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The leaf intercellular airspace is a tortuous environment consisting of cells of different shapes, packing densities, and orientation, all of which have an effect on the travelling distance of molecules from the stomata to the mesophyll cell surfaces. Tortuosity, the increase in displacement over the actual distance between two points, is typically defined as encompassing the whole leaf airspace, but heterogeneity in pore dimensions and orientation between the spongy and palisade mesophyll likely result in heterogeneity in tortuosity along different axes and would predict longer traveling distance along the path of least tortuosity, such as vertically within the columnar cell matrix of the palisade layer. Here, we compare a previously established geometric method to a random walk approach, novel for this analysis in plant leaves, in four different Eucalyptus species. The random walk method allowed us to quantify directional tortuosity across the whole leaf profile, and separately for the spongy and palisade mesophyll. For all species tortuosity was higher in the palisade mesophyll than the spongy mesophyll and horizontal (parallel to the epidermis) tortuosity was consistently higher than vertical (from epidermis to epidermis) tortuosity. We demonstrate that a random walk approach improves on previous geometric approaches and is valuable for investigating CO 2 and H 2 O transport within leaves., (© 2021 John Wiley & Sons Ltd.)

The pulse of the tree (diurnal cycle of stem radius fluctuations) has been widely studied as a way of analyzing tree responses to the environment, including the phenotypic plasticity of tree-water relationships in particular. However, the genetic basis of this daily phenotype and its interplay with the environment remain largely unexplored. We characterized the genetic and environmental determinants of this response, by monitoring daily stem radius fluctuation (dSRF) on 210 trees from a Eucalyptus urophylla × E. grandis full-sib family over 2 years. The dSRF signal was broken down into hydraulic capacitance, assessed as the daily amplitude of shrinkage (DA), and net growth, estimated as the change in maximum radius between two consecutive days (ΔR). The environmental determinants of these two traits were clearly different: DA was positively correlated with atmospheric variables relating to water demand, while ΔR was associated with soil water content. The heritability for these two traits ranged from low to moderate over time, revealing a time-dependent or environment-dependent complex genetic determinism. We identified 686 and 384 daily quantitative trait loci (QTL) representing 32 and 31 QTL regions for DA and ΔR, respectively. The identification of gene networks underlying the 27 major genomics regions for both traits generated additional hypotheses concerning the biological mechanisms involved in response to water demand and supply. This study highlights that environmentally induced changes in daily stem radius fluctuation are genetically controlled in trees and suggests that these daily responses integrated over time shape the genetic architecture of mature traits., (© 2020 Society for Experimental Biology and John Wiley & Sons Ltd.)

Genomic Best Linear Unbiased Prediction (GBLUP) in tree breeding typically only uses information from genotyped trees. However, information from phenotyped but non-genotyped trees can also be highly valuable. The single-step GBLUP approach (ssGBLUP) allows genomic prediction to take into account both genotyped and non-genotyped trees simultaneously in a single evaluation. In this study, we investigated the advantage, in terms of breeding value accuracy and bias, of including phenotypic observation from non-genotyped trees in a standard tree GBLUP evaluation. We compared the efficiency of the conventional pedigree-based (ABLUP), GBLUP and ssGBLUP approaches to evaluate eight growth and wood quality traits in a Eucalyptus hybrid population, genotyped with 33,398 single nucleotide polymorphisms (SNPs) using the EucHIP60k. Theoretical accuracies, predictive ability and bias were calculated by ten-fold cross validation on all traits. The use of additional phenotypic information from non-genotyped trees by means of ssGBLUP provided higher predictive ability (from 37% to 75%) and lower prediction bias (from 21% to 73%) for the genetic component of non-phenotyped but genotyped trees when compared to GBLUP. The increase (decrease) in the prediction accuracy (bias) became stronger as trait heritability decreased. We concluded that ssGBLUP is a promising breeding tool to improve accuracies and bias over classical GBLUP for genomic evaluation in Eucalyptus breeding practice., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Premise: Cambial activity in some tropical trees varies intra-annually, with the formation of xylem rings. Identification of the climatic factors that regulate cambial activity is important for understanding the growth of such species. We analyzed the relationship between climatic factors and cambial activity in four tropical hardwoods, Acacia mangium, Tectona grandis, Eucalyptus urophylla, and Neolamarckia cadamba in Yogyakarta, Java Island, Indonesia, which has a rainy season (November-June) and a dry season (July-October)., Methods: Small blocks containing phloem, cambium, and xylem were collected from main stems in January 2014, October 2015 and October 2016, and examined with light microscopy for cambial cell division, fusiform cambial cells, and expanding xylem cells as evidence of cambial activity., Results: During the rainy season, when precipitation was high, cambium was active. By contrast, during the dry season in 2015, when there was no precipitation, cambium was dormant. However, in October 2016, during the so-called dry season, cambium was active, cell division was conspicuous, and a new xylem ring formation was initiated. The difference in cambial activity appeared to be related to an unusual pattern of precipitation during the typically dry months, from July to October, in 2016., Conclusions: Our results indicate that low or absent precipitation for 3 to 4 months induces cessation of cambial activity and temporal periodicity of wood formation in the four species studied. By contrast, in the event of continuing precipitation, cambial activity in the same trees may continue throughout the year. The frequency pattern of precipitation appears to be an important determinant of wood formation in tropical trees., (© 2019 Botanical Society of America.)

Seed shape, surface cells shape, arrangement, anticlinal wall pattern, and periclinal wall protuberances were recorded for nine species of Eucalyptus (Myrtaceae) using scanning electron microscopy to determine the importance of seed morphological characters as an additional tool for identification. Most of the seeds were found ovate and some seeds were elliptic and cuboid in shape. Almost reticulate regular seed surface patterns were observed. Four types of surface cells were examined; diamond, elliptic, oblong, and irregular. Majority of the seeds showed raised anticlinal wall level and diversity from wavy to puzzle in pattern. Periclinal wall may be glabrous or having protuberances that were rhombus and bullate in shape. Both macro- and micromorphological characters can provide basis for classification and delimitation of genus Eucalyptus. RESEARCH HIGHLIGHTS: Seeds quantitative characters of nine Eucalyptus species as seed length, width, and weight. Macromorphological characters of seeds including seed color, helium position, and seed shape. Micromorphological characters include seed surface, periclinal wall, and anticlinal wall investigation under scanning electron microscope. Ultra-seed sculpturing features as an additional tool in identification., (© 2018 Wiley Periodicals, Inc.)

Background: Wood basic density (WBD), the biomass of plant cell walls per unit volume, is an important trait for elite tree selection in kraft pulp production. Here, we investigated the correlation between WBD and wood volumes or wood properties using 98 open-pollinated, 2.4 to 2.8 year-old hybrid Eucalyptus (Eucalyptus urophylla x E. grandis). Transcript levels of lignocellulose biosynthesis-related genes were studied., Results: The progeny plants had average WBD of 516 kg/m 3 with normal distribution and did not show any correlations between WBD and wood volume or components of α-cellulose, hemicellulose and Klason lignin content. Transcriptomic analysis of two groups of five plants each with high (570-609 kg/m 3 ) or low (378-409 kg/m 3 ) WBD was carried out by RNA-Seq analysis with total RNAs extracted from developing xylem tissues at a breast height. Lignocellulose biosynthesis-related genes, such as cellulose synthase, invertase, cinnamate-4-hydroxylase and cinnamoyl-CoA reductase showed higher transcript levels in the high WBD group. Among plant cell wall modifying genes, increased transcript levels of several expansin and xyloglucan endo-transglycosylase/hydrolase genes were also found in high WBD plants. Interestingly, strong transcript levels of several cytoskeleton genes encoding tubulin, actin and myosin were observed in high WBD plants. Furthermore, we also found elevated transcript levels of genes encoding NAC, MYB, basic helix-loop-helix, homeodomain, WRKY and LIM transcription factors in the high WBD plants. All these results indicate that the high WBD in plants has been associated with the increased transcription of many genes related to lignocellulose formation., Conclusions: Most lignocellulose biosynthesis related genes exhibited a tendency to transcribe at relatively higher level in high WBD plants. These results suggest that lignocellulose biosynthesis-related genes may be associated with WBD.

