Freeze-preservation of buds from Scots pine trees

A procedure has been developed for freeze-preservation of buds of the Scots pine (Pinus sylvestris L.). Instead of liquid nitrogen, cold storage in –80°C was used. The partly dormant material used in the experiments was obtained directly from a natural stand in Northern Finland and no prefreezing or cryoprotectants for preconditioning were used. Cooling velocity was 1°C/min up to a terminal freezing temperature of –39°C, after which the buds were immersed in liquid nitrogen at –196°C for 10 minutes. The material was then transferred to a deepfreezer at –80°C and stored up to 6 months. After rapid thawing, the buds were sterilized and their viability was tested by FDA staining and by culturing meristems on 1/2 MS medium for at least two weeks. All the freezing experiments were performed during March and April. The best survival of buds (90–100%) was achieved at the beginning of April, after which a pronounced decline in survival occurred obviously due to a rise in the water content of the buds.     

Ectomycorrhizal development on Scots pine (Pinus sylvestris L.) seedlings in different soils

In the present study ectomycorrhizal development of Laccaria bicolor, Rhizopogon luteolus and Suillus bovinus associated with Scots pine (Pinus sylvestris) seedings was studied as affected by primary stand humus, secondary stand humus, podsolic sandy soil or peat in perspex growth chambers. After 9 weeks, ectomycorrhizal development with S. bovinus was significantly greater in peat and primary stand humus than in secondary stand humus or podsolic sandy soil. Ectomycorrhizal development with R. luteolus in secondary stand humus was higher than in primary stand humus. Degree of ectomycorrhizal development of L. bicolor, R. lutuelus and S. bovinus on Scots pine was related to potassium concentration, organic matter content and pH of the soils suggesting that chemical composition of the soils affects ectomycorrhizal development.     

Seasonal changes in explant viability and contamination of tissue cultures from mature Scots pine

Explants from 10 to 40-year-old Scots pine trees (Pinus sylvestris L.) were cultured in vitro. Material was collected from Northern Finland once or twice a week during 1984–1987. excised shoot meristems and lower parts of the buds formed soft callus on modified MS medium. A seasonal effect was observed in the explant viability and degree of contamination. Callus proliferation was highest from explants collected in December and January and during the growing season from April to July, and lowest in February and during the autumn from September to November. It seemed that the bud metabolism at each particular time was rather persistent and affected the outcome of the experiments. Contamination was significantly higher from December to April. Organogenesis occurred only rarely.     

Variation of specific leaf area and upscaling to leaf area index in mature Scots pine

Reliable and objective estimations of specific leaf area (SLA) and leaf area index (LAI) are essential for accurate estimates of the canopy carbon gain of trees. The variation in SLA with needle age and position in the crown was investigated for a 73-year-old Scots pine (Pinus sylvestris L.) stand in the Belgian Campine region. Allometric equations describing the projected needle area of the entire crown were developed, and used to estimate stand needle area. SLA (cm² g⁻¹) as significantly influenced by the position in the crown and by needle age (current-year versus 1-year-old needles). SLA increased significantly from the top to the bottom of the crown, and was significantly higher near the interior of the crown as compared to the crown edge. SLA of current-year needles was significantly higher than that of 1-year-old needles. Allometric relationships of projected needle area with different tree characteristics showed that stem diameter at breast height (DBH), tree height and crown depth were reliable predictors of projected needle area at the tree level. The allometric relationships between DBH and projected needle area at the tree level were used to predict stand-level needle area and estimate LAI. The LAI was 1.06 (m² m⁻²) for current-year needles and 0.47 for 1-year-old needles, yielding a total stand LAI of 1.53.     

