Effects of drought and waterlogging on ultrastructure of Scots pine and Norway spruce needles

Effects of water stress on needle ultrastructure of 2-year-old Scots pine (Pinus sylvestris L.) and 5-year-old Norway spruce [Picea abies (L.) Karst.] seedlings were studied in greenhouse experiments. Drought stress was induced by leaving seedlings without watering, and waterlogging stress was produced by submerging the seedling containers in water. Needle samples for ultrastructural analyses were collected several times during the experiments, and samples for nutrient analyses at the end of the experiments. In drought stress, plasmolysis of mesophyll and transfusion parenchyma tissues, aggregation of chloroplast stroma and its separation from thylakoids and decreased size and abundance of starch grains in needles of both species were observed. The concentration of lipid bodies around the chloroplasts were detected in pine needles. Calcium and water concentrations in spruce needles were lower by the end of the experiments compared to controls. In waterlogging treatment, swelling of phloem cells in pine needles and large starch grains, slight swelling of thylakoids and increased translucency of plastoglobuli in chloroplasts of both species studied were observed. The phosphorus concentration in pine needles was higher while phosphorus, calcium and magnesium concentrations in spruce needles were lower in the waterlogging treatments compared to controls. Typical symptoms induced by drought stress, e. g. aggregation of chloroplast stroma and its separation from thylakoids, were detected, but, in waterlogging stress, ultrastructural symptoms appeared to be related to the developing nutrient imbalance of needles.     

Endophyte communities vary in the needles of Norway spruce clones

Endophytic fungi show no symptoms of their presence but can influence the performance and vitality of host trees. The potential use of endophytes to indicate vitality has been previously realized, but a standard protocol has yet to be developed due to an incomplete understanding of the factors that regulate endophyte communities. Using a culture-free molecular approach, we examined the extent to which host genotype influences the abundance, species richness, and community composition of endophytic fungi in Norway spruce needles. Briefly, total DNA was extracted from the surface-sterilized needles of 30 clones grown in a nursery field and the copy number of the fungal internal transcribed spacer (ITS) region of ribosomal DNA was estimated by quantitative PCR. Fungal species richness and community composition were determined by denaturing gradient gel electrophoresis and DNA sequencing. We found that community structure and ITS copy number varied among spruce clones, whereas species richness did not. Host traits interacting with endophyte communities included needle surface area and the location of cuttings in the experimental area. Although Lophodermium piceae is considered the dominant needle endophyte of Norway spruce, we detected this species in only 33 % of samples. The most frequently observed fungus (66 %) was the potentially pathogenic Phoma herbarum. Interestingly, ITS copy number of endophytic fungi correlated negatively with the richness of ectomycorrhizal fungi and thus potential interactions between fungal communities and their influence on the host tree are discussed. Our results suggest that in addition to environmental factors, endophyte communities of spruce needles are determined by host tree identity and needle surface area.     

Needle chemistry in young Norway spruce stands after application of crushed wood ash

Nutrient concentrations in current and 1-year-old needles were analyzed annually for 5 years after application of hardened wood ash in 1–4-year-old Norway spruce (Picea abies (L.) Karst.) stands within a range of climate and fertility gradients. At each site, 3000 kg ha^–1 hardened wood ash of two types, Nymölla and Perstorp, was applied in a randomized block design. Wood ash Nymölla contained 12 kg ha^–1 P, 30 kg ha^–1 K, 891 kg ha^–1 Ca, 72 kg ha^–1 Mg and wood ash Perstorp contained 12 kg ha^–1 P, 60 kg ha^–1 K, 486 kg ha^–1 Ca, and 60 kg ha^–1 Mg. The ash was intended to compensate for nutrients removed at the preceding harvest when logging residues were collected and removed from the site (whole-tree harvesting). The climate gradient included four climate zones throughout Sweden and each of these included a fertility gradient of three sites classified according to their ground vegetation type. There were no effects on nutrient concentrations in the needles 1 year after the application of wood ash. Five years after ash application, the concentrations of P, K and Ca in current and 1-year-old needles were higher than in the control plots. The results were consistent over all stands, irrespective of climate zone and fertility status. P and K concentrations were higher in spruce needles from plots treated with Perstorp wood ash, whereas Ca concentrations were higher in those of Nymölla treated plots. Analyses across all study sites revealed a treatment effect in terms of increased ratios of P:N, K:N and Ca:N in 1-year-old needles. The ratio P:N tended to increase with time in the Perstorp wood ash treatment compared with the control. The needle concentrations of Mg and S were not affected by the ash applications. The increase in needle nutrient concentrations after application of hardened wood ash suggests that wood ash recycling could be used in order to replace nutrients removed at whole-tree harvesting.     