Despite a wealth of eco-physiological assessments of plant response to extreme drought, few studies have addressed the interactive effects of global change factors on traits driving mortality. To understand the interaction between hydraulic and carbon metabolic traits influencing tree mortality, which may be independently influenced by atmospheric [CO2] and temperature, we grew Eucalyptus sideroxylon A. Cunn. ex Woolls from seed in a full-factorial [CO2] (280, 400 and 640 μmol mol-1, Cp, Ca and Ce, respectively) and temperature (ambient and ambient +4 °C, Ta and Te, respectively) experiment. Prior to drought, growth across treatment combinations resulted in significant variation in physiological and morphological traits, including photosynthesis (Asat), respiration (Rd), stomatal conductance, carbohydrate storage, biomass and leaf area (LA). Ce increased Asat, LA and leaf carbohydrate concentration compared with Ca, while Cp generated the opposite response; Te reduced Rd. However, upon imposition of drought, Te hastened mortality (9 days sooner compared with Ta), while Ce significantly exacerbated drought stress when combined with Te. Across treatments, earlier time-to-mortality was mainly associated with lower (more negative) leaf water potential (Ψl) during the initial drought phase, along with higher water loss across the first 3 weeks of water limitation. Among many variables, Ψl was more important than carbon status in predicting time-to-mortality across treatments, yet leaf starch was associated with residual variation within treatments. These results highlight the need to carefully consider the integration, interaction and hierarchy of traits contributing to mortality, along with their responses to environmental drivers. Both morphological traits, which influence soil resource extraction, and physiological traits, which affect water-for-carbon exchange to the atmosphere, must be considered to adequately predict plant response to drought. Researchers have struggled with assessing the relative importance of hydraulic and carbon metabolic traits in determining mortality, yet an integrated trait, time-dependent framework provides considerable insight into the risk of death from drought for trees.

Background and Aims: Many studies exist in the literature dealing with mathematical representations of root systems, categorized, for example, as pure structure description, partial derivative equations or functional-structural plant models. However, in these studies, root architecture modelling has seldom been carried out at the organ level with the inclusion of environmental influences that can be integrated into a whole plant characterization., Methods: We have conducted a multidisciplinary study on root systems including field observations, architectural analysis, and formal and mathematical modelling. This integrative and coherent approach leads to a generic model (DigR) and its software simulator. Architecture analysis applied to root systems helps at root type classification and architectural unit design for each species. Roots belonging to a particular type share dynamic and morphological characteristics which consist of topological and geometric features. The DigR simulator is integrated into the Xplo environment, with a user interface to input parameter values and make output ready for dynamic 3-D visualization, statistical analysis and saving to standard formats. DigR is simulated in a quasi-parallel computing algorithm and may be used either as a standalone tool or integrated into other simulation platforms. The software is open-source and free to download at http://amapstudio.cirad.fr/soft/xplo/download., Key Results: DigR is based on three key points: (1) a root-system architectural analysis, (2) root type classification and modelling and (3) a restricted set of 23 root type parameters with flexible values indexed in terms of root position. Genericity and botanical accuracy of the model is demonstrated for growth, branching, mortality and reiteration processes, and for different root architectures. Plugin examples demonstrate the model's versatility at simulating plastic responses to environmental constraints. Outputs of the model include diverse root system structures such as tap-root, fasciculate, tuberous, nodulated and clustered root systems., Conclusions: DigR is based on plant architecture analysis which leads to specific root type classification and organization that are directly linked to field measurements. The open source simulator of the model has been included within a friendly user environment. DigR accuracy and versatility are demonstrated for growth simulations of complex root systems for both annual and perennial plants.

This study focuses on the effects of different thinning regimes on clonal Eucalyptus plantations growth. Four different trials, planted in 1999 and located in Bahia and Espírito Santo States, were used. Aside from thinning, initial planting density, and post thinning fertilization application were also evaluated. Before canopy closure, and therefore before excessive competition between trees took place, it was found that stands planted under low densities (667 trees per hectare) presented a lower mortality proportion when compared to stand planted under higher densities (1111 trees per hectare). However, diameter growth prior to thinning operations was not statistically different between these two densities, presenting an overall mean of 4.9 cm/year. After canopy closure and the application of the thinning treatments, it was found that thinning regimes beginning early in the life of the stand and leaving a low number of residual trees presented the highest diameter and height growth. Unthinned treatments and thinning regimes late in the life of the stand (after 5.5 years), leaving a large number of residual trees presented the highest values of basal area production. The choice of the best thinning regime for Eucalyptus clonal material will vary according to the plantation objective.

Wood density can be considered as a measure of the internal wood structure, and it is usually used as a proxy measure of other mechanical and functional traits. Eucalyptus is one of the most important commercial forestry genera worldwide, but the relationship between wood density and vulnerability to cavitation in this genus has been little studied. The analysis is hampered by, among other things, its anatomical complexity, so it becomes necessary to address more complex techniques and analyses to elucidate the way in which the different anatomical elements are functionally integrated. In this study, vulnerability to cavitation in two races of Eucalyptus globulus Labill. with different wood density was evaluated through Path analysis, a multivariate method that allows evaluation of descriptive models of causal relationship between variables. A model relating anatomical variables with wood properties and functional parameters was proposed and tested. We found significant differences in wood basic density and vulnerability to cavitation between races. The main exogenous variables predicting vulnerability to cavitation were vessel hydraulic diameter and fibre wall fraction. Fibre wall fraction showed a direct impact on wood basic density and the slope of vulnerability curve, and an indirect and negative effect over the pressure imposing 50% of conductivity loss (P50) through them. Hydraulic diameter showed a direct negative effect on P50, but an indirect and positive influence over this variable through wood density on one hand, and through maximum hydraulic conductivity (ks max) and slope on the other. Our results highlight the complexity of the relationship between xylem efficiency and safety in species with solitary vessels such as Eucalyptus spp., with no evident compromise at the intraspecific level., (© The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background and Aims: Sapwood traits like vessel diameter and intervessel pit characteristics play key roles in maintaining hydraulic integrity of trees. Surprisingly little is known about how sapwood traits covary with tree height and how such trait-based variation could affect the efficiency of water transport in tall trees. This study presents a detailed analysis of structural and functional traits along the vertical axes of tall Eucalyptus grandis trees., Methods: To assess a wide range of anatomical and physiological traits, light and electron microscopy was used, as well as field measurements of tree architecture, water use, stem water potential and leaf area distribution., Key Results: Strong apical dominance of water transport resulted in increased volumetric water supply per unit leaf area with tree height. This was realized by continued narrowing (from 250 to 20 µm) and an exponential increase in frequency (from 600 to 13 000 cm-2) of vessels towards the apex. The widest vessels were detected at least 4 m above the stem base, where they were associated with the thickest intervessel pit membranes. In addition, this study established the lower limit of pit membrane thickness in tall E. grandis at ~375 nm. This minimum thickness was maintained over a large distance in the upper stem, where vessel diameters continued to narrow., Conclusions: The analyses of xylem ultrastructure revealed complex, synchronized trait covariation and trade-offs with increasing height in E. grandis. Anatomical traits related to xylem vessels and those related to architecture of pit membranes were found to increase efficiency and apical dominance of water transport. This study underlines the importance of studying tree hydraulic functioning at organismal scale. Results presented here will improve understanding height-dependent structure-function patterns in tall trees., (© The Author(s) 2018. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

This study evaluated the quality of heartwood and sapwood from mature trees of three species of Eucalyptus, by means of the qualification of their proportion, determination of basic and apparent density using non-destructive attenuation of gamma radiation technique and calculation of the density uniformity index. Six trees of each species (Eucalyptus grandis - 18 years old, Eucalyptus tereticornis - 35 years old and Corymbia citriodora - 28 years old) were used in the experimental program. The heartwood and sapwood were delimited by macroscopic analysis and the calculation of areas and percentage of heartwood and sapwood were performed using digital image. The uniformity index was calculated following methodology which numerically quantifies the dispersion of punctual density values of the wood around the mean density along the radius. The percentage of the heartwood was higher than the sapwood in all species studied. The density results showed no statistical difference between heartwood and sapwood. Differently from the density results, in all species studied there was statistical differences between uniformity indexes for heartwood and sapwood regions, making justifiable the inclusion of the density uniformity index as a quality parameter for Eucalyptus wood.