Fatty acids,starch and biomass of Scots pine needles and roots in open-air ozone exposure

Scots pine (Pinus sylvestris L.) seedlings were fumigated with 1.2–1.5 x ambient ozone (cumulative exposure) over 2 seasons in an open-air experiment. Starch and fatty acid concentrations were analyzed in needle and root tissue in the summer, autumn and early winter. Seedling growth was determined by measuring the height of the stem and the total shoot and root biomass. Significant decreases in growth were found in exposed seedlings, even though visible symptoms were lacking. Almost significant reductions in needle and root starch concentrations were found. In the ozone treated foliage, significant increases in myristic acid (140) were detected, but the major fatty acids remained unchanged. Fatty acid ratios showed that the degree of unsaturation decreased in treated needles in the summer. In the roots of ozone treated seedlings, changes in fatty acids were different from those in the foliage. Decreases of the main root fatty acids (160, 180, 181, 18:2, 183) were detected in the summer. These results show that Scots pine is susceptible to enhanced levels of ozone. If the tropospheric ozone levels continue to increase it may have deleterious effects on Scots pine forests in Finland.     

Evaporative demand determines branchiness of Scots pine

Analysis of the branch area/stem area ratio of Scots pine growing in different climatic conditions in Europe and Siberia indicates that the branch area supported by a stem increases in warmer and drier conditions. The ratio was significantly correlated with several climatic variables, especially with potential evapotranspiration (E _p). The ratio was negatively correlated with stand density (d _s). A regression model combining E _p and d _s accounted for 85% of the total variation. These trends are believed to reflect hydraulic segmentation of trees and may represent a strategy to avoid cavitation in the tree, especially in the branches.     

Photoreactivation of the UV light effects on growth of Scots pine (Pinus sylvestris L.) seedlings

Summary Ultraviolet-B light (UV-B) and ultraviolet-A light (UV-A) at higher doses exert a strong inhibitory (toxic) effect on axis growth in Scots pine (Pinus sylvestris L.) seedlings. This effect is unrelated to control of growth rate by phytochrome. Rather, after a toxic UV dose growth of the pine seedling no longer responded to phytochrome. Both, the effect of UV-B as well as the inhibiting effect of UV-A could be photoreactivated by blue light (B). The action of UV-A was 2 fold: (i) it exerted a toxic effect which could be photoreactivated by B, and (ii) applied after UV-B it photoreactivated to some extent the toxic UV-B effect. Obviously, the UV-A range causes a toxic effect, and at the same time is capable of photoreactivating the toxic UV effect. At higher doses the toxic effect prevails.     

Studies of crystallinity of Scots pine and Norway spruce cellulose

The variation in the mass fraction of crystalline cellulose (crystallinity of wood), the intrinsic crystallinity of cellulose, and the thickness of cellulose crystallites in early wood of Norway spruce [Picea abies (L.) Karst.], and Scots pine (Pinus sylvestris L.) grown in Finland were studied using wide angle X-ray scattering and nuclear magnetic resonance spectroscopy. The mass fraction of crystalline cellulose in wood increased slightly with the distance from the pith and was about 30±4% in mature wood of both species. The crystallinity of cellulose and the thickness of cellulose crystallites were almost constant for both species. The crystallinity of cellulose was 52±3% for both species and the average thickness of the cellulose crystallites was 32±1 Å and 31±1 Å for Norway spruce and Scots pine, respectively. The mass fraction of cellulose in wood, calculated from the crystallinity values, increased with the distance from the pith for both species.     

Aluminium causes nutrient imbalance and structural changes in the needles of Scots pine without inducing clear root injuries

The effects of aluminium chloride (AICI3) treatments (50 and 150 mg/l) on 3-year-old Scots pine (Pinus sylvestris L.) seedlings were studied in a sand culture during 2 growing periods in an open field experiment. Even by the end of the first growing period, a decline was observed in the concentrations of Ca, Mg and P within the needles, and of Ca and Mg in the roots. After the second growing period, increased N and K concentrations were observed in the needles of Al-treated seedlings. Both the needles and roots of Al-treated seedlings showed, after the second growing period, a decline in growth and increased concentrations of AI as the amount of AICI3 in the nutrient solution increased. Al-induced changes in needle structure were found to be symptomatic of a nutrient imbalance, particularly of Mg and P. Al-stress did not result in any observable changes in root anatomy or in the number of mycorrhizas. Scots pine proved to be rather resistant to Al-stress, indicating that direct Al-injuries are not likely in the field, though Al-stress may be a contributing factor in the formation of nutrient imbalances.     