Differences in growth characteristics and dynamics of elements absorbed in seedlings of three spruce species raised on serpentine soil in northern Japan

BACKGROUND AND AIMS: Serpentine soils are characterized by the presence of heavy metals (Ni and Cr) and excess Mg; these elements often suppress plant growth. Picea glehnii is nevertheless distributed widely on serpentine soils in northern Japan. Growth characteristics were compared among P. glehnii, Picea jezoensis (distributed in the same region) and Picea abies (planted for timber production), and concentrations of elements in various tissues over time and the amount of ectomycorrhizal infection in short roots were evaluated. METHODS: Seedlings of three spruce species were planted in two types of experimental plots, comprising serpentine soil and brown forest (non-serpentine) soil, and these seedlings were grown for 3 years. Growth, ectomycorrhizal infection of short roots, and elemental composition of tissues were examined. KEY RESULTS: The total dry mass of P. glehnii planted on serpentine soil was almost the same as on brown forest soil, and a large number of needles survived to reach later age classes. By contrast, growth of P. jezoensis and P. abies in serpentine soil was significantly less than in brown forest soil, and needle shedding was accelerated. Moreover, roots of seedlings of P. glehnii on serpentine soil were highly infected with ectomycorrhiza, and the concentration of Ni in needles and roots of P. glehnii was the lowest of the three species. CONCLUSIONS: Picea glehnii has a high ability to maintain a low concentration of Ni, and the ectomycorrhizal infection may have the positive effect of excluding Ni. As a result, P. glehnii is more tolerant than the other spruce species to serpentine soil conditions.     

Effects of Cd and Zn on the development of annual xylem rings of young Norway spruce (Picea abies) plants

Three-year-old spruce (Picea abies) saplings were planted and cultivated for 2 years in pots with 3 1 substrate, consisting of a homogenized mixture of sand, peat and forest soil with a high organic content (volume ratio 11.52). This substrate was amended with 10–180 mol Cd [kg soil dry weight (DW)]^–1, 50–7500 mol Zn (kg soil DW)^–1 (determined with 1 M ammonium acetate extracts) or combinations of both elements. Annual xylem growth rings in stems of plants treated with 50 mol Cd (kg soil DW)^–1 or 7500 mol Zn (kg soil DW)^–1 were significantly narrower than in control plants. Growth reductions were more pronounced in the second year of the experiment. The contents of Cd and Zn in stem wood and needles were positively correlated with the substrate concentrations. The Mg contents of the spruce needles were inversely correlated with soil concentrations of Cd and Zn. Root development was impeded at moderate concentrations of Cd (50 mol kg^–1) or Zn (1000 mol kg^–1) in the substrate. The adverse effects of potentially toxic trace elements, like Cd or Zn, on xylem growth of spruce plants are discussed with regard to possible growth reductions in forest trees under field conditions.     

Effect of nitrogen on drought strain and nutrient uptake in Norway spruce Picea abies (L.) Karst.) trees

A field lysimeter study was established with the aim of investigating the effect of nitrogen availability upon drought strain in Norway spruce trees. Forest soil (Typic Udipsamment) was filled in lysimeters 1 m in diameter and 1 m deep. Small trees of Norway spruce from five different clones were planted in the lysimeters. Roofs under the canopy of the trees ensured full control of water and nutrient input. Three levels of nitrogen were given to the trees during five years; ambient rainwater, and five and fifteen times this N concentration, respectively. Additional N was given as NH₄NO₃ in irrigation water. Mean annual N-addition during the five years corresponded to 5, 27 and 82 kg per ha and year for the three treatments, respectively. During the third and fifth growth season drought was artificially induced. In addition to a watered control, two levels of drought were applied, representing water deprivation for 2 and 3 months, respectively, in 1990 and 3 and 4 months, respectively in 1992. A higher water consumption in the nitrogen fertilized trees during the droughts resulted in a significantly lower pre-dawn shoot water potential compared to the trees receiving ambient rain N. The interaction between drought and nitrogen fertilization was clear also for photosynthesis and transpiration. A decrease in height- and diameter increment caused by drought was most pronounced in the 82 kg N ha^–1 yr^–1 treatment. A water strain integral showed a strong positive correlation to the needle biomass of the trees. Foliar concentrations of several nutrients decreased significantly with increasing drought strain in the trees. Concentration of potassium and boron were especially low and visual symptoms of deficiency occurred.     

