Growth and Dormancy in Norway Spruce Ecotypes (Picea abies) I. Interaction of Photoperiod and Temperature

Growth and dormancy as affected by photoperiod and temperature have been studied in Norway spruce ecotypes of different latitudinal and altitudinal origin. First-year seedlings were used. In all ecotypes apical growth cessation and terminal bud formation occurred within 2 weeks after exposure to SD at temperatures of 18 to 24°C. At lower temperatures or at near-critical photoperiods the response was delayed. The critical photoperiod for apical growth cessation varied from 21 hours in ecotype Steinkjer, Norway (64°N) to about 15 hours in ecotype Lankowitz, Austria (47°04′N). High-elevation ecotypes also had longer critical pholoperiods than low-elevation ecotypes from the same latitude. A detectable growth depression resulted from as little as 1 or 2 SDs of 10 hours, and with 4 or more SDs apical growth cessation took place. In contrast to the situation in the shoot, root growth was not affected by photoperiod. Accordingly, the top:root ratio is drastically affected by photoperiod. The critical photoperiod for cambial growth was shorter than that for apical growth in all ecotypes and cambial growth cessation was delayed for several weeks compared with cessation of apical growth. A transition to formation of late-wood tracheids with thick walls and narrow lumens took place upon exposure to SD. The photoperiodic effects were significantly modified by temperature, but the critical photoperiods were only slightly changed by temperature in the range of 12 to 24°C. However, a 10-hour “night” at 4°C caused growth cessation in continuous light in four ecotypes tested. Temperature optimum for apical growth under non-limiting photoperiods (24 hours) was 21°C in all ecotypes, but with little difference among 18,21 and 24°C. The Q₁₀ for apical growth was 3.5 in the temperature range 12 to 18°C. The growth potential as determined in 24-hour photoperiods was not significantly different among the various ecotypes except for one northern eco-type which was clearly inferior to the others. However, the growth of ecotype Steinkjer (64°N) was greatly suppressed even by the long midsummer days at 59°40′N, thus demonstrating the misleading impression one gets of the growth potential of northern ecotypes when they are moved southwards.     

Effects of Growth Retardants on Norway Spruce (Picea abies)

Young seedlings of Picea abies Karst, grown in nutrient solution were treated with the growth retardants Amo-1618, B-995, and CCC. These were added to the nutrient medium. B-995 and CCC retarded root and shoot growth in the concentrations 100, 10, and 1 mg/l. Growth was almost entirely inhibited by 300 mg/l, obviously due to toxicity. The effects of Amo-1618 were similar but more varying. GA counteracted the effects of all the retardants on shoot growth, but not on root growth.     

Induced Resistance to Pathogenic Fungi in Norway Spruce

Norway spruce (Picea abies) trees (approximately 16 m high) of a single clone were used to study the effects of fungal infection and wounding on induction of resistance to the bark beetle-associated bluestain fungus Ceratocystis polonica. A dose-response experiment was designed involving three different dosages of fungal (fungus and wound) and sterile agar (wound) pretreatment inoculations (10, 50, or 100 inoculations/m² on the stem between 0.8 and 2.0 m high). Three weeks after pretreatment, trees were challenged with a massive C. polonica inoculation (400 inoculations/m²). Control trees that received no pretreatment were heavily colonized and killed by the challenge inoculation. The high and medium fungal pretreatments reduced subsequent fungal colonization success by 76% to 97% relative to the control, and fungal pretreatments protected the trees much more efficiently than sterile agar pretreatments. The protection was demonstrated to be local and not systemic in a subsequent experiment, where trees were pretreated with the medium fungal dosage on the lower bole and challenge inoculated further up the stem. Protection was also demonstrated to be pathogen nonspecific, as trees that had been pretreated with a medium dosage of the root rot fungus Heterobasidion annosum showed enhanced resistance to challenge inoculation with C. polonica.     

Biodegradation of Trichloroacetic Acid in Norway Spruce/Soil System

Trichloroacetic acid (TCA) belongs to secondary atmospheric pollutants affecting the forest health. Distribution of [1,2-¹⁴C]TCA-residues and TCA biodegradation were investigated in 4-year-old nursery-grown trees of Norway spruce [Picea abies (L.) Karst.] in the whole plant/soil system. Radioactivity was monitored in needles, wood, roots and soil as well as in the air. During two weeks of exposure TCA was continuously degraded, especially in the soil. Estimates of radioactivity balance showed loss of radioactivity into the atmosphere in the form of ¹⁴CO₂; unincorporated [1,2-¹⁴C]TCA, chloroform, carbon monoxide and methane were not detected at all. TCA degradation to CO₂ was indicated also in the spruce needles. Moreover, it was found that soil litter contained [1,2-¹⁴C]TCA unavailable to microorganisms.     