This study focuses on the effects of different thinning regimes on clonal Eucalyptus plantations growth. Four different trials, planted in 1999 and located in Bahia and Espírito Santo States, were used. Aside from thinning, initial planting density, and post thinning fertilization application were also evaluated. Before canopy closure, and therefore before excessive competition between trees took place, it was found that stands planted under low densities (667 trees per hectare) presented a lower mortality proportion when compared to stand planted under higher densities (1111 trees per hectare). However, diameter growth prior to thinning operations was not statistically different between these two densities, presenting an overall mean of 4.9 cm/year. After canopy closure and the application of the thinning treatments, it was found that thinning regimes beginning early in the life of the stand and leaving a low number of residual trees presented the highest diameter and height growth. Unthinned treatments and thinning regimes late in the life of the stand (after 5.5 years), leaving a large number of residual trees presented the highest values of basal area production. The choice of the best thinning regime for Eucalyptus clonal material will vary according to the plantation objective.

Dengue, chikungunya, and yellow fever are important vector-borne diseases transmitted by female mosquitoes when they feed on humans. The use of repellents based on natural products is an alternative for personal protection against these diseases. Application of chemicals with larvicidal activity is another strategy for controlling the mosquito population. The repellent and larvicidal activities of the essential oil from Eucalyptus nitens were tested against Aedes aegypti and Aedes albopictus, the main vectors of these arboviruses. The essential oil was extracted by hydrodistillation and then analyzed by gas chromatography-mass spectrometry. The main components of Eucalyptus nitens essential oil were found to be terpenes such as 1,8-cineole and p-cymene, followed by β-triketones and alkyl esters. The repellent activity of the essential oil against both species was significantly higher when compared with the main component, 1,8-cineole, alone. These results indicate that the repellent effect of E. nitens is not due only to the main component, 1,8-cineole, but also that other compounds may be responsible. Aedes aegypti was found to be more tolerant to the essential oil larvicidal effects than Ae. albopictus (Ae. aegypti LC50 = 52.83 ppm, Ae. albopictus LC 50 = 28.19 ppm). The repellent and larvicidal activity could be associated to the presence of cyclic β-triketones such as flavesone, leptospermone, and isoleptospermone., (© The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Reticulate evolution by hybridization is considered a common process shaping the evolution of many plant species, however, reticulation could also be due to incomplete lineage sorting in biodiverse systems. For our study we selected a group of closely related plant taxa with contrasting yet partially overlapping geographic distributions and different population sizes, to distinguish between reticulated patterns due to hybridization and incomplete lineage sorting. We predicted that sympatric or proximal populations of different species are more likely to have gene flow than geographically distant populations of the same widespread species. Furthermore, for species with restricted distributions, and therefore, small effective population sizes, we predicted complete lineage sorting. Eastern grey box eucalypt species (Eucalyptus supraspecies Moluccanae) provide an ideal system to explore patterns of reticulate evolution. They form a diverse, recently evolved and phylogenetically undefined group within Eucalyptus, with overlapping morphological features and hybridization in nature. We used a multi-faceted approach, combining analyses of chloroplast and nuclear DNA, as well as seedling morphology, flowering time and ecological spatial differentiation in order to test for species delimitation and reticulate evolution in this group. The multiple layers of results were consistent and suggested a lack of monophyly at different hierarchical levels due to multidirectional gene flow among several species, challenging species delimitation. Chloroplast and nuclear haplotypes were shared among different species in geographic proximity, consistent with hybridization zones. Furthermore, species with restricted distributions appeared better resolved due to lineage sorting in the absence of hybridization. We conclude that a combination of molecular, morphological and ecological approaches is required to disentangle patterns of reticulate evolution in the box eucalypts., (Published by Elsevier Inc.)

Leaf veins supply the mesophyll with water that evaporates when stomata are open to allow CO 2 uptake for photosynthesis. Theoretical analyses suggest that water is optimally distributed in the mesophyll when the lateral distance between veins (d x ) is equal to the distance from these veins to the epidermis (d y ), expressed as d x :d y ≈ 1. Although this theory is supported by observations of many derived angiosperms, we hypothesize that plants in arid environments may reduce d x :d y below unity owing to climate-specific functional adaptations of increased leaf thickness and increased vein density. To test our hypothesis, we assembled leaf hydraulic, morphological, and photosynthetic traits of 68 species from the Eucalyptus and Corymbia genera (termed eucalypts) along an aridity gradient in southwestern Australia. We inferred the potential gas-exchange advantage of reducing d x beyond d y using a model that links leaf morphology and hydraulics to photosynthesis. Our observations reveal that eucalypts in arid environments have thick amphistomatous leaves with high vein densities, resulting in d x :d y ratios that range from 1.6 to 0.15 along the aridity gradient. Our model suggests that, as leaves become thicker, the effect of reducing d x beyond d y is to offset the reduction in leaf gas exchange that would result from maintaining d x :d y at unity. This apparent overinvestment in leaf venation may be explained from the selective pressure of aridity, under which traits associated with long leaf life span, high hydraulic and thermal capacitances, and high potential rates of leaf water transport confer a competitive advantage., (© 2016 American Society of Plant Biologists. All Rights Reserved.)

Premise of the Study: Plants are routinely subjected to multiple environmental stressors, and the ability to respond to these stressors determines species survival and ecological breadth. Despite stressors such as wind and dust significantly influencing plant development, morphology, and chemistry, the combined influence of these factors is yet to be investigated., Methods: We used a manipulative glasshouse approach to compare the morphological, physiological, and biomechanical responses of Eucalyptus tereticornis to the independent and combined effects of wind and dust., Key Results: Wind decreased both E. tereticornis height and stem flexural stiffness. Additionally, wind had no effect on leaf physiology, nor did dust have any significant effect on any of the traits measured., Conclusions: Our results suggest that wind and dust in combination may have an additive effect on several plant traits and provide new insight into the effects and importance of studying wind, dust, and different stress combinations., (© 2016 Botanical Society of America.)

It is widely accepted that substantial nighttime sap flux (J s,n) or transpiration (E) occurs in most plants, but the physiological implications are poorly known. It has been hypothesized that J s,n or E serves to enhance nitrogen uptake or deliver oxygen; however, no clear evidence is currently available. In this study, sap flux (J s) in Eucalyptus grandis × urophylla with apparent stem photosynthesis was measured, including control trees which were covered by aluminum foil (approximately 1/3 of tree height) to block stem photosynthesis. We hypothesized that the nighttime water flux would be suppressed in trees with lower stem photosynthesis. The results showed that the green tissue degraded after 3 months, demonstrating a decrease in stem photosynthesis. The daytime J s decreased by 21.47%, while J s,n decreased by 12.03% in covered trees as compared to that of control, and the difference was statistically significant (P < 0.01). The linear quantile regression model showed that J s,n decreased for a given daytime transpiration water loss, indicating that J s,n was suppressed by lower stem photosynthesis in covered trees. Predawn (ψ pd) of covered trees was marginally higher than that of control while lower at predawn stomatal conductance (g s), indicating a suppressed water flux in covered trees. There was no difference in leaf carbon content and δ(13)C between the two groups, while leaf nitrogen content and δ(15)N were significantly higher in covered trees than that of the control (P < 0.05), indicating that J s,n was not used for nitrogen uptake. These results suggest that J s,n may act as an oxygen pathway since green tissue has a higher respiration or oxygen demand than non-green tissue. Thus, this study demonstrated the physiological implications of J s,n and the possible benefits of nighttime water use or E by the tree.