The effect of drought on mycorrhizal production and very fine root system development of Norway spruce under natural and experimental conditions

Mycorrhizal growth rates were measured monthly, using a new method, in two neighouring plots of a natural spruce stand. One of the plots was irrigated while the other suffered from drought during the late summer and autumn months. Drought did not completely stop mycorrhizal growth. It caused a higher rate of root dormancy and a reduced elongation rate of the parent roots but an increased development of new mucorrhizal last order laterals. Thus, the branching density of the very fine root system was increased, even though fewer growing mycorrhizae were found in the non-irrigated plot during the dry period. Similar results were observed in a water-stress experiment with pot-cultures. After rewetting, elongation rate was stimulated and the number of growing mycorrhizae increased rapidly on the non-irrigated plot. Possible relationships between dry weight, distribution and branching density of growing fine root systems are presented.     

Cell structural changes in the needles of Norway spruce exposed to long-term ozone and drought

Effects of ozone and/or drought on Norway spruce needles werestudied using light microscopy and electron microscopy. Saplingswere exposed to ozone in open-top chambers during 1992–1995and also to drought in the late summers of 1993–1995.Samples from current and previous year needles were collectedfive times during 1995. Ozone increased the numbers of peroxisomesand mitochondria, which suggests that defence mechanisms againstoxidative stress were active. The results from peroxisomes suggestthat the oxidative stress was more pronounced in the upper sideof the needles, and those from mitochondria that defence wasmore active in the younger needle generation. Possibly due tothe good nitrogen status and the active defence, no ozone-specificchloroplast alterations were seen. At the end of the season,older needles from ozone treatments had smaller central vacuolescompared with other needles. Cytoplasmic vacuoles around thenucleus were increased by ozone in the beginning of the experiment,and did not increase towards the end of the season as in thecontrols. These results from vacuoles may indicate that ozoneaffected the osmotic properties of the cells. Decreased numberand underdevelopment of sclerenchyma cells and proliferationof tonoplast were related to nutrient imbalance, which was enhancedby drought. Larger vascular cylinders and more effective starchaccumulation before and after the drought periods compensatedfor the reduced water status. Numbers of peroxisomes and mitochondriawere increased in the drought-exposed needles before the onsetof drought treatments of the study year, i.e. these changeswere memory effects. Interactions between ozone and droughtwere few.     

Genotype dependent blue and red light inhibition of the proliferation of the embryogenic tissue of Norway spruce

Summary The influence of light quality on the proliferation of embryogenic tissue of three genotypes of Norway spruce (Picea abies [L.] Karst), with different capacities for mature somatic embryo production, was studied. The proliferating tissues were subjected to light from commercially available light sources: Philips TLD Warm White 36W/29, Philips TLD Blue 18W/18, Philips TLD Red 36W/15, Osram L Fluora 36W/77 and Sylvania Far Red 7080, for 18 h a day with the photon flux (PAR) at 30 μmol m⁻²s⁻¹. The effect of light quality on the growth of embryogenic tissue was strongly genotype dependent. In genotype 164-4 tissue proliferation was strongly inhibited by blue and red light. Genotype 86∶52 reacted in a similar way, but not as strongly as 164-4, whereas the tissue of genotype 186-3 was almost insensitive to light quality and grew fast in all light conditions.     

Effects of ozone on starch accumulation in Norway spruce (Picea abies)

Summary Fumigation with 100 g/m³ and 200 g/m³ ozone in closed-top fumigation chambers induced starch accumulation in chloroplasts of Norway spruce. This accumulation was probably due to a partial inhibition of the starch translocation at night. The intensity of the effect was dependent on the season and the age of the needles. The accumulation was reversed in winter. It is therefore unlikely that such an effect has much significance for plant health.     

