Physiological effects of a long term exposure to low concentrations of NH3, NO2 and SO2 on Douglas fir (Pseudotsuga menziesii)

The above-ground parts of two years old seedlings of Douglas fir (Pseudotsuga menziesii) were exposed to filtered air, NH₃, NO₂₊, SO₂ (66, 96 and 95 μg m^?3, respectively), to a mixture of NO₂+NH₃ (55 + 82 μg m^?3) or SO₂+NO₂ (128 + 129 μg m^?3), for 8 months in fumigation chambers. Both chlorophyll fluorescence and gas exchange measurements were carried out on shoots which had sprouted at the beginning of the exposure period. The chlorophyll fluorescence measurements were performed after 3 and 5 months of exposure (average shoot age 70 and 140 days, respectively). Light response curves of electron transport rate (J) were determined, in which J was deduced from chlorophyll fluorescence. In addition, light response curves of net CO₂ assimilation were determined after 5 months of exposure. After 3 months of exposure (average shoot age 70 days) all exposure treatments showed a lower maximum electron transport rate (J_max) as compared to the control shoots (filtered air). A large reduction (45%) was observed for shoots exposed to SO₂+NO₂. During the exposure period between 3 and 5 months (average shoot age 70 and 140 days, respectively) a decrease of J_max was observed for all treatments. J_max had further declined some time after termination of the exposure, when average shoot age was 310 days. Shoots exposed to SO₂ and SO₂+NO₂ also showed a reduction in maximum net CO₂ assimilation (P_max) as compared to the control shoots. However, shoots exposed to NO₂ showed no reduction and even a higher P_max was observed for shoots exposed to NH₃ or NO₂+NH₃. Needles of these treatments also showed a higher chlorophyll content which might explain the contradictory results obtained for these treatments: the increased amount of photosynthetic units counteracts the reduction in J_max and consequently no reduction in P_max is measured. Shoots exposed to SO₂ and SO₂+NO₂ also showed a reduction in maximum stomatal conductance (g_s). However, the stomatal opening was larger than could be expected on basis of their (maximum) CO₂ assimilation rate. Consequently, water use efficiency of these shoots was lower than that of the control shoots. Also shoots exposed to NO₂ had a lower water use efficiency due to a significantly higher maximum g_s. Shoots exposed to NH₃ showed a high transpiration rate in the dark, indicating imperfect stomatal closure.     

Physiological effects of long-term exposure to low and moderate concentrations of atmospheric NH3 on poplar leaves

Abstract. Poplar shoots ( Populus euramericana L.) obtained from cuttings were exposed for 6 or 8 weeks to NH₃ concentrations of 50 and 100 μgm⁻³ or filtered air in fumigation chambers. After this exposure the rates of NH₃ uptake, transpiration, CO₂ assimilation and respiration of leaves were measured using a leaf chamber. During the long-term exposure also modulated chlorophyll fluorescence measurements were carried out to obtain information about the photosynthetic performance of individual leaves. Both fluorescence and leaf chamber measurements showed a higher photosynthetic activity of leaves exposed to 100 μg NH₃ m⁻³. These leaves showed also a larger leaf conductance and a larger uptake rate of NH₃ than leaves exposed to 50 μg m⁻³ NH₃ or filtered air. The long-term NH₃ exposure did not induce an internal resistance against NH₃ transport in the leaf, nor did it affect the leaf cuticle. So, not only at a short time exposure, but also at a long-term exposure NH₃ uptake into leaves can be calculated from data on the boundary layer and stomatal resistance for H₂O and ambient NH₃-concentration. Furthermore, the NH₃ exposure had no effect on the relation between CO₂-assimilation and stomatal conductance, indicating that NH₃ in concentrations up to 100 μg m⁻³ has no direct effect on stomatal behaviour; for example, by affecting the guard or contiguous cells of the stomata.     