Food reserves of Scots pine (Pinus sylvestris L.)

Summary The amounts of starch, soluble sugars, triacylglycerols, diacylglycerols and free fatty acids were studied in Scots pine (Pinus sylvestris L.) during an annual cycle in current-year needles and in 1-, 2- and 3-year-old needles collected shortly after bud break. Determination of the compounds was performed using specific enzymatic assays, capillary gas chromatography and thin layer chromatography. Newly emerging needles contained relatively large amounts of starch, but only trace amounts of fat. During autumn and winter, fat content rose, while starch content decreased; amounts of both these reserve materials were very high the next spring shortly before bud break and decreased again during shoot elongation. Concentration of intermediates in triacylglycerol biosynthesis (diacylglycerols and free fatty acids), were low in summer and high in winter. The same pattern was observed for fructose and glucose (the predominant soluble sugars), galactose/arabinose and raffinose/melibiose. In contrast, sucrose concentrations were highest in spring and in autumn. Mature needles of different ages collected in May showed significant differences only in their triacylglycerol and starch content. Concentration changes of reserve materials are discussed in relation to season, mobilization and translocation processes, dormancy, frost resistance and the possibility of carbohydrate-fat interconversions.     

Effect of provenance on free amino acid and chemical composition of Scots pine needles

Free amino acid (16 amino acids) and chemical composition (N, P, K, Mg, Ca, S, Fe, Mn, Cu, Zn) of Scots pine (Pinus sylvestris L.) needles were compared between six provenances in three different experimental areas. The main free amino acids in the needles were in the sequence of quantity; glutamic acid, glutamine, arginine and γ-aminobutyric acid. There were no significant differences in the concentrations of phenylalanine, γ-aminobutyric acid, methionine, proline and threonine in pine needles between the sites or between the provenances. Significant differences in the foliar concentrations of alanine and leucine were found between the sites and in the foliar concentrations of isoleucine, glutamine, glycine and tyrosine between the provenances. The concentrations of aspartic acid, glutamic acid, arginine and lysine were significantly affected by the sites and the provenances. The foliar nutrients, except copper, had statistically significant differences, both between the sites and between the provenances. Calcium did not differ between the provenances. This revised version was published online in June 2006 with corrections to the Cover Date.     

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.     

Root biomass and rooting profile of naturally regenerated beech in mid-elevation Scots pine woodlands

The natural establishment of shade-tolerant forest species such as beech (Fagus sylvatica L.) occurs in naturally regenerated Scots pine (Pinus sylvestris L.) woodlands that develop on former pastures and cultivated lands. To examine possible effects of underground competition in beech establishment, we studied the root biomass and the rooting profile of 53 mixed Scots pine-natural beech woodlands in French mid-elevation volcanic areas. Stands were arranged along a maturation gradient. Roots were sampled using the root-auger technique (0 to 75 cm depth every 15 cm). In addition, 23 young beeches were uprooted to study the entire root system. Total beech fine-root biomass was closely correlated with most beech aerial characteristics, (e.g., height, diameter and girth), and correlated moderately with tree age. However, it correlated poorly with basic competition indices such as stand density and basal area. Conversely, competition indices including vertical dimensions of competing trees were correlated with the underground biomass, probably as a result of redundancy with beech height. The rooting profile (fine roots, < 5 mm) of beech and pine were quite similar, and did not change significantly along the stand maturation gradient. Beech has a heart-shaped root system while pine is more plate-like and dimorphic. Beech fine-root biomass progressively surpassed pine biomass throughout the soil layers, thus confirming that it is dynamic and competitive in mature mixed stands. The coexistence of the root systems of beech and pine in the same soil layers presumably results in strong underground competition.     