Concentrations of nutritional and trace elements in needles of Norway spruce (Picea abies [L.] Karst.) as functions of the needle age class

The endogenous concentrations of the essential elements Ca, Cl, Cu, Fe, K, Mg, Mn, N, P and Zn, and of the nonessential elements Al, As, Ba, Br, Co, Cr, Cs, Hg, I, La, Na, Rb, Sb, Sc, Si and Sr were determined in 5 successive needle age classes. 40 mature spruce trees from 6 different sites were investigated individually. A given element usually shows smooth changes with the needle age class t. Trees on a given site usually have a similar dynamic behaviour. The same holds for the different site means. The concentrations can be approximated by functions c=f(t). Three different types of functions are required to describe the dynamic behaviour of 3 groups of elements that increase with t, and one for the elements that decrease with t. A given element usually can be described by the same type of function at all sites, even if its concentration differs widely. Exceptions are Mn, Co and Zn, which change from a decreasing function at low concentrations to an increasing function at high concentrations. Further irregulatities are found at some sites with Ca, Sr and Ba. These findings are corroborated by a multivariate statistical analysis.     

Major and trace elements in the twig axes of Norway spruce and the relationship between twig axis and needles

Summary The concentrations of 23 elements have been determined by instrumental neutron activation analysis in the needles and axes of 1-year-old spruce twigs from 12 different sites. Although the twigs had been washed with toluene and tetrahydrofuran prior to analysis, it was found that the amounts of eight elements (Al, Cr, Eu, Fe, La, Sc, Th and V) present were predominantly due to aerosol particles still being retained on the surface of the axes. The results of the remaining 15 elements (Ba, Br, Ca, Cl, Co, Cs, Hg, K, Mg, Mn, Na, P, Rb, Sr and Zn) were not or only slightly influenced by this effect and are considered to represent their inherent concentrations within the axes. With most of these latter 15 elements there are highly significant and linear correlations between the concentrations in the needles and in the axis, which testify to the great similarity of needles and axis of a twig. The axis/needles ratios are constant for every element (except K), but the individual elements show considerable differentiation, as evidenced by their ratios, which have values between 0.5 and 2.8. Inter-element correlations in the axes are mainly found within two groups of elements. With Na, Cl and Br these correlations are probably due to different pollution levels of the different sites, whereas with Mn, Co, Rb and Cs they are due to the pH of the soil. The activity of ¹³⁷Cs from the Chernobyl fallout represents independent confirmation of the results obtained with the stable elements. With twigs sprouted before the deposition of the fallout, ¹³⁷Cs acts as a tracer for aerosol particles, whereas it forms a tracer for the stable Cs inherent to the plant with twigs sprouted after the deposition.     

Water relations in Norway spruce trees growing at ambient and elevated CO2 concentrations

Water relations were studied in Norway spruce [Picea abies (L.) Karst.] trees grown at ambient (AC, 350 μmol mol⁻¹) and elevated (EC, 700 μmol mol⁻¹) CO₂ concentrations under temperate water stress. The results suggested that both crown position and variability in atmospheric CO₂ concentration are responsible for different patterns of crown water relations. Mean hourly sap flux density (FSA) showed higher values in upper crown position in comparison with the whole crown in both AC and EC treatments. Mean soil-to-leaf hydraulic conductance (G_Tsa) was 1.4 times higher for the upper crown than that calculated across the whole crown for the trees in AC. However, G_Tsa did not vary significantly with crown position in EC trees, suggesting that elevated CO₂ may mitigate differences in hydraulic supply for different canopy layers. The trees in EC treatment exhibited significantly higher values of F_SA measured on the whole crown level and slightly higher soil water content compared to AC treatment, suggesting more economical use of soil water and therefore an advantage under water-limited conditions.     

Effect of drought stress on chlorophyll a fluorescence and electrical admittance of shoots in Norway spruce seedlings

Effects of mild and severe soil drought on the water status of needles, chlorophyll a fluorescence, shoot electrical admittance, and concentrations of photosynthetic pigments in needles of seedlings of Picea abies (L.) Karst. were examined under controlled greenhouse conditions. Drought stress reduced shoot admittance linearly with a decrease in shoot water potential (_w) and increase in water deficit (WD) and led to a decrease in concentrations of chlorophyll a, b and carotenoids. Severe water stress (shoot _w=–2.4 MPa) had a negative effect on chlorophyll a fluorescence parameters including PSII activity (F_v/F_m), and the vitality index (R_fd). Variations in these parameters suggest an inhibition of the photosynthetic electron transport in spruce needles. Water stress led to a decrease in the mobility of electrolytes in tissues, which was reflected by decreased shoot electrical admittance. After re-watering for 21 days the WD in needles decreased and the shoot water potential increased. In the re-watered plants, the chloroplast function was restored and chlorophyll a fluorescence returned to a similar level as in the control plants. This improved hydraulic adjustment in the seedlings triggered a positive effect on ion flow in the tissues and increased shoot electrical admittance. We conclude that the shoot electrical admittance and photosynthetic electron transport in leaves are closely linked to changes in water status and their decrease is among the initial responses of seedlings to water stress.     