The sub-dermal secretory cavities (glands) embedded within the leaves of Eucalyptus (Myrtaceae) were once thought to be the exclusive repositories of monoterpene and sesquiterpene oils. Recent research has debunked this theory and shown that abundant non-volatile compounds also occur within foliar glands. In particular, glands of four species in subgenus Eucalyptus contain the biologically active flavanone pinocembrin. Pinocembrin shows great promise as a pharmaceutical and is predominantly plant-sourced, so Eucalyptus could be a potential commercial source of such compounds. To explore this we quantified and assessed the purity of pinocembrin in glands of 11 species of E. subg. Eucalyptus using Electro-Spray Ionisation Liquid Chromatography Mass Spectrometry of acetonitrile extracts and Gas Chromatography Mass Spectrometry analyses of hexane extracts of isolated glands which were free from other leaf tissues. Our results showed that the glands of subgenus Eucalyptus contain numerous flavanones that are structurally related to pinocembrin and often present in much greater abundance. The maximum concentration of pinocembrin was 2 mg g-1 dry leaf found in E. stellulata, whereas that of dimethylpinocembrin (5,7-dimethoxyflavanone) was 10 mg g-1 in E. oreades and that of pinostrobin (5-hydroxy-7-methoxyflavanone) was 12 mg g-1 in E. nitida. We also found that the flavanones are exclusively located within the foliar glands rather than distributed throughout leaf tissues. The flavanones differ from the non-methylated pinocembrin in the degree and positions of methylation. This finding is particularly important given the attractiveness of methylated flavonoids as pharmaceuticals and therapeutics. Another important finding was that glands of some members of the subgenus also contain flavanone O-glucosides and flavanone-β-triketone conjugates. In addition, glands contain free β-triketones, β-triketone heterodimers and chromone C-glucosides. Therefore, the foliar glands of this taxonomically distinct group of plants are a rich source of a range of flavonoids and other biologically active compounds with great commercial potential.

Current theory presumes that natural selection on vascular traits is controlled by a trade-off between efficiency and safety of hydraulic architecture. Hence, traits linked to efficiency, such as vessel diameter, should show biogeographic patterns; but critical tests of these predictions are rare, largely owing to confounding effects of environment, tree size and phylogeny. Using wood sampled from a phylogenetically constrained set of 28 Eucalyptus species, collected from a wide gradient of aridity across Australia, we show that hydraulic architecture reflects adaptive radiation of this genus in response to variation in climate. With increasing aridity, vessel diameters narrow, their frequency increases with a distribution that becomes gradually positively skewed and sapwood density increases while the theoretical hydraulic conductivity declines. Differences in these hydraulic traits appear largely genotypic in origin rather than environmentally plastic. Data reported here reflect long-term adaptation of hydraulic architecture to water availability. Rapidly changing climates, on the other hand, present significant challenges to the ability of eucalypts to adapt their vasculature., (© 2016 John Wiley & Sons Ltd/CNRS.)

Water stress modifies plant above- vs belowground biomass allocation, i.e., morphological plasticity. It is known that all species and genotypes reduce their growth rate in response to stress, but in the case of water stress it is unclear whether the magnitude of such reduction is linked to the genotype's growth potential, and whether the reduction can be largely attributed to morphological adjustments such as plant allocation and leaf and root anatomy. We subjected seedlings of six seed sources, three from each of Eucalyptus camaldulensis (potentially fast growing) and E. globulus (inherently slow growing), to three experimental water regimes. Biomass, leaf area and root length were measured in a 6-month glasshouse experiment. We then performed functional growth analysis of relative growth rate (RGR), and aboveground (leaf area ratio (LAR), specific leaf area (SLA) and leaf mass ratio (LMR)) and belowground (root length ratio (RLR), specific root length (SRL) and root mass ratio (RMR)) morphological components. Total biomass, root biomass and leaf area were reduced for all Eucalyptus provenances according to drought intensity. All populations exhibited drought plasticity, while those of greater growth potential (RGRmax) had a larger reduction in growth (discounting the effect of size). A positive correlation was observed between drought sensitivity and RGRmax. Aboveground, drought reduced LAR and LMR; under severe drought a negative correlation was found between LMR and RGRmax. Belowground, drought reduced SRL but increased RMR, resulting in no change in RLR. Under severe drought, a negative correlation was found between RLR, SRL and RGRmax. Our evidence strongly supports the classic ecophysiological trade-off between growth potential and drought tolerance for woody seedlings. It also suggests that slow growers would have a low capacity to adjust their morphology. For shoots, this constraint on plasticity was best observed in partition (i.e., LMR) whereas for roots it was clearest in morphology/anatomy (i.e., SRL). Thus, a low RGRmax would limit plastic response to drought not only at the whole plant level but also at the organ and even the tissue level., (© The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Eucalyptus trees are among the most important species for industrial forestry worldwide. However, as with most forest trees, flowering does not begin for one to several years after planting which can limit the rate of conventional and molecular breeding. To speed flowering, we transformed a Eucalyptus grandis × urophylla hybrid (SP7) with a variety of constructs that enable overexpression of FLOWERING LOCUS T (FT). We found that FT expression led to very early flowering, with events showing floral buds within 1-5 months of transplanting to the glasshouse. The most rapid flowering was observed when the cauliflower mosaic virus 35S promoter was used to drive the Arabidopsis thaliana FT gene (AtFT). Early flowering was also observed with AtFT overexpression from a 409S ubiquitin promoter and under heat induction conditions with Populus trichocarpa FT1 (PtFT1) under control of a heat-shock promoter. Early flowering trees grew robustly, but exhibited a highly branched phenotype compared to the strong apical dominance of nonflowering transgenic and control trees. AtFT-induced flowers were morphologically normal and produced viable pollen grains and viable self- and cross-pollinated seeds. Many self-seedlings inherited AtFT and flowered early. FT overexpression-induced flowering in Eucalyptus may be a valuable means for accelerating breeding and genetic studies as the transgene can be easily segregated away in progeny, restoring normal growth and form., (© 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.)