Physiological effects of long term exposure to low concentrations of SO2 and NH3 on poplar leaves

Shoots of poplar (Populus euramericana L. cv. Flevo) were exposed to filtered air, SO₂, NH₃ or a mixture of SO₂ and NH₃ for 7 weeks in fumigation chambers. After this exposure gas exchange measurements were carried out using a leaf chamber. As compared to leaves exposed to filtered air, leaves pretreated with 112 μg m^?3 SO₂ showed a small reduction in maximum CO₂ assimilation rate (P_max) and stomatal conductance (g_s). They also showed a slightly higher quantum yield and dark respiration. In addition, the fluorescence measurements indicated that the Calvin cycle of the leaves pretreated with 112 μg m^?3 SO₂ was more rapidly activated after transition from dark to light. An exposure to 64 μg m^?3 NH₃ had a positive effect on P_max, stomatal conductance and NH₃ uptake of the leaves. This positive effect was counteracted by an SO₂ concentration of 45 μg m^?3. The exposure treatments appeared to have no effect on the relationship between net CO₂-assimilation and g_s. Also, no injury of the leaf cuticle or of epidermal cells was observed. Resistance analysis showed that NH₃ transfer into the leaf can be estimated from data on the boundary layer and stomatal resistance for H₂O transfer and NH₃ concentration at the leaf surface, irrespective of whether the leaves are exposed for a short or long time to NH₃ or to a mixture of NH₃ and SO₂. In contrast SO₂ uptake into the leaves was only partly correlated to the stomatal resistance. The results suggest a large additional uptake of this gas by the leaves. The possibility of a difference in path length between SO₂ and H₂O molecules is proposed.     

Detection and identification of gibberellins in Douglas fir (Pseudotsuga menziesii) shoots

Extracts of Douglas fir ( Pseudotsuga menziesii [Mirb.] Franco) shoots were purified by reversed and normal phase HPLC; gibberellin (GA)-like compounds detected by radioimmunoassay with antibodies against GA₄ and the Tan-ginbozu dwarf rice micro-drop biossay were analyzed by GC-MS. Three major components were identified as GA₄, GA₇, and GA₉ while smaller amounts of GA¹, GA₃ and putative GA⁹-glucosyl ester were also present.     

Uptake and translocation of manganese in seedlings of two varieties of Douglas fir (Pseudotsuga menziesii var. viridis and glauca)

Douglas fir (Pseudotsuga menziesii) variety glauca (DFG) but not the variety viridis (DFV) showed symptoms of manganese (Mn) toxicity in some field sites. We hypothesized that these two varieties differed in Mn metabolism. To test this hypothesis, biomass partitioning, Mn concentrations, subcellular localization and 54Mn-transport were investigated. Total Mn uptake was three-times higher in DFG than in DFV. DFV retained > 90% of 54Mn in roots, whereas > 60% was transported to the shoot in DFG. The epidermis was probably the most efficient Mn barrier since DFV contained lower Mn concentrations in cortical cells and vacuoles of roots than DFG. In both varieties, xylem loading was restricted and phloem transport was low. However, sieve cells still contained high Mn concentrations. DFV displayed higher biomass production and higher shoot : root ratios than DFG. Our results clearly show that both varieties of Douglas fir differ significantly in Mn-uptake and allocation patterns rendering DFG more vulnerable to Mn toxicity.     

Multiple-host fungi are the most frequent and abundant ectomycorrhizal types in a mixed stand of Douglas fir (Pseudotsuga menziesii) and bishop pine (Pinus muricata)

The ectomycorrhizal fungal associations of Douglas fir ( Pseudotsuga menziesii D. Don) and bishop pine ( Pinus muricata D. Don) were investigated in a mixed forest stand. We identified fungi directly from field-collected ectomycorrhizal (ECM) root tips using PCR-based methods. Sixteen species of fungi were found, of which twelve associated with both hosts. Rhizopogon parksii Smith was specific to Douglas fir. Three other species colonized only one of the hosts, but were too infrequent to draw conclusions about specificity. Seventy-four percent of the biomass of ECM root tips sampled in the stand were colonized by members of the Thelephoraceae and Russulaceae. All 12 species of fungi that associated with both tree species did so within a 10×40 cm soil volume, suggesting that individual fungal genotypes linked the putatively competing tree hosts.     

