Characterization of an acetylated heteroxylan from Eucalyptus globulus Labill

A heteroxylan was isolated from Eucalyptus globulus wood by extraction of peracetic acid delignified holocellulose with dimethyl sulfoxide. Besides (1-->4)-linked beta-D-xylopyranosyl units of the backbone and short side chains of terminal (1-->2)-linked 4-O-methyl-alpha-D-glucuronosyl residues (MeGlcA) in a 1:10 molar ratio, this hemicellulose contained galactosyl and glucosyl units attached at O-2 of MeGlcA originating from rhamnoarabinogalactan and glucan backbones, respectively. About 30% of MeGlcA units were branched at O-2. The O-acetyl-(4-O-methylglucurono)xylan showed an acetylation degree of 0.61, as determined by 1H NMR spectroscopy, and a weight-average molecular weight (M(w)) of about 36 kDa (P=1.05) as revealed from size-exclusion chromatography (SEC) analysis. About half of the beta-D-xylopyranosyl units of the backbone were found as acetylated moieties at O-3 (34 mol%), O-2 (15 mol%) or O-2,3 (6 mol%). Practically, all beta-D-xylopyranosyl units linked at O-2 with MeGlcA residues were 3-O-acetylated (10 mol%).     

Growth dynamics of superficial roots in Portuguese plantations ofEucalyptus globulus Labill. studied with a mesh bag technique

Summary The variation in growth of the fine roots of blue gum (Eucalyptus globulus labill. ssp.globulus) in the 0–40 cm soil layer was studied from March 1982 to March 1983 at Quinta do Furaduoro, Óbidos, Portugal. A mesh bag method was used; bags of nylon net were inserted into a clay soil and a sandy soil and filled with root-free soil. They were resampled after 2, 4, 6 and 12 months in both places and, in a separate series in the sandy soil every second month throughout the year.The ingrowth of roots was high during the winter months but there was also a surprisingly high ingrowth during the spring-early summer period. There was also some root growth during the driest part of the yearviz. July–September.The amount of fine roots reached a maximum of about 260 g dw m^–2 after about 6 months in the sandy soil, whereas it took at least 12 months to reach the somewhat higher level of 450 g dw m^–2 in the clay soil. At that level the decomposition of dead roots was expected to equal the formation of new roots. Dead roots appeared after only 2 months. There was a higher proportion of dead roots in the clay soil than in the sandy soil, 35% as compared with 20% on an average, which indicates a slower decomposition or a higher mortality at equal decomposition rates in the clay than and in the sandy soil. The present data gives an indication of a minimum fine root production in mature Eucalyptus stands of at least 600 g dw m^–2 yr^–1.     

In vitro studies on Eucalyptus globulus rooting ability

Summary Micropropagation has the potential to quickly introduce selected genotypes of adult Eucalyptus globulus clones and it is now widely used in Portugal as a part of genetic improvement programs. Several clones have been established and multiplied in vitro. The different clones have individual requirements for successful rooting. Rejuvenation was achieved at different periods after culture initiation for the different clones. Subculturing preceding rooting in multiplication medium supplemented with riboflavin and cholene chloride allowed the increase of rooting ability for several clones tested. Removal of boron from the rooting medium increased rooting by 10%. Indolebutyric acid (IBA) dipping before transfer to the rooting medium resulted in a rooting percentage of 80–95% for the best clones tested. Acclimatization was performed without difficulties (90–95% success) and the rooted plants were either planted directly or used as mother plants for further cutting production, depending on the needs. The results described in this paper increase the commercial feasibility of the micropropagation system for E. globulus.     

Boron requirements for Eucalyptus globulus seedlings

Although boron (B) deficiency limits the productivity of eucalypts in plantations in many parts of the world, the concentrations of foliar B used in the diagnosis of B deficiency vary greatly among studies. There has been a lack of reliable diagnosis standards for B deficiency in Eucalyptus species. Therefore, the present study investigated the relationship between internal and external B concentrations and growth of Eucalyptus globulus, the main commercial temperate eucalypt species. Seedlings were grown in a B-buffered solution culture (Amberlite IRA 743) from 0.03 to 8.35 M B. Boron deficiency symptoms appeared at day 5 in the nutrient solution containing less than 0.27 M B. The external critical B concentrations, estimated for the growth of shoots and roots, were 1.08 and 0.99 M B, respectively. The internal critical B concentration range in the youngest fully expanded leaf (YFEL) for shoot growth was 12–16 mg B kg^–1 dry weight. The internal critical B concentrations estimated in the present study have been successfully used in the diagnosis of B deficiency in E. globulus trees up to three years of age in south-east Asia.     