Growth Substances and Dormancy in Ceratophyllum demersum

Ceratophyllum demersum L. occurs in winter in the dormant form, in summer in the vegetative form. Factors that affect growth and dormancy in Ceratophyllum were studied. After several weeks of severe winter conditions the plants changed from dormant to quiescent state. Under natural conditions Ceratophyllum plants remain quiescent for several months, due to unfavourable growth conditions. Experimentally the dormant could also be broken by high and low temperature treatments (shocks), and most effectively by addition of GA, An attempt to induce dormancy in full grown plants by the addition of ABA under extreme summer or winter conditions proved unsuccessful. The IAA and ABA contents in the plants were measured during the year. In winter the concentration of ABA was high and that of IAA low, whereas in summer the IAA concentration increased and that of ABA was variable. IAA only slightly antagonized the inhibition of growth by ABA. Both the growth regulators were readily taken up from the culture medium, as was confirmed by a study with the radioactive labelled compounds. The uptake rate of IAA was significantly higher than that of ABA. being 762 μg and 3.26, μg per plant in 24 h, respectively. GA, was found to have a strong antagonistic effect on the ABA induced growth inhibition. The total GA activity in dormant and quiescent plants was similar, in full grown plants it was much lower. In the dormant state a large part of GA was in a bound form, whereas during quiescence relatively more GA occurred in a free state in the plants.     

Gibberellin-like Substances from Norway Spruce (Picea abies)

By use of methanol extraction, two different consecutive partition procedures, repeated polyvinylpyrrolidone column chromatography, silicic acid partition column chromatography and the dwarf rice, lettuce, and barley half-seed bioassays, several gibberellin-like substances were detected in elongating shoots of Picea abies (L.) Karst. No significant differences in the content of gibberellin-like substances could be detected between juvenile and mature trees. The shoot axes were found to contain fewer gibberellin-like substances than the needles.     

Spatial Patterns in Hyphal Growth and Substrate Exploitation within Norway Spruce Stems Colonized by the Pathogenic White-Rot Fungus Heterobasidion parviporum

In Norway spruce, a fungistatic reaction zone with a high pH and enrichment of phenolics is formed in the sapwood facing heartwood colonized by the white-rot fungus Heterobasidion parviporum. Fungal penetration of the reaction zone eventually results in expansion of this xylem defense. To obtain information about mechanisms operating upon heartwood and reaction zone colonization by the pathogen, hyphal growth and wood degradation were investigated using real-time PCR, microscopy, and comparative wood density analysis of naturally colonized trees with extensive stem decay. The hyphae associated with delignified wood at stump level were devoid of any extracellular matrix, whereas incipient decay at the top of decay columns was characterized by a carbohydrate-rich hyphal sheath attaching hyphae to tracheid walls. The amount of pathogen DNA peaked in aniline wood, a narrow darkened tissue at the colony border apparently representing a compromised region of the reaction zone. Vigorous production of pathogen conidiophores occurred in this region. Colonization of aniline wood was characterized by hyphal growth within polyphenolic lumen deposits in tracheids and rays, and the hyphae were fully encased in a carbohydrate-rich extracellular matrix. Together, these data indicate that the interaction of the fungus with the reaction zone involves a local concentration of fungal biomass that forms an efficient translocation channel for nutrients. Finally, the enhanced production of the hyphal sheath may be instrumental in lateral expansion of the decay column beyond the reaction zone boundary.To grow to great heights, trees continually replace their water- and nutrient-conducting elements. Older elements, such as the heartwood that is formed in many trees, gradually become nonconductive. In contrast to the living sapwood, heartwood lacks active defense mechanisms against microbes. However, lignin, the polymer coating cell wall polysaccharides, is highly resistant to microbial degradation. In fact, white-rot fungi, besides having evolved the ability to tolerate or detoxify the secondary metabolites accumulating in heartwood, are the only organisms capable of efficiently degrading lignin. Following establishment in the heartwood of living trees, the colonies of pathogenic white-rot fungi expand and eventually also threaten the conductive sapwood.The white-rot fungus Heterobasidion annosum sensu lato, composed of three species with overlapping geographic distributions and host ranges in Europe (23), is the most important pathogen of Norway spruce (Picea abies L. Karst) in boreal forests. Primary infection of Norway spruce stands by H. annosum sensu lato takes place through fresh thinning stumps or wounds on roots and at the base of the stem. Basidiospores landing on these entrance points give rise to mycelia which colonize the root systems, and eventually the fungus spreads into the stem heartwood. At sites infested with Heterobasidion parviporum, a species primarily restricted to Norway spruce, roots of saplings can become infected by the fungus after around 10 years of growth (25). Stem colonization usually initiates only after the heartwood has started to develop, which in Norway spruce takes place in trees 25 to 40 years old (17). Due to relatively rapid axial spread within heartwood, the decay column caused by H. annosum sensu lato often is up to 10 m high in the stems of mature Norway spruce trees.In response to sapwood challenge by an expanding heartwood-based colony of H. annosum sensu lato, Norway spruce forms a so-called reaction zone (RZ) in the border area between healthy sapwood and colonized heartwood. This xylem defense is characterized by high pH due to increased carbonate content and enrichment of phenolic compounds, particularly lignans, some of which have shown antifungal properties in bioassays (14, 30, 31). Although several wood decay fungi are able to eventually penetrate the RZ regions formed in trees, the strategies employed by fungi to breach these unique defense barriers are poorly understood (24). The purpose of this study was to obtain information about the mechanisms operating in heartwood colonization and expansion of the decay column via penetration of the RZ. To do this, we examined spatial growth of H. parviporum and the associated substrate exploitation patterns within naturally colonized mature stems of Norway spruce.     