Background and Aims: Experimental drought is well documented to induce a decline in photosynthetic capacity. However, if given time to acclimate to low water availability, the photosynthetic responses of plants to low soil moisture content may differ from those found in short-term experiments. This study aims to test whether plants acclimate to long-term water stress by modifying the functional relationships between photosynthetic traits and water stress, and whether species of contrasting habitat differ in their degree of acclimation., Methods: Three Eucalyptus taxa from xeric and riparian habitats were compared with regard to their gas exchange responses under short- and long-term drought. Photosynthetic parameters were measured after 2 and 4 months of watering treatments, namely field capacity or partial drought. At 4 months, all plants were watered to field capacity, then watering was stopped. Further measurements were made during the subsequent 'drying-down', continuing until stomata were closed., Key Results: Two months of partial drought consistently reduced assimilation rate, stomatal sensitivity parameters (g1), apparent maximum Rubisco activity (V'(cmax)) and maximum electron transport rate (J'(max)). Eucalyptus occidentalis from the xeric habitat showed the smallest decline in V'(cmax) and J'(max); however, after 4 months, V'(cmax) and J'(max) had recovered. Species differed in their degree of V'(cmax) acclimation. Eucalyptus occidentalis showed significant acclimation of the pre-dawn leaf water potential at which the V'(cmax) and 'true' V(cmax) (accounting for mesophyll conductance) declined most steeply during drying-down., Conclusions: The findings indicate carbon loss under prolonged drought could be over-estimated without accounting for acclimation. In particular, (1) species from contrasting habitats differed in the magnitude of V'(cmax) reduction in short-term drought; (2) long-term drought allowed the possibility of acclimation, such that V'(cmax) reduction was mitigated; (3) xeric species showed a greater degree of V'(cmax) acclimation; and (4) photosynthetic acclimation involves hydraulic adjustments to reduce water loss while maintaining photosynthesis., (© The Author 2015. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Adventitious rooting (AR) is essential in clonal propagation. Eucalyptus globulus is relevant for the cellulose industry due to its low lignin content. However, several useful clones are recalcitrant to AR, often requiring exogenous auxin, adding cost to clonal garden operations. In contrast, E. grandis is an easy-to-root species widely used in clonal forestry. Aiming at contributing to the elucidation of recalcitrance causes in E. globulus, we conducted a comparative analysis with these two species differing in rooting competence, combining gene expression and anatomical techniques. Recalcitrance in E. globulus is reversed by exposure to exogenous indole-3-acetic acid (IAA), which promotes important gene expression modifications in both species. The endogenous content of IAA was significantly higher in E. grandis than in E. globulus. The cambium zone was identified as an active area during AR, concentrating the first cell divisions. Immunolocalization assay showed auxin accumulation in cambium cells, further indicating the importance of this region for rooting. We then performed a cambium zone-specific gene expression analysis during AR using laser microdissection. The results indicated that the auxin-related genes TOPLESS and IAA12/BODENLOS and the cytokinin-related gene ARR1may act as negative regulators of AR, possibly contributing to the hard-to-root phenotype of E. globulus., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Tree invasions have substantial impacts on biodiversity and ecosystem functioning, and trees that are dispersed by animals are more likely to become invasive. In addition, hybridisation between plants is well documented as a source of new weeds, as hybrids gain new characteristics that allow them to become invasive. Corymbia torelliana is an invasive tree with an unusual animal dispersal mechanism: seed dispersal by stingless bees, that hybridizes readily with other species. We examined hybrids between C. torelliana and C. citriodora subsp. citriodora to determine whether hybrids have inherited the seed dispersal characteristics of C. torelliana that allow bee dispersal. Some hybrid fruits displayed the characteristic hollowness, resin production and resin chemistry associated with seed dispersal by bees. However, we did not observe bees foraging on any hybrid fruits until they had been damaged. We conclude that C. torelliana and C. citriodora subsp. citriodora hybrids can inherit some fruit characters that are associated with dispersal by bees, but we did not find a hybrid with the complete set of characters that would enable bee dispersal. However, around 20,000 hybrids have been planted in Australia, and ongoing monitoring is necessary to identify any hybrids that may become invasive.

The cerambycid beetle, Phoracantha semipunctata F., was introduced into California in the mid-1980s and killed large numbers of Eucalyptus host trees. The populations of the borer declined to very low levels in the mid-1990s following the establishment of the congener, Phoracantha recurva Newman, and the intentional introduction of the egg parasitoid, Avetianella longoi Siscaro. The distributions of the beetles overlap in the Australian native range, but one species has replaced the other in the adventive range in California. One possible explanation is differential susceptibility to natural enemies introduced for biological control. An alternative explanation for the reduced abundance of P. semipunctata is asymmetric interspecific competition between the two species. To test this hypothesis, equal larval densities of each species were introduced into host logs. In all cases, more P. recurva adults emerged than P. semipunctata adults, but the presence of congeners did not have a different effect than the presence of an equal density of conspecific individuals. Neither the temporal order of introduction or bark thickness altered the outcome of potential competitive interactions. Consequently, it appears that the ecological replacement of one borer with another in the adventive environment in southern California may not be a result of bottom-up intraguild competitive interactions. The top-down effects of natural enemies on P. semipunctata have most likely led to its decline., (© The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Recent advances in modelling the architecture and function of the plant hydraulic network have led to improvements in predicting and interpreting the consequences of functional trait variation on CO2 uptake and water loss. We build upon one such model to make novel predictions for scaling of the total specific hydraulic conductance of leaves and shoots (kL and kSH , respectively) and variation in the partitioning of hydraulic conductance. Consistent with theory, we observed isometric (slope = 1) scaling between kL and kSH across several independently collected datasets and a lower ratio of kL and kSH , termed the leaf-to-shoot conductance ratio (CLSCR ), in arid environments and in woody species. Isometric scaling of kL and kSH supports the concept that hydraulic design is coordinated across the plant. We propose that CLSCR is an important adaptive trait that represents the trade-off between efficiency and safety at the scale of the whole plant., (© 2015 John Wiley & Sons Ltd.)

Introgressive hybridization is increasingly recognized as having influenced the gene pools of large genera of plants, yet it is rarely invoked as an explanation for why closely related plant species do not co-occur. Here, we asked how the environment and tendency to interbreed relate to neighborhood co-occurrence patterns for Eucalyptus species in the Grampians National Park, Victoria, Australia. We identified species pairs that have experienced ongoing hybridization and introgression on the basis of the extent of incongruence between chloroplast DNA (JLA+ region) and nuclear ribosomal DNA (internal transcribed spacer region) phylogenies, geographic patterns of gene sharing, and field observation of intermediate morphologies. Co-occurrence, trait data (specific leaf area [SLA], maximum height, and seed mass), and environmental data were measured in plots distributed along environmental gradients. Trait and habitat similarity influenced species co-occurrence the most overall (e.g., co-occurring species had similar SLA). Reproductively compatible species were an exception; they rarely co-occurred despite being functionally similar. The negative effect of reproductive compatibility was stronger than the positive effect of SLA on co-occurrence. Our results emphasize the dominant roles of the environment and the importance of evolution in structuring local assemblages. We argue that the mechanism responsible for preventing closely related species from co-occurring in this system is reproductive interference rather than competitive exclusion. Reproductive interference should be considered more generally as a potential cause of phylogenetic overdispersion.

Tension wood has distinct physical and chemical properties, including altered fibre properties, cell wall composition and ultrastructure. It serves as a good system for investigating the genetic regulation of secondary cell wall biosynthesis and wood formation. The reference genome sequence for Eucalyptus grandis allows investigation of the global transcriptional reprogramming that accompanies tension wood formation in this global wood fibre crop. We report the first comprehensive analysis of physicochemical wood property changes in tension wood of Eucalyptus measured in a hybrid (E. grandis × Eucalyptus urophylla) clone, as well as genome-wide gene expression changes in xylem tissues 3 wk post-induction using RNA sequencing. We found that Eucalyptus tension wood in field-grown trees is characterized by an increase in cellulose, a reduction in lignin, xylose and mannose, and a marked increase in galactose. Gene expression profiling in tension wood-forming tissue showed corresponding down-regulation of monolignol biosynthetic genes, and differential expression of several carbohydrate active enzymes. We conclude that alterations of cell wall traits induced by tension wood formation in Eucalyptus are a consequence of a combination of down-regulation of lignin biosynthesis and hemicellulose remodelling, rather than the often proposed up-regulation of the cellulose biosynthetic pathway., (© 2014 University of Pretoria New Phytologist © 2014 New Phytologist Trust.)

We investigated physiological and morphological responses to flooding and recovery in Eucalyptus camaldulensis subsp. refulgens, a riparian tree species from a dryland region prone to intense episodic floods. Seedlings in soil flooded for 88 d produced extensive adventitious roots, displayed stem hypertrophy (stem diameter increased by 93%) and increased root porosity owing to aerenchyma formation. Net photosynthesis (Pn) and stomatal conductance (gs) were maintained for at least 2 weeks of soil flooding, contrasting with previous studies of other subspecies of E. camaldulensis. Gradual declines followed in both gs (30% less than controls) and Pn (19% less). Total leaf soluble sugars did not differ between flooded and control plants. Root mass did not recover 32 d after flooding ceased, but gs was not lower than controls, suggesting the root system was able to functionally compensate. However, the limited root growth during recovery after flooding was surprising given the importance of extensive root systems in dryland environments. We conclude that early flood tolerance could be an adaptation to capitalize on scarce water resources in a water-limited environment. Overall, our findings highlight the need to assess flooding responses in relation to a species' fitness for particular flood regimes or ecological niches., (© 2014 John Wiley & Sons Ltd.)