Experimental manipulation of water and nutrient input to a Norway spruce plantation at Klosterhede,Denmark

Water and nutrient supply to forest trees are major factors controlling tree growth and forest vitality. Therefore, changes in the supply of water and nutrients to the trees may be important contributing reasons to the forest damages observed in Europe. Such changes may be caused by several factors, e.g. air pollution, soil acidification and climate change. The present study investigates possible effects on tree growth, growth related parameters and nutrition related to changes in the water and nutrient supply. Water and nutrient supply to the forest soil was manipulated in three roof covered plots. The treatments consisted of 1) summer drought, 2) irrigation and 3) combined irrigation and fertilizer application (fertigation). The results from the roof covered treatment plots were compared to a control plot without roof. Increased supply of water during the spring and early summer increased the diameter growth, whereas application of nutrients in addition to irrigation had no additional effect on tree growth. Addition of nutrients increased the needle content of P, Mn, Ca and Mg. Extended summer drought for 2 months had no effect on the tree growth or other growth parameters, but drought reduced the root development in the upper soil layer. Furthermore, drought induced significant stress symptoms by increasing the cone shredding substantially.     

Impact of warming and drought on carbon balance related to wood formation in black spruce

Background and Aims

Wood formation in trees represents a carbon sink that can be modified in the case of stress. The way carbon metabolism constrains growth during stress periods (high temperature and water deficit) is now under debate. In this study, the amounts of non-structural carbohydrates (NSCs) for xylogenesis in black spruce, Picea mariana, saplings were assessed under high temperature and drought in order to determine the role of sugar mobilization for osmotic purposes and its consequences for secondary growth.

Methods

Four-year-old saplings of black spruce in a greenhouse were subjected to different thermal conditions with respect to the outside air temperature (T0) in 2010 (2 and 5 °C higher than T0) and 2011 (6 °C warmer than T0 during the day or night) with a dry period of about 1 month in June of each year. Wood formation together with starch, NSCs and leaf parameters (water potential and photosynthesis) were monitored from May to September.

Key Results

With the exception of raffinose, the amounts of soluble sugars were not modified in the cambium even if gas exchange and photosynthesis were greatly reduced during drought. Raffinose increased more than pinitol under a pre-dawn water potential of less than –1 Mpa, presumably because this compound is better suited than polyol for replacing water and capturing free radicals, and its degradation into simple sugar is easier. Warming decreased the starch storage in the xylem as well the available hexose pool in the cambium and the xylem, probably because of an increase in respiration.

Conclusions

Radial stem growth was reduced during drought due to the mobilization of NSCs for osmotic purposes and due to the lack of cell turgor. Thus plant water status during wood formation can influence the NSCs available for growth in the cambium and xylem.     

Rubidium and cesium in spruce needles

The endogenous concentrations of Rb and Cs have been determined in needles of 56 trees (Norway spruce,Picea abies) from eight different sites. Analysis was done by instrumental neutron activation. Concentrations were found to have a very large range (3-28,000 ng Cs/g and 1–190 μg Rb/g). The values on a given site have a tendency toward a log-normal distribution. There is a significant correlation between Rb and Cs, but the correlation is not linear. The concentrations of both elements are a function of the needle age. They decrease smoothly, approaching a constant value, when going from needle age class 1 to 5. It is shown that one algebraic function describes this biodynamic behavior on all sites and at all concentration levels. The function and its parameters are discussed.     