Indirect Measurement of Pore Size and Permeability in Scots Pine and Norway Spruce

Five experiments to investigate the water relationships of Scotspine and Norway spruce sapwood are described. Field exposureof stakes, and their moisture characteristic relating wood moisturecontent to the water potential of the surroundings showed thatspruce had a lower moisture content than pine under the sameconditions. A pore size distribution derived from the characteristicsshowed more large pores in spruce than in pine, but in sprucethere were also more small pores. The inference was that thelarge pores of spruce were readily accessible to water, allowinga fast rate of water uptake, as found in the wick action andswelling investigations, while the small pores were not accessible,resulting in a low rate of water uptake and a low rate of swelling.The possibility is briefly discussed that it is the pore sizedistribution of the two woods which determines their water relationships,and the difference in pore size which determines their treatability. Key words: Pinus sylvestris L, Picea abies (L) Karst, Pore size, Water uptake     

Accumulation of a Metallothionein-like mRNA in Norway Spruce Under Environmental Stress

Differential screening of a Norway spruce ( Picea abies L., Karst) cDNA library with probes from damaged and symptomless trees identified a cDNA clone of a 650 nt metallothionein-like (MTL) mRNA as damage-related. Northern blot analysis revealed an about six-fold increase of the MTL mRNA in damaged trees in 1 year and a two-fold increase in the consecutive year. In the N-terminal domain this MTL mRNA comprises typical CysXCys motifs of a subgroup of class II MTL proteins found in kiwi and a few other plant species. In the C-terminal domain, however, the Norway spruce MTL protein is different to the kiwi subgroup and also distinct from a novel Douglas fir metallothionein-related protein, respectively. Hence, the cDNA identified during this study may represent a new type of MTL protein in plants. The accumulation of this MTL mRNA in needles of trees growing in an area of high ozone, sulphur dioxide and nitrogen oxide input and showing clear symptoms of forest decline may be a senescence effect or may play a role in the metal-mediated regulation of the intracellular activated oxygen concentration.     

Changes in Sulfur Metabolism During Needle Development of Norway Spruce*

In a field-study with approx. 150-year-old spruce trees, seasonal changes in thiol composition and content, sulfate content, and oxtractable activities of enzymes of sulfate reduction and assimilation were analyzed in needles and buds and were related to developmental processes during flushing. GSH was the predominant thiol in spruce needles throughout the year, with maximum contents of more than 400nmol g FW^?1 during winter and minimum contents of less than 200 nmol g FW^?1 during summer. Negative correlation of changes in GSI1 contents during winter and spring with air temperature showed a low correlation coefficient (r =– 0.55), suggesting minor significance of GSH in frost protection. Drastic changes in thiol contents during flushing suggested that GSH accumulated in the previous year's needles during winter supports the growing needles with reduced sulfur. Sulfate and other substrates for sulfate assimilation appeared to be available in buds and the new needles. An external supply with reduced sulfur may still be required because of insufficient activities of enzymes of sulfate reduction and assimilation, esp. ATP-sulfurylase and APS-sulfotransferase.     

Effect of Norway Spruce Planting Density on Shoot Morphological Parameters

Temporal and spatial variations of shoot structural parameters, including shoot silhouette to projected needle area ratio, are very important, e.g., for the correction of leaf area index estimated by indirect methods. Here we bring few examples of their evolution within mountain spruce monoculture planted in two different densities.     