Hydraulic control of stomatal conductance in Douglas fir [Pseudotsuga menziesii (Mirb.) Franco] and alder [Alnus rubra (Bong)] seedlings

Experiments were conducted on 1-year-old Douglas fir [Pseudotsuga menziesii (Mirb.) Franco] and 2- to 3-month-old alder [Alnus rubra (Bong)] seedlings growing in drying soils to determine the relative influence of root and leaf water status on stomatal conductance (g_c). The water status of shoots was manipulated independently of that of the roots using a pressure chamber that enclosed the root system. Pressurizing the chamber increases the turgor of cells in the shoot but not in the roots. Seedling shoots were enclosed in a whole-plant cuvette and transpiration and net photosynthesis rates measured continuously. In both species, stomatal closure in response to soil drying was progressively reversed with increasing pressurization. Responses occurred within minutes of pressurization and measurements almost immediately returned to pre-pressurization levels when the pressure was released. Even in wet soils there was a significant increase in g_c with pressurization. In Douglas fir, the stomatal response to pressurization was the same for seedlings grown in dry soils for up to 120 d as for those subjected to drought stress over 40 to 60 d. The stomatal conductance of both Douglas fir and alder seedlings was less sensitive to root chamber pressure at higher vapour pressure deficits (D), and stomatal closure in response to increasing D from 1.04 to 2.06 kPa was only partially reversed by pressurization. Our results are in contrast to those of other studies on herbaceous species, even though we followed the same experimental approach. They suggest that it is not always appropriate to invoke a ‘feedforward’ model of short-term stomatal response to soil drying, whereby chemical messengers from the roots bring about stomatal closure.     

Stomatal and non-stomatal limitation to photosynthesis in two trembling aspen (Populus tremuloides Michx.) clones exposed to elevated CO2 and/or O3

Leaf gas exchange parameters and the content of ribulose‐1,5‐bisphosphate carboxylase/oxygenase (Rubisco) in the leaves of two 2‐year‐old aspen (Populus tremuloides Michx.) clones (no. 216, ozone tolerant and no. 259, ozone sensitive) were determined to estimate the relative stomatal and mesophyll limitations to photosynthesis and to determine how these limitations were altered by exposure to elevated CO₂ and/or O₃. The plants were exposed either to ambient air (control), elevated CO₂ (560 p.p.m.) elevated O₃ (55 p.p.b.) or a mixture of elevated CO₂ and O₃ in a free air CO₂ enrichment (FACE) facility located near Rhinelander, Wisconsin, USA. Light‐saturated photosynthesis and stomatal conductance were measured in all leaves of the current terminal and of two lateral branches (one from the upper and one from the lower canopy) to detect possible age‐related variation in relative stomatal limitation (leaf age is described as a function of leaf plastochron index). Photosynthesis was increased by elevated CO₂ and decreased by O₃ at both control and elevated CO₂. The relative stomatal limitation to photosynthesis (l_s) was in both clones about 10% under control and elevated O₃. Exposure to elevated CO₂ + O₃ in both clones and to elevated CO₂ in clone 259, decreased l_s even further – to about 5%. The corresponding changes in Rubisco content and the stability of C_i/C_a ratio suggest that the changes in photosynthesis in response to elevated CO₂ and O₃ were primarily triggered by altered mesophyll processes in the two aspen clones of contrasting O₃ tolerance. The changes in stomatal conductance seem to be a secondary response, maintaining stable C_i under the given treatment, that indicates close coupling between stomatal and mesophyll processes.     

Kinetics of NH4+ and K+ uptake by ectomycorrhizal fungi. Effect of NH4+ on K+ uptake

NH₄⁺ and K⁺ uptake experiments have been conducted with 3 ectomycorrhizal fungi, originating from Douglas fir (Pseudotsuga menziesii (Mirb.] Franco) stands. At concentrations up to 250 μM, uptake of both NH₄⁺ and K⁺ follow Michaelis-Menten kinetics. Laccaria bicolor (Maire) P. D. Orton, Lactarius rufus (Scop.) Fr. and Lactarius hepaticus Plowr. ap. Boud. exhibit K_m values for NH₄⁺ uptake of 6, 35, and 55 μM, respectively, and K_m values for K⁺ uptake of 24, 18, and 96 μM, respectively. Addition of 100 μM NH₄⁺ raises the K_m of K⁺ uptake by L. bicolor to 35 μM, while the V_max remains unchanged. It is argued that the increase of K_m is possibly caused by depolarization of the plasma membrane. It is not due to a competitive inhibition of K⁺ by NH₄⁺ since the apparent inhibitor constant is much higher than the K_m, for NH₄⁺ uptake. The possibility that NH₄⁺ and K⁺ are taken up by the same carrier can be excluded. The K_m, values for K⁺ uptake in the two other fungi are not significantly affected by 100 μM NH₄⁺. Except for a direct effect of NH₄⁺ on influx of K⁺ into the cells, there may also be an indirect effect after prolonged incubation of the cells in the presence of 100 μM NH₄⁺.     

