestris L.) and Blue spruce (Picea pungens Engelm.) at the Kahleberg. Twigs still attached to the trees were enclosed in a flow-through gas exchange cuvette. H2S was detected as the predominant reduced sulfur compound emitted from the twigs. The mean H2S emission rate from twigs of Norway spruce varied between 0.04 pmol kg-1 dw s-1 at Würzburg and 6.21 pmol kg-1 dw s-1 at the Kahleberg. Comparing different species at the Kahleberg, the mean H2S emission rate was almost the same from twigs of Norway spruce (6.2 pmol kg-1 dw s-1) and Blue Spruce trees (5.9 pmol kg-1 dw s-1) but it was approximately 18 times higher for Scotch pine (110 pmol kg-1 dw s-1). The percentage of SO2-exclusion via H2S-emission of the tree species investigated at the Kahleberg is calculated on the basis of data on SO2 fluxes. It is very small for Norway spruce and Blue spruce. However, for Scotch pine, H2S emission contributes about 10% to the detoxification of SO2.

Notes: Abstract Monthly uptake rates and the annual deposition of gaseous SO2 via the stomata of six Norway spruce canopies (Picea abies (L.) Karst.) in Germany (Königstein im Taunus, Witzenhausen, Grebenau, Frankenberg, Spessart, Fürth im Odenwald) were calculated (i) from statistical response functions of stomatal aperture depending on meteorological data, and (ii) from the synchronously measured SO2 immission at these stands. The stomatal response functions had been derived on the basis of thorough stomatal water conductance measurements in the field. Calculations of the SO2 conductance of spruce twigs and SO2 uptake rates via stomata need continuously measured complete data sets of the (i) light intensity, (ii) air temperature, (iii) air humidity and (iv) SO2 concentration in spruce forests from all the year. These data were recorded half hourly in different German spruce forests. The apparent needle water vapour pressure difference and transpiration rates were calculated from meteorological data. Additional use of canopy through flow data in dry years allowed the estimation of the mean stomatal conductance for H2O and SO2 of whole spruce canopies. The annual SO2 uptake of a mean unit needle surface in spruce forests was 32% of the SO2 uptake rate of exposed needles at the top of spruce crowns. There is significant SO2 uptake all the year. The mean SO2 dose at all sites and years received through the stomata was (0.25±0.07) μmol SO2 m-2 (total needle surface) (nPa Pa-1)-1 (annual mean of SO2 immission; 1 nPa (SO2) Pa-1 (air) = 1 ppb) day-1 (vegetation period per year). Comparison of calculated SO2 uptake rates into needles with measured SO4 2- accumulation rates in needles from the mentioned sites and additionally from Würzburg, Schneeberg (Fichtelgebirge) and from three sites in the eastern Erzgebirge (Höckendorf, Kahleberg, Oberbärenburg) revealed that oxidative SO2 detoxification (SO4 2- formation) dominates only at sites with high SO2 immission and short vegetation periods. Under these conditions 70 to 90% of the annual stomatal SO2 uptake is detoxified via SO4 2- accumulation in needles. Cations are needed for neutralization of accumulating SO4 2- which are inavailable to support growth. Thus, SO2 induces a dominant and competitive additional nutrient cation demand, cation deficiency symptoms and enhanced needle loss (“spruce decline symptoms”) mainly at sites, where the ratio R=(SO2 immission): (length of the vegetation period) is higher than R=0.07 nPa Pa-1 day-1. Correlation analysis of the relative needle loss versus the SO2-dependent SO4 2- formation rate revealed a significant increase of needle loss at the 98% level (Student). At sites with small SO2 immission and long vegetation periods (R〈0.07 nPa Pa-1 day-1) reductive SO2 detoxification via growth (and/or phloem export of SO4 2-) is not kinetically overburdened. Under these conditions only 30% of the annual SO2 uptake is detoxified via SO4 2- formation and spruce decline is small or absent. On the basis of the critical value R≈0.07 nPa Pa-1 day-1 recommended SO2 immission limits can be deduced on a mere ecophysiological basis. These deduced values are close to the proposed SO2 immission limits of the IUFRO, WHO and the UNECE.