Coarse and Fine Control and Annual Changes of Sucrose-Phosphate Synthase in Norway Spruce Needles

Annual changes of activity of sucrose-phosphate synthase (SPS) from spruce (Picea abies [L.] Karst.) needles were studied with respect to three regulatory levels: metabolic fine control, covalent modification (phosphorylation), and protein amount. Glucose-6-phosphate served as an allosteric activator of spruce SPS by shifting the Michaelis constant for the substrate fructose-6-phosphate from 4.2 to 0.59 mM, whereas inorganic phosphate competitively inhibited this activation. The affinity for the other substrate, UDP-glucose, was unaffected. Incubation of the crude extract with ATP resulted in a time- and concentration-dependent decrease of the maximal velocity of SPS. This inactivation was sensitive to staurosporine, a potent protein kinase inhibitor, indicating the participation of a protein kinase. Probing SPS protein with heterologous antibodies showed that the subunit of spruce SPS is an approximately 139-kD protein and that changes in the extractable activity during the course of a year were correlated with the amount of SPS protein. High SPS activities in winter were paralleled by increased levels of the activator glucose-6-phosphate and the substrate fructose-6-phosphate, indicating a high capacity for sucrose synthesis that may be necessary to maintain photosynthetic CO2 fixation in cold-hardened spruce needles.     

Induction of Adventitious Buds on Embryos of Norway Spruce Grown in vitro

Adventitious buds were induced when isolated embryos of Norway spruce (Picea abies L.) were cultured on a defined medium containing 2iP. Anatomically different bud primordia were formed at high and low cytoldnin concentrations. The highest percentage of embryos forming bud primordia was obtained after 4–5 weeks on 2iP medium. Buds developed after transfer of the induced embryos to medium without growth regulators. Many of the buds developed into elongated shoots. No root primordia were observed in any of the induced embryos.     

Actin Distribution in Mitotic Apparatus of Somatic Embryo Cells of Norway Spruce

F-actin distribution was studied in mitotic cells of embryogenic suspension culture of Norway spruce [Picea abies (L.) Karst.]. Actin was present in dividing cells of embryo head during whole mitosis. Transient co-localization of actin microfilaments with preprophase band of microtubules was observed. Weak actin staining occurred with non-kinetochor microtubular fibers in metaphase spindle. F-actin was not localized with kinetochore microtubular fibres in metaphase as well as with shortening kinetochore fibres in late anaphase. On the other hand, abundant actin microfilaments array was formed in the area of late anaphase spindle in equatorial level of the cell between separating chromatids. F-actin was also present in phragmoplast area in telophase. This revised version was published online in July 2006 with corrections to the Cover Date.     

Apoplastic Peroxidases and Lignification in Needles of Norway Spruce (Picea abies L.)

The objective of the present study was to investigate the correlation of soluble apoplastic peroxidase activity with lignification in needles of field-grown Norway spruce (Picea abies L.) trees. Apoplastic peroxidases (EC 1.11.1.7) were obtained by vacuum infiltration of needles. The lignin content of isolated cell walls was determined by the acetyl bromide method. Accumulation of lignin and seasonal variations of apoplastic peroxidase activities were studied in the first year of needle development. The major phase of lignification started after bud break and was terminated about 4 weeks later. This phase correlated with a transient increase in apoplastic guaiacol and coniferyl alcohol peroxidase activity. NADH oxidase activity, which is thought to sustain peroxidase activity by production of H2O2, peaked sharply after bud break and decreased during the lignification period. Histochemical localization of peroxidase with guaiacol indicated that high activities were present in lignifying cell walls. In mature needles, lignin was localized in walls of most needle tissues including mesophyll cells, and corresponded to 80 to 130 [mu]mol lignin monomers/g needle dry weight. Isoelectric focusing of apoplastic washing fluids and activity staining with guaiacol showed the presence of strongly alkaline peroxidases (isoelectric point [greater than or equal to] 9) in all developmental stages investigated. New isozymes with isoelectric points of 7.1 and 8.1 appeared during the major phase of lignification. These isozymes disappeared after lignification was terminated. A strong increase in peroxidase activity in autumn was associated with the appearance of acidic peroxidases (isoelectric point [less than or equal to] 3). These results suggest that soluble alkaline apoplastic peroxidases participate in lignin formation. Soluble acidic apoplastic peroxidases were apparently unrelated to developmentally regulated lignification in spruce needles.     

Changes in Sapwood of Roots of Norway Spruce, Attacked by Fomes annosus

Acetone extracts of sapwood and reaction zone of spruce roots attacked by Fomes annosus, collected in February, June and October, were separated into resinous and phenolic fractions. The fractions were further separated by column, thin layer and gas liquid chromatography, followed by biological tests, using Fomes annosus and other rot fungi. The reaction zone contained quantitatively less light petroleum soluble compounds than the sapwood but more acids. The phenolic content was about ten times higher in the reaction zone than in the sapwood. Nine lignans and one simple phenol (4-methylcatechol) were identified and quantitatively estimated in the reaction zone. The resinous fraction of the extract from the reaction zone as well as some of the lignans and 4-methylcatechol inhibited fungal growth, in some cases followed by detoxification and continued growth. The predominant lignan, hydroxymatairesinol, had no effect on Fomes annosus or five other wood degrading fungi. About 15 unidentified phenols were observed, some of them probably of importance as inhibitors, either alone or in combination with other phenols.