Early ovule development following self- and cross-pollinations in Eucalyptus globulus Labill. ssp. globulus

The study was conducted to identify the self-incompatibility mechanism in Eucalyptus globulus ssp. globulus. Controlled self- and cross-pollinations were conducted on individual flowers from three mature trees that had self-incompatibility levels of 76, 99.6 and 100%. Flowers were harvested at 4, 6 and 8 weeks after pollination. Embryology was investigated by bright field microscopy on material harvested at 4 and 6 weeks after pollination. Fertilization had taken place at 4 weeks after pollination with zygotes and free nuclear endosperm visible. There was a greater proportion of healthy, fertilized ovules in the cross- compared with the self-pollination treatment, and approx. half the ovules examined from both pollen treatments were not fertilized or were degenerating. By 6 weeks after pollination a few zygotes were starting to divide. The number of healthy, fertilized ovules was still greater in the cross-pollination treatment, but the number of healthy fertilized ovules was lower in both treatments compared with 4 weeks after pollination, and many ovules were degenerating. Fertilized ovules were significantly larger than non-fertilized or degenerating ovules and this difference was detectable by eye at 6 and 8 weeks after pollination. The mechanism of self-incompatibility appears to have both late pre- and post-zygotic components.     

Ontogeny of in vitro rooting processes in Eucalyptus globulus

Summary In the present work, histological changes observed at the base of Eucalyptus globulus shoots in in vitro culture are described. Shoots were placed on solidified Murashige and Skoog medium containing half the original salt concentration, the complete vitamin composition, 9.8 μM indolebutyric acid (IBA), and 30 gI⁻¹ agar, and were incubated in the dark for the first 7 d, followed by a 16-h photoperiod. In vitro-generated roots could be originated either from old vascular tissue or from newly formed xylem. The influence of the preexistent tissues on the neoformation process appeared to be varied. The medulla did not intervene directly, although there were abundant cellular divisions in response to the induction medium. On the other hand, the interruptions observed in the vascular cylinder of the stem suggested an influence of the interfascicular parenchyma, and therefore the medulla could have participated in the differentiation process. However, the cortical parenchyma showed most of the changes that lead to the formation of adventitious roots of E. globulus growing in vitro. Histological analysis suggests that vascular rays can also be formed in direct contact with the central cylinder of the stem, although they mainly originate from the cortical parenchyma.     

Leaf and root growth dynamics in Eucalyptus globulus seedlings grown in drying soil

Summary Seedlings of Eucalyptus globulus growing in soil columns were subjected to a 24 day soil drying treatment. Water and solute potentials of both young expanding and fully expanded leaves declined under reduced soil water availability, while slightly higher turgor was sustained by the fully expanded leaves. Although leaf area of unwatered seedlings was smaller, the corresponding leaf dry weight was quite similar to that of well-watered seedlings. Soon after rewatering, leaf area of plants experiencing water shortage was comparable to that of well-watered plants. It seems that a difference in wall properties between juvenile and mature leaves allows for an effective pattern of water use by eucalypt plants growing in drying soil. Some stomatal opening is sustained and therefore, presumably, some carbon may be fixed, keeping the carbon balance of the whole plant positive, and allowing a continuous cell division despite the limited water supply. The highest root density of both well-watered and unwatered plants was found in the upper soil layers. However, root growth of unwatered seedlings was gradually increased in the deeper soil layers, where thicker root apices and higher soil water depletion rates per unit root length were recorded. As a consequence, root absorbing surface area was as large in unwatered plants as in well-watered plants.     