Life expectancy of spruce needles under extremely high air pollution stress: performance of trees in the Ore Mountains

Summary Photosynthetic performance of Norway spruce needles [Picea abies (L.) Karst.] was measured over a 1-year period. The trees grew in an area of heavy air pollution and forest decline on a mountain ridge in the eastern Ore Mountains (Czech Republic). Photosynthetic capacity, as well as light use efficiency, decreased dramatically with time, starting in July (2 months after bud-break) to finally reach zero (respiration only) by February of the following year. Two months later all needles from upper crown parts were shed. Needles from lower crown parts, on the other hand, were undamaged. The chlorophyll and Mg content decreased transiently during the cold season, with Mg reaching deficiency thresholds during winter. However, total sulfur, as well as organic and sulfate S increased with time. The increase was higher in needles from the upper parts of the crown, which were exposed to windy air throughout the year, than in the lower parts of the crown, which were covered by grass during summer and by snow during most of the winter.     

Nitrogen uptake in a Norway spruce stand following ammonium sulphate application,fertigation, irrigation,drought and nitrogen-free-fertilisation

The above-ground accumulation of N,N uptake and litter quality resulting from improved or deteriorated availability of water and nutrients in a 25 year old Norway spruce stand in SW Sweden (as part of the Skogaby project) is presented. Treatment include irrigation; artificial drought; ammonium sulphate addition; N-free-fertilisation and irrigation with liquid fertilisers including a complete set of nutrients according to the Ingested principle (fertigation). At start of the experiment the stand contained 86.5 t dry mass and 352 kg N ha⁻¹. The following three years the annual N uptake in untreated trees was 32 kg N ha⁻¹ to be compared with the annual N throughfall of 17 kg ha⁻¹. Simultaneously, the treatment with ammonium sulphate and liquid fertilisation resulted in 48 and 56 kg ha⁻¹ y⁻¹, respectively, in treatment specific N-uptake following an application of 100 kg N ha⁻¹ y⁻¹. Addition of a N-free fertiliser resulted in improved N-uptake by 19 kg N ha⁻¹ y⁻¹ and irrigation by 10 kg N ha⁻¹ y⁻¹, compared to control. A linear relation between total above-ground dry mass production and N-uptake was found for trees growing with similar water availability. Dry mass production increased with increased water availability given the same N-uptake. It is concluded that the studied stand this far is not N saturated', as N fertilisation resulted in both increased N uptake and increased growth. Addition of a N-free-fertiliser resulted in increased uptake of N compared to the control, indicating an increased mineralisation rate or uptake capacity of the root system. The linear relation between N uptake and biomass production shows that at this study site N is a highly limiting factor for growth.     

Nitrate reductase in needles,roots and trunk wood of spruce trees [Picea abies (L.) Karst.]

Summary Nitrate reductase (EC 1.6.6.2) activity (NRA), as measured by an in vivo assay, is present in needle leaves and mycorrhizal fine root tips of adult Norway spruce [Picea abies (L.) Karst.] in at least equal amounts on a fresh weight basis, in both adult and 5-year-old trees. NRA could also be demonstrated in trunk wood of deroted trees after fertilization with 5 mM , exhibiting a longitudinal profile in the trunk. Inducibility in needles can more efficiently be achieved by NO₂ (100 g·m^-3) than by 5 mM nitrate, which is effective only in root-amputated trees. A remarkably high level of needle-NRA in unfertilized trees, which are characterized by a very low level of nitrate in the xylem sap, suggests that NRA in spruce needles may in part be constitutive. Organic-N is a major nitrogen source for the needles even in root-amputated trees, indicating pronounced exchange processes between ray parenchyma and trunk xylem, which in turn are modified by the nitrogen source fed to the trunk stump. Intact trees exhibit a very similar amino acid composition of the xylem sap, regardless of whether or has been fed. The amino acid pattern of the needles is not thrown out of balance by flooding with and , which occurs in fertilized derooted trees. This indicates a distinct potential for homoeostasis of nitrogen entrance-metabolism (i.e. NRA and glutamine synthetase activity) in the needles. In the ectomycorrhiza/fine root-system (EMC), marked differences in NRA were observed depending on root-tip diameter and along the longitudinal profile of the fine roots. EMC-nitrate reductase is strongly enhanced by . Needle-NRA exhibits a circannual rhythm. An early summer maximum is followed by a December minimum. This activity pattern matches well the transitory increase of soluble nitrogen in spring and the total protein maximum in winter. In an indirect way assimilatory NRA may well contribute to nitrogen overfertilization (by consumption of NO_X) as one possible cause of the contemporary decline of spruce populations.