Parameterization of the CO2 and H2O gas exchange of several temperate deciduous broad-leaved trees at the leaf scale considering seasonal changes

A combined model to simulate CO₂ and H₂O gas exchange at the leaf scale was parameterized using data obtained from in situ leaf‐scale observations of diurnal and seasonal changes in the CO₂ and H₂O gas exchange of four temperate deciduous broad‐leaved trees using a porometric method. The model consists of a Ball et al. type stomatal conductance submodel [Ball, Woodrow & Berry, pp. 221–224 in Progress in Photosynthesis Research (ed. I. Biggins), Martinus‐Nijhoff Publishers, Dordrecht, The Netherlands, 1987] and a Farquhar et al. type biochemical submodel of photosynthesis (Farquhar, von Caemmerer & Berry, Planta 149, 78–90, 1980). In these submodels, several parameters were optimized for each tree species as representative of the quantitative characteristics related to gas exchange. The results show that the seasonal physiological changes of V_cmax25 in the biochemical model of photosynthesis should be used to estimate the long‐term CO₂ gas exchange. For R_d25 in the biochemical model of photosynthesis and m in the Ball et al. type stomatal conductance model, the difference should be counted during the leaf expansion period.     

Effects of elevated CO2 and/or O3 on growth, development and physiology of wheat (Triticum aestivum L.)

Two cultivars of spring wheat (Triticum aestivum L. cvs. Alexandria and Hanno) and three cultivars of winter wheat (cvs. Riband, Mercia and Haven) were grown at two concentrations of CO₂ [ambient (355 pmol mol^?1) and elevated (708 μmol mol^?1)] under two O₃ regimes [clean air (< 5 nmol mol^?1 O₃) and polluted air (15 nmol mol^?1 O₃ at night rising to a midday maximum of 75 nmol mol^?1)] in a phytotron at the University of Newcastle-upon-Tyne. Between the two-leaf stage and anthesis, measurements of leaf gas-exchange, non-structural carbohydrate content, visible O₃ damage, growth, dry matter partitioning, yield components and root development were made in order to examine responses to elevated CO₂ and/or O₃. Growth at elevated CO₂ resulted in a sustained increase in the rate of CO₂ assimilation, but after roughly 6 weeks' exposure there was evidence of a slight decline in the photosynthetic rate (c.-15%) measured under growth conditions which was most pronounced in the winter cultivars. Enhanced rates of CO₂ assimilation were accompanied by a decrease in stomatal conductance which improved the instantaneous water use efficiency of individual leaves. CO₂ enrichment stimulated shoot and root growth to an equivalent extent, and increased tillering and yield components, however, non-structural carbohydrates still accumulated in source leaves. In contrast, long-term exposure to O₃ resulted in a decreased CO₂ assimilation rate (c. -13%), partial stomatal closure, and the accumulation of fructan and starch in leaves in the light. These effects were manifested in decreased rates of shoot and root growth, with root growth more severely affected than shoot growth. In the combined treatment growth of O₃-treated plants was enhanced by elevated CO₂, but there was little evidence that CO₂ enrichment afforded additional protection against O₃ damage. The reduction in growth induced by O₃ at elevated CO₂ was similar to that induced by O₃ at ambient CO₂ despite additive effects of the individual gases on stomatal conductance that would be expected to reduce the O₃ flux by 20%, and also CO₂-induced increases in the provision of substrates for detoxification and repair processes. These observations suggest that CO₂ enrichment may render plants more susceptible to O₃ damage at the cellular level. Possible mechanisms are discussed.