Genotype by environment interaction for growth of Eucalyptus globulus in Australia

This study assessed the genotype by environment (G × E) interaction for diameter growth in 15 Eucalyptus globulus progeny trials in Australia. Single-site analyses revealed significant subrace and family-within-subrace variance in all trials. Across-site subrace () and family () correlations were estimated by linear mixed model analyses of pairs of trials. Using a factor analytic structure for subrace and family random terms in a multi-environment mixed model analysis, best linear unbiased predictions of subrace effects were obtained for each trial. These were then averaged for each of four states (Victoria, Tasmania, South Australia and Western Australia) and across all sites. Statistically significant G × E interaction was detected, and weighted means across states for and were 0.73 and 0.76, respectively. Nevertheless, the three subraces from the Otway Ranges were both fast growing and relatively stable in their ranks over all sites. We evaluated the sensitivity of subraces to changing environmental conditions, on the basis of random coefficient models regressing subrace performance on selected trial climatic variables. The results suggested differential susceptibility of subraces to water, light and (to a less extent) temperature stresses during summer. Moreover, using multivariate techniques to visualize and interpret the across-site correlation structure for subrace effects, we could identify site clusters of reduced G × E interaction related to soil water availability and evaporative demand during summer. A revised site-type classification using these factors should allow a better capture of genetic gains from breeding and deployment.     

Within-tree variation in cambial anatomy and xylem cell differentiation in Eucalyptus globulus

Summary Microscopic examination of the vascular cambium of Eucalyptus globulus, from four trees, at nine height levels, revealed a non-storied cambial structure with both ray and fusiform initials. Average fusiform initial length increased from 443 m at 70% of tree height from the base to a maximum of 484 m at 10% of tree height. At 2.5% of tree height average fusiform initial length sharply declined to 438 m. A strong positive correlation (r = 0.757, df = 7, P <0.05) existed between average fusiform initial length and average fibre length at each height, giving quantitative support to the notion that variation in fusiform initial length is an important mechanism influencing fibre length in hardwoods. However, 62% of the longitudinal variation in average fibre length was explained by the extent of elongation during differentiation, which increased linearly down the stem. At 70% of tree height, the average length of fibres was 1.84 times that of the fusiform initials, compared to 2.11 times at 2.5% of tree height. Concomitant with this longitudinal gradient of elongation was a gradient of increasing tangential area of the cambium occupied by cambial rays. This was accounted for by dimensional increases in the ray initial cells with decreasing height. Possible mechanisms that regulate the elongation of developing fibres and the proportion of cambial rays are discussed.     

Variation of heartwood and sapwood in 18-year-old Eucalyptus globulus trees grown with different spacings

Heartwood and sapwood development was studied in 18-year-old Eucalyptus globulus trees from pulpwood plantations with different spacings (3 × 2, 3 × 3, 4 × 3, 4 × 4 and 4 × 5 m), on cross-sectional discs taken at breast height. The trees possessed a large proportion of heartwood, on average 60% of the wood cross-sectional surface. Spacing was a statistically significant source of variation of heartwood area, which ranged between 99 and 206 cm² for the closer (3 × 2) and wider (4 × 5) spacings, respectively. There was a positive and high statistical significant correlation between heartwood diameter and tree diameter (heartwood diameter = −0.272 + 0.616 dbh; r ² = 0.77; P < 0.001), and larger trees contained more heartwood regardless of spacing. Heartwood proportion in cross-section remained practically constant between spacings but increased with tree diameter class: 55.1, 62.2, 65.0 and 69.5% for diameter at breast height classes <15, 15–20, 20–25 and >25 cm, respectively. The sapwood width did not depend on tree diameter growth and remained practically constant at an average of 18 mm (range 15–21 mm), but sapwood area showed a good linear regression with tree diameter. Therefore, tree growth enhancement factors, such as wide spacings, will induce formation of larger heartwoods that can negatively impact raw-material quality for pulping. The increase in heartwood in relation with tree dimensions should therefore be taken into account when designing forest management guidelines.     