Notes: Abstract At six sites in central Germany consequences of SO2, NOX and O3 deposition and of acid precipitation on canopy throughfall of sulphate, nitrate, ammonium, organic acids and of metal cations from Norway spruce crowns were investigated in the field. Measured canopy throughfall rates (mmol ion kg-1 needle dw a-1 are separated in (i) “background” ion throughfall rates in clean air and (ii) trace gas-(or acid interception)-dependent throughfall rates at ambient trace gas concentrations. Based on synchronously measured pollution, precipitation and canopy throughfall data, statistical response functions are given, which allow the separate estimation of annual rates of sulphur and nitrogen deposition into spruce canopies if only annual means of SO2 or NO2 concentrations in air are known. The specific SO2 deposition rate of (0.841±0.214) mmol S kg-1 needle dw a-1 (nPa SO2 Pa-1)-1 is 2.3 times higher than the specific stomatal SO2 uptake. The NO2-dependent nitrogen deposition of (2.464±0.707) mmol N kg-1 needle dw a-1 (nPa NO2 Pa-1)-1 is 2.2 times higher than the specific stomatal NOX (NO2+NO) uptake. These ratios (2.3≈2.2) are explained by the percentage of annual hours with open needle stomata. The shape of observed “epicuticular” SO2 and NOX deposition curves and of stomatal SO2 and NOX uptake curves are congruent. As for stomatal NOX uptake, there is an apparent compensation point at (5 to 8) nPa NO2 Pa-1. There is significant SO2-dependent canopy throughfall of Ca〉K〉Al〉Mg〉Fe in this order of relative importance. NOX deposition in spruce canopies reduces K+ throughfall and it weakly promotes throughfall of Mn2+ and Zn2+. There was no significant codeposition of sulphate and ammonium and no ion exchange of intercepted H3O+ with nutrient cations at the measured ambient pH values of the precipitation water. In the presence of O3, throughfall of Mn2+ is reduced and throughfall of K+, Ca2+ and Al3+ is enhanced. In the cooperative presence of SO2, NO2 and O3 pollution in the field there is a 1.3-fold increase of the annual K+ demand and a 1.5-fold Mg2+ demand of spruce canopies relative to the situation in clean air. This trace gas-dependent additional cation demand of spruce canopies corresponds to a needle loss percentage of (23 to 33)% if the additional K+ and Mg2+ throughfall could not be recycled in spruce ecosystems. Observed canopy thinning ranges from (13 to 26)% at the investigated six spruce stands.

Notes: Abstract Jasmonic acid (JA) permeates the plasma membrane of mesophyll cells by diffusion as the lipophilic undissociated JAH molecule probably without the participation of a saturable uptake component. The mesophyll plasma membrane is nearly impermeable to the JA anion. The permeability coefficients of JA and several JA derivatives (its methyl ester (JAMe), 7-iso-cucurbic acid (7-iso-CA), 6-epi-7-iso-cucurbic acid (6-epi-7-iso-CA), and both stereoisomers of the JA leucine conjugate ((+)-JA-Leu and (-)-JA-Leu)) were determined and used in a simplified mathematical model to predict stressdependent JA redistribution between cytosol and apoplast in comparison with ABA. The redistribution of JA takes place similar to ABA; however, its velocity is much higher because of the high JA membrane permeability. When the permeability coefficients for the mesophyll plasma membrane are plotted double-logarithmically against the ratio of the distribution coefficient and the molecular ratio to the power of 1.5 (KDMr −1.5), two straight lines result for two different classes of compounds. The permeability coefficients of JA conjugates are lower than that of the free acid by approximately one order of magnitude, but they are still significantly higher than that of ABA.