Long-term needle litterfall of a Scots pine Pinus sylvestris stand: relation to temperature factors

Summary Needle litterfall of a Scots pine was caught over 24 years (1962–1986) with litter-traps in a Scots pine stand in southeastern Finland. The age of the trees averaged 111 years in 1962. The stand was naturally recruited and only minor silvicultural treatments occurred during its history. Litterfall showed great year-to-year variation, the minimum being 18 g/m² (in 1968) and maximum 213 g/m² (in 1973). There was no overall trend in the amount of litterfall, and the age of the stand was thus not important in determining the needle fall. We used time domain time series analysis (ARIMA) and standard climatic data (temperature, precipitation) to investigate the relationship of litterfall to climatic factors. Mean July temperature was clearly correlated with needle litterfall. High temperature in July coincided with enhanced litterfall in the same and the next year. Litterfall enhanced litterfall in the same and the next year. Litterfall increased also after high temperatures during March–April, but only in the same year. In addition to these the litterfall had a 4-year self-dependency. This is approximately the same as the mean longevity of needles in the study area. Altogether the time series model we propose covers about 90% of the variance of the original time series.     

Impact of Douglas-fir and Scots pine seedlings on plagioclase weathering under acidic conditions

The weathering of soil minerals in forest ecosystems increases nutrient availability for the trees. The rate of such weathering and its relative contribution to forest tree nutrition, is a major issue when evaluating present and potential forest stand productivity and sustainability. The current paper examines the weathering rate of plagioclase with and without Douglas-fir or Scots pine seedlings, in a laboratory experiment at pH 3–4 and 25 °C. All nutrients, with the exception of Ca, were supplied in sufficient amounts in a nutrient solution. The objective of the experiment was to evaluate the potential of trees to mobilise Ca from the mineral plagioclase that contained 12% of Ca. Amounts of nutrients supplied in the nutrient solution, amounts accumulated in the living tissue of the seedlings and amounts leached from the experimental vessels, were measured. A weathering balance, accounting for leached + accumulated − supplied amounts, was established. Bio-induced weathering, defined as the weathering increase in the presence of trees, relative to the weathering rate without trees (geochemical weathering; control vessels), under the present experimental conditions, explained on average, 40% of total weathering (biological + geochemical). These conditions appeared more beneficial to Scots pine (higher relative growth rate, higher Ca incorporation) than to Douglas-fir.     

Characterization of cDNAs encoding CuZn-superoxide dismutases in Scots pine

A Scots pine (Pinus sylvestris L.) cDNA library was screened with two heterologous cDNA probes (P31 and T10) encoding cytosolic and chloroplastic superoxide dismutases (SOD) from tomato. Several positive clones for cytosolic and chloroplastic superoxide dismutases were isolated, subcloned, mapped and sequenced. One of the cDNA clones (PS3) had a full-length open reading frame of 465 bp corresponding to 154 amino acid residues and showed approximately 85% homology with the amino acid sequences of angiosperm cytosolic SOD counterparts. Another cDNA clone (PST13) was incomplete, but encoded a putative protein with 93% homology to pea and tomato chloroplastic superoxide dismutase. The derived amino acid sequence from both cDNA clones matched the corresponding N-terminal amino acid sequence of the purified mature SOD isozymes. Northern blot hybridizations showed that, cytosolic and chloroplastic CuZn-SOD are expressed at different levels in Scots pine organs. Sequence data and Southern blot hybridization confirm that CuZn-SODs in Scots pine belong to a multigene family. The results are discussed in relation to earlier observations of CuZn-SODs in plants.     

Rooting strategy of naturally regenerated beech in Silver birch and Scots pine woodlands