Detection and stability of quantitative trait loci (QTL) in Eucalyptus globulus

Eucalyptus globulus Labill. ssp. globulus is an important tree species for the pulp and paper industry, and several breeding programmes throughout the world are striving to improve key traits such as growth and wood density. This study aimed to detect quantitative trait loci (QTL) for growth, wood density, relative bark thickness and early flowering in a single full-sib E. globulus family grown across seven sites. Growth was measured a number of times over a 6-year period, enabling temporal stability of growth QTL to be studied. Ten putative QTL (LOD > 2.0) were detected in the single family, which was of moderate size. Based on permutations of the trait data, six of these QTL were significant at the experimentwise significance level of 0.1 for at least one of the four models implemented in analysis to remove site effects. For wood density, two putative QTL explained 20% of the variance for the trait, indicating that a small number of QTL might explain a reasonable proportion of the trait variance. One of these QTL was found to be independent of QTL for growth whereas the second QTL co-segregated with a QTL for relative incremental growth. The marker nearest to this QTL was associated with fast growth but low wood density. A putative growth QTL at year 6 was found to be relatively stable across ages. In addition, it was found that residuals from models based on measurements from across all families across all sites in the trial detected QTL with greater experimentwise significance.     

The relationship of the female reproductive success of Eucalyptus globulus to the endogenous properties of the flower

Low capsule and seed set is a major factor limiting seed production in Eucalyptus globulus seed orchards. Controlled pollination studies showed that the reproductive success (number of seeds produced per flower pollinated) was primarily determined by the female. We aimed to identify the factors contributing to the differences in reproductive success between female genotypes in terms of the physical and anatomical properties of the flower. We studied pairs of genotypes of high and low reproductive success from each of three races (Furneaux Group, Strzelecki Ranges and Western Otways) growing in a seed orchard. Controlled pollinations were performed on six females and along with flower physical measurements, pollen tube growth and seed set were assessed. Overall tree reproductive success was positively correlated with flower size, ovule numbers, style size, cross-sectional area of conductive tissue within the style (all of which were inter-correlated) and the proportion of pollen tubes reaching the bottom of the style. Significant positive correlations of reproductive success and flower physical properties between different ramets of the same genotypes across seasons suggests a genetic basis to the variation observed. The majority of pollen tube attrition occurred within the first millimetre of the cut style and appeared to be associated with differences in style physiology. When examined as pairs within races the difference in reproductive success for the Western Otways pair was simply explained by differences in flower size and the number of ovules per flower. Physical features did not differ significantly for the Strzelecki Ranges pair, but the proportion of pollen tubes reaching the bottom of the style was lower in the less reproductively successful genotype, suggesting an endogenous physiological constraint to pollen tube growth. The difference in reproductive success between the females from the Furneaux Group was associated with a combination of these factors.     

Decomposition of Eucalyptus globulus leaves and three native leaf species (Alnus glutinosa, Castanea sativa and Quercus faginea) in a Portuguese low order stream

Leaf decomposition of the exotic evergreen Eucalyptus globulus (eucalyptus), and three native deciduous tree species, Alnus glutinosa (alder), Castanea sativa (chestnut) and Quercus faginea (oak), was compared in a second order stream in Central Portugal. Changes in dry weight, nitrogen and polyphenolic compounds and microbial colonization were periodically assessed for three months.Negative exponential curves fit the leaf weight loss with time for all leaf species. Mass loss rate was in the order alder (K = 0.0161) > chestnut (K = 0.0079) > eucalyptus (K = 0.0068) > oak (K = 0.0037). Microbial colonization followed the same pattern as breakdown rates. Evidence of fungal colonization was observed in alder after 3 days in the stream, whereas it took 21 days in oak leaves to have fungal colonization. Fungal diversity was leaf species-dependent and increased with time. In all cases, percent nitrogen per unit leaf weight increased, at least, at the initial stages of decay while soluble polyphenolics (expressed as percentage per unit leaf weight) decreased rapidly in the first month of leaves immersion.Intrinsic factors such as nitrogen and polyphenolic content may explain differences in leaf decomposition. The possible incorporation of eucalyptus litter into secondary production in a reasonable time span is suggested, although community balance and structure might be affected by differences in allochthonous patterns determined by eucalyptus monocultures.     