Understanding the regulation of water use at the whole-tree scale is critical to advancing the utility of physiological ecology, for example in its role in predictive hydrology of forested catchments. For three eucalypt species that dominate high-elevation catchments in south-eastern Australia, we examined if whole-tree water use could be related to three widely discussed regulators of water use: stomatal anatomy, sensitivity of stomata [i.e. stomatal conductance (g(s))] to environmental influences, and sapwood area. While daily tree water use varied sixfold among species, sap velocity and sapwood area varied in parallel. Combined, stomatal structure and physiology could not explain differences in species-specific water use. Species which exhibited the fastest (Eucalyptus delegatensis) and slowest (Eucalyptus pauciflora) rates of water use both exhibited greater capacity for physiological control of g(s) [indicated by sensitivity to vapour pressure deficit (VPD)] and a reduced capacity to limit g(s) anatomically [indicated by greater potential g(s) (g(max))]. Conversely, g(s) was insensitive to VPD and g(max) was lowest for Eucalyptus radiata, the species showing intermediate rates of water use. Improved knowledge of stomatal anatomy will help us to understand the capacity of species to regulate leaf-level water loss, but seems likely to remain of limited use for explaining rates of whole-tree water use in montane eucalypts at the catchment scale.

The transfer of water from phloem into xylem is thought to mitigate increasing hydraulic tension in the vascular system of trees during the diel cycle of transpiration. Although a putative plant function, to date there is no direct evidence of such water transfer or the contributing pathways. Here, we trace the radial flow of water from the phloem into the xylem and investigate its diel variation. Introducing a fluorescent dye (0.1% [w/w] fluorescein) into the phloem water of the tree species Eucalyptus saligna allowed localization of the dye in phloem and xylem tissues using confocal laser scanning microscopy. Our results show that the majority of water transferred between the two tissues is facilitated via the symplast of horizontal ray parenchyma cells. The method also permitted assessment of the radial transfer of water during the diel cycle, where changes in water potential gradients between phloem and xylem determine the extent and direction of radial transfer. When injected during the morning, when xylem water potential rapidly declined, fluorescein was translocated, on average, farther into mature xylem (447 ± 188 µm) compared with nighttime, when xylem water potential was close to zero (155 ± 42 µm). These findings provide empirical evidence to support theoretical predictions of the role of phloem-xylem water transfer in the hydraulic functioning of plants. This method enables investigation of the role of phloem tissue as a dynamic capacitor for water storage and transfer and its contribution toward the maintenance of the functional integrity of xylem in trees., (© 2015 American Society of Plant Biologists. All Rights Reserved.)

These past years have seen an enormous development of the area of natural antioxidants and antimicrobials. Eucalyptus globulus is widely cultivated in subtropical and Mediterranean regions in intensive short rotation coppice plantations. In the Portuguese context, E. globulus is the third species in terms of forest area. The stump is the basal part of the tree, including the near-the-ground stem portion and the woody roots that remain after stem felling. The purpose of this work was to study the phytochemical profile and to evaluate the antioxidant and antimicrobial properties of several crude stump wood and stump bark extracts of E. globulus, comparing it with similar extracts of E. globulus wood (industrial chips). The results showed the presence of high concentrations of total phenolic compounds (>200 mg GAE/g extract) and flavonoids (>10 mg QE/g extract) in E. globulus stump extracts. Generally the stump wood extracts stands out from the other ones, presenting the highest percentages of inhibition of linoleic acid oxidation. It was also possible to conclude that the extracts were more active against Gram-positive bacteria, presenting low MIC values. This study thus provides information supporting the economic valorization of E. globulus stump wood.

Large trees are critical components of forest ecosystems, but are declining in many forests worldwide. We predicted that growth of large trees is more vulnerable than that of small trees to high temperatures, because respiration and tissue maintenance costs increase with temperature more rapidly than does photosynthesis and these costs may be disproportionately greater in large trees. Using 5 00 000 measurements of eucalypt growth across temperate Australia, we found that high temperatures do appear to impose a larger growth penalty on large trees than on small ones. Average stem diameter growth rates at 21 °C compared with 11 °C mean annual temperature were 57% lower for large trees (58 cm stem diameter), but only 29% lower for small trees (18 cm diameter). While our results are consistent with an impaired carbon budget for large trees at warmer sites, we cannot discount causes such as hydraulic stress. We conclude that slower growth rates will impede recovery from extreme events, exacerbating the effects of higher temperatures, increased drought stress and more frequent fire on the tall eucalypt forests of southern Australia., (© 2014 John Wiley & Sons Ltd.)

Widespread species often occur across a range of climatic conditions, through a combination of local genetic adaptations and phenotypic plasticity. Species with greater phenotypic plasticity are likely to be better positioned to cope with rapid anthropogenic climate changes, while those displaying strong local adaptations might benefit from translocations to assist the movement of adaptive genes as the climate changes. Eucalyptus tricarpa occurs across a climatic gradient in south-eastern Australia, a region of increasing aridity, and we hypothesized that this species would display local adaptation to climate. We measured morphological and physiological traits reflecting climate responses in nine provenances from sites of 460 to 1040 mm annual rainfall, in their natural habitat and in common gardens near each end of the gradient. Local adaptation was evident in functional traits and differential growth rates in the common gardens. Some traits displayed complex combinations of plasticity and genetic divergence among provenances, including clinal variation in plasticity itself. Provenances from drier locations were more plastic in leaf thickness, whereas leaf size was more plastic in provenances from higher rainfall locations. Leaf density and stomatal physiology (as indicated by δ(13)C and δ(18)O) were highly and uniformly plastic. In addition to variation in mean trait values, genetic variation in trait plasticity may play a role in climate adaptation., (© 2013 John Wiley & Sons Ltd.)

Following disturbance many woody species are capable of resprouting new foliage, resulting in a reduced leaf-to-sapwood area ratio and altered canopy structure. We hypothesized that such changes would promote adjustments in leaf physiology, resulting in higher rates of transpiration per unit leaf area, consistent with the mechanistic framework proposed by Whitehead et al. (Whitehead D, Jarvis PG, Waring RH (1984) Stomatal conductance, transpiration and resistance to water uptake in a Pinus sylvestris spacing experiment. Can J For Res 14:692-700). We tested this in Eucalyptus obliqua L'Hér following a wildfire by comparing trees with unburnt canopies with trees that had been subject to 100% canopy scorch and were recovering their leaf area via resprouting. In resprouting trees, foliage was distributed along the trunk and on lateral branches, resulting in shorter hydraulic path lengths. We evaluated measurements of whole-tree transpiration and structural and physiological traits expected to drive any changes in transpiration. We used these structural and physiological measurements to parameterize the Whitehead et al. equation, and found that the expected ratio of transpiration per unit leaf area between resprouting and unburnt trees was 3.41. This is similar to the observed ratio of transpiration per unit leaf area, measured from sapflow observations, which was 2.89 (i.e., resprouting trees had 188% higher transpiration per unit leaf area). Foliage at low heights (<2 m) was found to be significantly different to foliage in the tree crown (14-18 m) in a number of traits, including higher specific leaf area, midday leaf water potential and higher rates of stomatal conductance and photosynthesis. We conclude that these post-fire adjustments in resprouting trees help to drive increased stomatal conductance and hydraulic efficiency, promoting the rapid return of tree-scale transpiration towards pre-disturbance levels. These transient patterns in canopy transpiration have important implications for modelling stand-level water fluxes in forests capable of resprouting, which is frequently done on the basis of the leaf area index.

Although vast areas in tropical regions have weathered soils with low potassium (K) levels, little is known about the effects of K supply on the photosynthetic physiology of trees. This study assessed the effects of K and sodium (Na) supply on the diffusional and biochemical limitations to photosynthesis in Eucalyptus grandis leaves. A field experiment comparing treatments receiving K (+K) or Na (+Na) with a control treatment (C) was set up in a K-deficient soil. The net CO2 assimilation rates were twice as high in +K and 1.6 times higher in +Na than in the C as a result of lower stomatal and mesophyll resistance to CO2 diffusion and higher photosynthetic capacity. The starch content was higher and soluble sugar was lower in +K than in C and +Na, suggesting that K starvation disturbed carbon storage and transport. The specific leaf area, leaf thickness, parenchyma thickness, stomatal size and intercellular air spaces increased in +K and +Na compared to C. Nitrogen and chlorophyll concentrations were also higher in +K and +Na than in C. These results suggest a strong relationship between the K and Na supply to E. grandis trees and the functional and structural limitations to CO2 assimilation rates., (© 2013 John Wiley & Sons Ltd.)