This study investigated the belowground development and strategy of late-successional European beech (Fagus sylvatica L.) in ageing natural Scots pine (Pinus sylvestris L.) and Silver birch (Betula pendula Roth.) woodlands in a French volcanic mid-elevation area. For this purpose root biomass, root profile and fine-root architecture of competitor trees were examined in 53 mixed pine–beech and 42 birch–beech woodlands along a stand maturation gradient, using the root auger technique (0–75-cm). The total beech fine-root biomass highly correlated with aerial dimensions such as stem height and girth, whereas it moderately correlated with its age, thus indicating the effects of competition. Basic stand biometric data such as stand density and basal area had no significant effect on beech root biomass. Conversely, competition indices taking into account the vertical dimensions of competitor trees were efficient, probably due to redundancy with beech height. At similar age and height, beeches under birch had a greater belowground development than beeches under pine. Each species exhibited specific rooting pattern and plasticity of fine-root architecture along the gradients of stand maturation and competition. Beech had a heart-shaped rooting habit in both mixings, which strongly increased along stand maturation. Its fine-root system adopted a foraging strategy to respond to increasing stand competition. The Scots pine fine-root system was plate-like and showed a low morphological plasticity, thus presumably a conservative strategy. Silver birch exhibited a high biomass and a foraging capacity in the topsoil but a loose root system in the subsoil. The coexistence of pine and beech roots in the upper soil presumably leads to a high belowground competition. Beech root system becomes predominant throughout the soil profile and it adopts an efficient foraging strategy, but at the expense of its belowground development. Conversely, the niche partitioning strategy between beech and birch may explain why beech develops strongly belowground in spite of the fact that birch has a dense rooting and a competitive fine-root architecture. As a consequence, beech mid-term regeneration and development may be facilitated under birch as compared with pine.     

Spermidine and the ectomycorrhizal fungus Pisolithus tinctorius synergistically induce maturation of Scots pine embryogenic cultures

Exogenous spermidine (Spd) and the ectomycorrhizal (ECM) fungus Pisolithus tinctorius (Pers.) Coker and Couch had a synergistic effect on the maturation of Scots pine (Pinus sylvestris L.) somatic embryos. Induced maturation was expressed as a higher number of cell masses able to form embryos and a greater number of embryos formed per cell mass. In contrast, treatment with P. tinctorius alone on the hormone-free medium resulted in the lowest embryo-forming capacity. Retarded proliferation growth appeared to be required for maturation, but did not explain the synergistic effect of the fungus and exogenous Spd. Simultaneous treatment did not result in lower concentrations of putrescine (Put), Spd or spermine (Spm) in the embryogenic cell masses relative to the separate treatments. Our study is the first report on the use of a specific ECM fungus to induce maturation of somatic embryos, and it indicates that P. tinctorius was able to modify the maturation media in a way that, together with exogenous Spd, positively affected embryogenic cultures of Scots pine. Our study also shows that it is possible to enhance plant development other than root formation by using specific ECM fungi.     

Diurnal course of photochemical activity of winter-adapted Scots pine at subzero temperatures

The photochemical activity of native Central Siberian Scots pine trees (Pinus sylvestris L) was estimated from the middle of February to the middle of March 2001. We measured chlorophyll (Chl) fluorescence in attached intact needles from trees located approx. 30 km west of the Yenisey river (60°44′N, 89°09′E) near the village of Zotino. In this period, the air temperature varied between −39 °C and +7 °C. At temperatures below −10 °C, P. sylvestris needles did not exhibit any variable Chl fluorescence during the daylight period. During the night, however, the effective quantum yield of photosystem 2 (PS2) photochemistry, Φ₂ [Φ₂ = (F_m′ − F_t)/F_m′), increased from values near zero to values between 0.05 and 0.20 depending on the needle temperature and sample investigated. The increase started soon after dusk and lasted for 3–6 h depending on the temperature. A faster increase of Φ₂ was found for temperatures around −16 °C, and lower rates occurred at lower temperatures. Irrespective of the temperature, Φ₂ decreased rapidly to near zero values at dawn, when the photosynthetic photon flux density increased to about 1–5 μmol m⁻² s⁻¹, and remained near zero throughout the day. At temperatures higher than −10 °C, the diurnal decrease and the nocturnal increase of Φ₂ were less distinct or disappeared completely. Hence the winter-adapted Scots pine maintains some photochemical activity of PS2 even at extremely cold temperatures. The capacity of photochemical reactions below −10 °C is, however, very limited and PS2 photochemistry is saturated by an extremely low irradiance (less than 5 μmol m⁻² s⁻¹).