Impact of a eucalyptus (Eucalyptus globulus Labill.) plantation on the nutrient content and dynamics of coarse particulate organic matter (CPOM) in a small stream

Litterfall inputs, benthic storage and the transport of coarse particulate organic matter (CPOM) were studied in two headwater streams, one flowing through a mixed deciduous forest and one through a plantation of Eucalyptus globulus. Vertical and lateral traps, transported CPOM and benthic CPOM were sampled monthly to biweekly and sorted into four categories: leaves, twigs and bark, fruits and flowers and debris. The litterfall inputs were about 20% lower at the eucalyptus site but this reduction was unevenly distributed among the litter categories. The reduction of the nitrogen and phosphorus inputs was larger (50%) than that of CPOM because of the low nutrient concentration of the CPOM at the eucalyptus site. Transported CPOM was also lower at the eucalyptus site. Although total CPOM inputs to the stream were reduced in the eucalyptus plantation, benthic storage of CPOM was 50% higher due to (1) high inputs of CPOM and low discharge during summer, (2) more twig and bark inputs, (3) eucalyptus leaves being retained more efficiently in the stream than deciduous leaves (4) a lower discharge, which may in part be attributable to eucalyptus-induced changes in the hydrological cycle. Increased retention balanced lower nitrogen and phosphorus content of CPOM, so benthic storage of nitrogen and phosphorus was similar at both sites. This work demonstrates that the timing, quality and quantity of inputs and benthic storage of CPOM in streams changes substantially because of the substitution of natural deciduous forest with eucalyptus plantation. Maintenance of buffer strips of natural vegetation may be the best way to protect ecological functioning of small, forested streams.     

Impacts of silviculture on genetic diversity in the native forest species Eucalyptus sieberi

Potential impacts of regeneration practices ongenetic diversity in the Australian nativeforest species Eucalyptus sieberi L.A.S.Johnson. (silvertop ash) were assessed usingDNA markers. Three different silviculturaltreatments were examined: clear-felling withaerial re-sowing, and the seed tree system withsite preparation by either burning ormechanical disturbance. In addition, twounharvested stands were chosen as controls. Atotal sample of 825 trees were genotyped at 35Mendelian markers: 26 single-copy nuclear RFLPsand 9 microsatellites. No significantdifferences were found among the treatments inany of four population genetic statistics:allelic richness, effective number of alleles,expected heterozygosity and the panmictic index(f). Rare alleles were prevalent, and a MonteCarlo simulation showed that the apparent lossof four rare alleles from the saplingregenerants was highly statisticallysignificant. There was no evidence for recentbottlenecks from analyses of either the levelsof expected heterozygosity relative to thatexpected under mutation drift equilibrium, orthe allele frequency profiles. A dendrogram ofthe relationships between the sampledpopulations suggested that the seed tree systemmay result in the promotion of genetic drift(slight expansion of the dendrogram) whileaerial re-sowing of clear falls with the sameseedlot will lead to genetic homogenisation(contraction of the dendrogram). The apparentgenetic robustness of E. sieberi tonative forest regeneration practices isattributed to its local abundance combined withthe favourable properties of its reproductivebiology, as well as to the limitation that onlya single rotation was examined.     

Diagnosis of nitrogen deficiency and toxicity of Eucalyptus globulus seedlings by foliar analysis

The relationship between shoot growth and foliar nitrogen (N) in E. globulus seedlings was studied in the glasshouse to determine standard values for N deficiency and toxicity diagnosis. Seedlings were grown for 9 weeks in yellow sand, at 10 rates of N, applied as ammonium sulphate, calcium nitrate or ammonium nitrate. Shoot dry weight (DW) increased linearly with N rate for all forms of N in the deficiency range. Seedlings continued to respond to higher rates of ammonium and ammonium nitrate than to nitrate. Maximum shoot DW for nitrate fed plants and ammonium nitrate fed plants were 51% and 84% respectively of ammonium fed plants. Total N concentration in the youngest fully expanded leaf (YFEL) ranged from 1.0% to 3.3% in deficient and adequate plants. The critical N concentration for deficiency diagnosis (corresponding to 90% maximum yield) in the YFEL, determined from these growth response curves averaged over all N forms, was 2.6% N. For ammonium nitrate fed plants, total N concentration in the YFEL for the severely deficient, deficient, adequate, and toxic ranges were <1.4%, 1.4–2.5%, 2.6–3.5%, > 4.3%. High total N concentrations were associated with growth depression and toxicity symptoms, which differed with N form. For nitrate fed plants, a total N concentration above 3.3% in the YFEL was associated with severe growth depression, and leaf tip necrosis. The adequate concentration range for ammonium nitrate was similar to values found on a field trial with 7 month old E. globulus trees grown on an exforest site.     