Models of stomatal conductance (g(s)) are based on coupling between g(s) and CO(2) assimilation (A(net)), and it is often assumed that the slope of this relationship ('g(1) ') is constant across species. However, if different plant species have adapted to different access costs of water, then there will be differences in g(1) among species. We hypothesized that g(1) should vary among species adapted to different climates, and tested the theory and its linkage to plant hydraulics using four Eucalyptus species from different climatic origins in a common garden. Optimal stomatal theory predicts that species from sub-humid zones have a lower marginal water cost of C gain, hence lower g(1) than humid-zone species. In agreement with the theory that g(1) is related to tissue carbon costs for water supply, we found a relationship between wood density and g(1) across Eucalyptus species of contrasting climatic origins. There were significant reductions in the parameter g(1) during drought in humid but not sub-humid species, with the latter group maintaining g(1) in drought. There are strong differences in stomatal behaviour among related tree species in agreement with optimal stomatal theory, and these differences are consistent with the economics involved in water uptake and transport for carbon gain., (© 2012 Blackwell Publishing Ltd.)

Eucalyptus tetrodonta, a co-dominant tree species of tropical, northern Australian savannas, does not invade adjacent monsoon rain forest unless the forest is burnt intensely. Such facilitation by fire of seedling establishment is known as the "ashbed effect." Because the ashbed effect might involve disruption of common mycorrhizal networks, we hypothesized that in the absence of fire, intact rain forest arbuscular mycorrhizal (AM) networks inhibit E. tetrodonta seedlings. Although arbuscular mycorrhizas predominate in the rain forest, common tree species of the northern Australian savannas (including adult E. tetrodonta) host ectomycorrhizas. To test our hypothesis, we grew E. tetrodonta and Ceiba pentandra (an AM-responsive species used to confirm treatments) separately in microcosms of ambient or methyl-bromide fumigated rain forest soil with or without severing potential mycorrhizal fungus connections to an AM nurse plant, Litsea glutinosa. As expected, C. pentandra formed mycorrhizas in all treatments but had the most root colonization and grew fastest in ambient soil. E. tetrodonta seedlings also formed AM in all treatments, but severing hyphae in fumigated soil produced the least colonization and the best growth. Three of ten E. tetrodonta seedlings in ambient soil with intact network hyphae died. Because foliar chlorosis was symptomatic of iron deficiency, after 130 days we began to fertilize half the E. tetrodonta seedlings in ambient soil with an iron solution. Iron fertilization completely remedied chlorosis and stimulated leaf growth. Our microcosm results suggest that in intact rain forest, common AM networks mediate belowground competition and AM fungi may exacerbate iron deficiency, thereby enhancing resistance to E. tetrodonta invasion. Common AM networks-previously unrecognized as contributors to the ashbed effect-probably help to maintain the rain forest-savanna boundary.

The rise in atmospheric [CO(2)] is associated with increasing air temperature. However, studies on plant responses to interactive effects of [CO(2)] and temperature are limited, particularly for leaf structural attributes. In this study, Eucalyptus saligna plants were grown in sun-lit glasshouses differing in [CO(2)] (290, 400, and 650 µmol mol(-1)) and temperature (26 °C and 30 °C). Leaf anatomy and chloroplast parameters were assessed with three-dimensional confocal microscopy, and the interactive effects of [CO(2)] and temperature were quantified. The relative influence of leaf structural attributes and chemical properties on the variation of leaf mass per area (LMA) and photosynthesis within these climate regimes was also determined. Leaf thickness and mesophyll size increased in higher [CO(2)] but decreased at the warmer temperature; no treatment interaction was observed. In pre-industrial [CO(2)], warming reduced chloroplast diameter without altering chloroplast number per cell, but the opposite pattern (reduced chloroplast number per cell and unchanged chloroplast diameter) was observed in both current and projected [CO(2)]. The variation of LMA was primarily explained by total non-structural carbohydrate (TNC) concentration rather than leaf thickness. Leaf photosynthetic capacity (light- and [CO(2)]-saturated rate at 28 °C) and light-saturated photosynthesis (under growth [CO(2)] and temperature) were primarily determined by leaf nitrogen contents, while secondarily affected by chloroplast gas exchange surface area and chloroplast number per cell, respectively. In conclusion, leaf structural attributes are less important than TNC and nitrogen in affecting LMA and photosynthesis responses to the studied climate regimes, indicating that leaf structural attributes have limited capacity to adjust these functional traits in a changing climate.

Premise of the Study: Herein, we name, describe, and illustrate new macrofossil material representing Eucalyptus (Myrtaceae: Myrtoideae, Eucalypteae) from the diverse early Eocene Laguna del Hunco (LH) flora of Chubut Province, Patagonia, Argentina. We explore the significance of these fossils in light of understanding the fossil record of eucalypts and the biogeography of the Eucalypteae., Methods: Fossils representing vegetative and reproductive organs were collected from multiple LH localities over several field seasons. These specimens were prepared, photographed, and compared to extant Eucalyptus. Additional historical collections of Patagonian fossil Eucalyptus were also examined., Key Results: Vegetative and reproductive organs representing five different Eucalyptus taxa were identified in the LH paleoflora. One new taxon each representing leaves, flower buds, and infructescences with co-occurring, isolated capsules are described and named as new Eucalyptus species. Additionally, two flower types cf. Eucalyptus, represented by one specimen each, are illustrated and briefly described. The fossil species have unique characteristics that independently suggest each belongs within the Eucalypteae. The reproductive material is most similar morphologically to extant Eucalyptus, although it also shares many similarities to the closely related genus Corymbia., Conclusions: The LH fossil Eucalyptus material is among the few eucalypt macrofossils that have recently been named and described and are the oldest macrofossils that can presently be definitively ascribed to the Eucalypteae. They also represent the only credible description of Eucalyptus fossils occurring outside of Australasia and suggest a once broader geographic distribution for this group.

Background and Aims: Worldwide, many plant species are confined to open, shallow-soil, rocky habitats. Although several hypotheses have been proposed to explain this habitat specificity, none has been convincing. We suggest that the high level of endemism on shallow soils is related to the edaphic specialization needed to survive in these often extremely drought-prone habitats. Previous research has shown that species endemic to ironstone communities in SW Australia have a specialized root morphology that enhances their chance to access fissures in the underlying rock. Here we test the generality of these findings for species that are confined to a shallow-soil habitat that is of much greater global significance: granite outcrops., Methods: We compared temporal and spatial root growth and allocation of three endemic woody perennials of SW Australian granite outcrop communities with those of congeners occurring on nearby deeper soils. Seedlings of all species were grown in 1·2 m long custom-made containers with a transparent bottom that allowed monitoring of root growth over time., Key Results: The granite outcrop endemics mostly differed in a predictable way from their congeners from deeper soils. They generally invested a larger portion of their biomass in roots, distributed their roots faster and more evenly over the container and had a lower specific root length. In different species pairs the outcrop endemics achieved their apparent advantage by a different combination of the aforementioned traits., Conclusions: Our results are consistent with earlier work, indicating that species restricted to different types of drought-prone shallow-soil communities have undergone similar selection pressures. Although adaptive in their own habitat in terms of obtaining access to fissures in the underlying rock, these root system traits are likely to be maladaptive in deeper soil habitats. Therefore, our results may provide an explanation for the narrow endemism of many shallow-soil endemics.