Bioactive monoterpene glycosides conjugated with gallic acid from the leaves of Eucalyptus globulus

Two monoterpene glycosides, conjugated with gallic acid [globulusin A (1) and B (2)], together with four known compounds, cypellocarpin A (3), eucaglobulin (4), cuniloside (5) and (1S, 2S, 4R)-trans-2-hydroxy-1,8-cineole beta-d-glucopyranoside (6), were isolated from hot-water extracts of the leaves of Eucalyptus globulus. The structures of compounds 1 and 2 were determined by 1D, 2D NMR and MS spectroscopic analyses. The absolute stereochemistry of 1 was determined by correlating the spectroscopic data with those of synthetic compound 6 with a known configuration. Globulusin A (1) and B (2), cypellocarpin A (3) and eucaglobulin (4), scavenged DPPH free radicals and globulusin A (1) showed a higher antioxidant activity than the other tested compounds, with an IC50 of 3.8microM. Globulusin A (1) and eucaglobulin (4) concentration-dependently suppressed inflammatory cytokine production, tumor-necrosis factor-alpha and interleukin-1beta in cultured human myeloma THP-1 cells co-stimulated with phorbol myristate acetate. These compounds also inhibited melanogenesis in cultured murine melanoma B16F1 cells, without any significant cytotoxicity. These results suggested that globulusin A (1) and eucaglobulin (4), which were isolated as antioxidants from E. globulus, also had anti-inflammatory as well as anti-melanogenesis activity.     

Genetic control of somatic embryogenesis induction in <Emphasis Type="Italic">Eucalyptus globulus</Emphasis> Labill.

A reproducible protocol for somatic embryogenesis (SE) induction in Eucalyptus globulus from mature zygotic embryos is available since 2002. However, for the use of SE in tree breeding programs, the frequency of SE initiation needs to be improved and controlled, and this was investigated in 13 open-pollinated (OP) families over three consecutive years. A diallel mating design with five parent trees was used to study genetic control of SE induction. Results showed that SE induction varies across E. globulus families and over the years of seed production tested. Somatic embryogenesis was initiated on explants from 84% of the OP families tested in 2002 and 100% of the families tested in 2003 and 2004. The year 2003 gave best results for percentage of induction and total number of somatic embryos produced. Results concerning genetic control showed that SE induction is under the control of additive genetic effects, as 22.0% of variation in SE initiation was due to general combining ability (GCA) effect, whereas 6.4% was due to maternal effects. Neither specific combining ability (SCA) nor reciprocal effects were significant.     

Genetic parameters for growth, wood density and pulp yield in Eucalyptus globulus

Genetic variation and co-variation among the key pulpwood selection traits for Eucalyptus globulus were estimated for a range of sites in Portugal, with the aim of improving genetic parameters used to predict breeding values and correlated response to selection. The trials comprised clonally replicated full-sib families (eight trials) and unrelated clones (17 trials), and exhibited varying levels of pedigree connectivity. The traits studied were stem diameter at breast height, Pilodyn penetration (an indirect measure of wood basic density) and near infrared reflectance predicted pulp yield. Univariate and multivariate linear mixed models were fitted within and across sites, and estimates of additive genetic, total genetic, environmental and phenotypic variances and covariances were obtained. All traits studied exhibited significant levels of additive genetic variation. The average estimated within-site narrow-sense heritability was 0.19 ± 0.03 for diameter and 0.29 ± 0.03 for Pilodyn penetration, and the pooled estimate for predicted pulp yield was 0.42 ± 0.14. When they could be tested, dominance and epistatic effects were generally not statistically significant, although broad-sense heritability estimates were slightly higher than narrow-sense heritability estimates. Averaged across trials, positive additive (0.64 ± 0.08), total genetic (0.58 ± 0.04), environmental (0.38 ± 0.03) and phenotypic (0.43 ± 0.02) correlation estimates were consistently obtained between diameter and Pilodyn penetration. This data argues for at least some form of pleiotropic relationship between these two traits and that selection for fast growth will adversely affect wood density in this population. Estimates of the across-site genetic correlations for diameter and Pilodyn penetration were high, indicating that the genotype by environment interaction is low across the range of sites tested. This result supports the use of single aggregated selection criteria for growth and wood density across planting environments in Portugal, as opposed to having to select for performance in different environments.