Competition for light has driven forest trees to grow exceedingly tall, but the lack of a single universal limit to tree height indicates multiple interacting environmental limitations. Because soil nutrient availability is determined by both nutrient concentrations and soil water, water and nutrient availabilities may interact in determining realised nutrient availability and consequently tree height. In SW Australia, which is characterised by nutrient impoverished soils that support some of the world's tallest forests, total [P] and water availability were independently correlated with tree height (r = 0.42 and 0.39, respectively). However, interactions between water availability and each of total [P], pH and [Mg] contributed to a multiple linear regression model of tree height (r = 0.72). A boosted regression tree model showed that maximum tree height was correlated with water availability (24%), followed by soil properties including total P (11%), Mg (10%) and total N (9%), amongst others, and that there was an interaction between water availability and total [P] in determining maximum tree height. These interactions indicated a trade-off between water and P availability in determining maximum tree height in SW Australia. This is enabled by a species assemblage capable of growing tall and surviving (some) disturbances. The mechanism for this trade-off is suggested to be through water enabling mass-flow and diffusive mobility of P, particularly of relatively mobile organic P, although water interactions with microbial activity could also play a role.

Recovery following defoliation can be modified by co-occurring site resource limitations. The growth response of young Eucalyptus globulus saplings to two defoliation events was examined in an experimental plantation with combinations of low (-) or high (+) water (W) and nitrogen (N) resources. Artificial defoliation was applied at 3 and 9 months of age to remove ~40 and 55% of leaf area in the upper crown, respectively. At 18 months of age, height, stem diameter and leaf area were not significantly different between control and defoliated saplings, across all resource treatments. However, stem volume, bark volume and branch number were significantly increased in defoliated saplings, including a significant interaction with resource treatment. Total above-ground biomass of saplings in response to defoliation was significantly higher (almost double) than controls for the low water (N + W-) treatment only. Significantly increased foliar starch content (and a trend for increased soluble sugars) in the upper crown zone was found in the defoliated saplings of the N + W- treatment compared with the upper zone of control saplings. Foliar total non-structural carbohydrates were significantly correlated to stem biomass regardless of resource treatment or defoliation, and we suggest that foliar resources are most important for stem growth in E. globulus rather than stored carbon (C) from other tissues. After repeated defoliation and several months recovery, E. globulus saplings were generally not C limited in this study.

Background: Eucalyptus is an important genus in industrial plantations throughout the world and is grown for use as timber, pulp, paper and charcoal. Several breeding programmes have been launched worldwide to concomitantly improve growth performance and wood properties (WPs). In this study, an interspecific cross between Eucalyptus urophylla and E. grandis was used to identify major genomic regions (Quantitative Trait Loci, QTL) controlling the variability of WPs., Results: Linkage maps were generated for both parent species. A total of 117 QTLs were detected for a series of wood and end-use related traits, including chemical, technological, physical, mechanical and anatomical properties. The QTLs were mainly clustered into five linkage groups. In terms of distribution of QTL effects, our result agrees with the typical L-shape reported in most QTL studies, i.e. most WP QTLs had limited effects and only a few (13) had major effects (phenotypic variance explained > 15%). The co-locations of QTLs for different WPs as well as QTLs and candidate genes are discussed in terms of phenotypic correlations between traits, and of the function of the candidate genes. The major wood property QTL harbours a gene encoding a Cinnamoyl CoA reductase (CCR), a structural enzyme of the monolignol-specific biosynthesis pathway., Conclusions: Given the number of traits analysed, this study provides a comprehensive understanding of the genetic architecture of wood properties in this Eucalyptus full-sib pedigree. At the dawn of Eucalyptus genome sequence, it will provide a framework to identify the nature of genes underlying these important quantitative traits.

The patterns of hybridization and asymmetrical gene flow among species are important for understanding the processes that maintain distinct species. We examined the potential for asymmetrical gene flow in sympatric populations of Eucalyptus aggregata and Eucalyptus rubida, both long-lived trees of southern Australia. A total of 421 adults from three hybrid zones were genotyped with six microsatellite markers. We used genealogical assignments, admixture analysis and analyses of spatial genetic structure and spatial distribution of individuals, to assess patterns of interspecific gene flow within populations. A high number of admixed individuals were detected (13.9-40% of individuals), with hybrid populations consisting of F(1) and F(2) hybrids and backcrosses in both parental directions. Across the three sites, admixture proportions were skewed towards the E. aggregata genetic cluster (x=0.56-0.65), indicating that backcrossing towards E. aggregata is more frequent. Estimates of long-term migration rates also indicate asymmetric gene flow, with higher migration rates from E. aggregata to hybrids compared with E. rubida. Taken together, these results indicate a greater genetic input from E. aggregata into the hybrid populations. This asymmetry probably reflects differences in style lengths (E. rubida: ~7 mm, E. aggregata: ~4 mm), which can prevent pollen tubes of smaller-flowered species from fertilizing larger-flowered species. However, analyses of fine-scale genetic structure suggest that localized seed dispersal (<40 m) and greater clustering between hybrid and E. aggregata individuals may also contribute to directional gene flow. Our study highlights that floral traits and the spatial distributions of individuals can be useful predictors of the directionality of interspecific gene flow in plant populations.

Essential oils obtained by hydrodistillation from the different parts (stems, adult leaves, immature flowers and fruits) of Eucalyptus oleosa were screened for their antioxidant and antimicrobial properties and their chemical composition. According to GC-FID and GC-MS, the principal compound of the stem, immature flowers and the fruit oils was 1,8-cineole, representing 31.5%, 47.0% and 29.1%, respectively. Spathulenol (16.1%) and γ-eudesmol (15.0%) were the two principal compounds of adult leaves oil. In the DPPH (1,1-diphenyl-2-picrylhydrazyl) assay, the oils of the four parts showed moderate antioxidant activity. In the ABTS (2,2'-azinobis-3-ethylbenzothiazoline-6-sulphonate) assay, the most active part was the adult leaves, with a IC(50) value 13.0 ± 0.6 mg/L, followed by stems (IC(50) = 43.5 ± 1.4 mg/L). The essential oils showed a better antibacterial activity against Gram-positive and Gram-negative bacteria, and a significant antifungal activity also was observed against yeast-like fungi. A strong correlations between oxygenated monoterpenes and antimicrobial activity (especially 1,8-cineole) were noted (R2 = 0.99, 0.97 and 0.79 for B. subtilis, P. aeruginosa and C. albicans, respectively).

Background: Eucalyptus globulus and its hybrids are very important for the cellulose and paper industry mainly due to their low lignin content and frost resistance. However, rooting of cuttings of this species is recalcitrant and exogenous auxin application is often necessary for good root development. To date one of the most accurate methods available for gene expression analysis is quantitative reverse transcription-polymerase chain reaction (qPCR); however, reliable use of this technique requires reference genes for normalization. There is no single reference gene that can be regarded as universal for all experiments and biological materials. Thus, the identification of reliable reference genes must be done for every species and experimental approach. The present study aimed at identifying suitable control genes for normalization of gene expression associated with adventitious rooting in E. globulus microcuttings., Results: By the use of two distinct algorithms, geNorm and NormFinder, we have assessed gene expression stability of eleven candidate reference genes in E. globulus: 18S, ACT2, EF2, EUC12, H2B, IDH, SAND, TIP41, TUA, UBI and 33380. The candidate reference genes were evaluated in microccuttings rooted in vitro, in presence or absence of auxin, along six time-points spanning the process of adventitious rooting. Overall, the stability profiles of these genes determined with each one of the algorithms were very similar. Slight differences were observed in the most stable pair of genes indicated by each program: IDH and SAND for geNorm, and H2B and TUA for NormFinder. Both programs identified UBI and 18S as the most variable genes. To validate these results and select the most suitable reference genes, the expression profile of the ARGONAUTE1 gene was evaluated in relation to the most stable candidate genes indicated by each algorithm., Conclusion: Our study showed that expression stability varied between putative reference genes tested in E. globulus. Based on the AGO1 relative expression profile obtained using the genes suggested by the algorithms, H2B and TUA were considered as the most suitable reference genes for expression studies in E. globulus adventitious rooting. UBI and 18S were unsuitable for use as controls in qPCR related to this process. These findings will enable more accurate and reliable normalization of qPCR results for gene expression studies in this economically important woody plant, particularly related to rooting and clonal propagation.

*Recent research suggests that increasing conduit tapering progressively reduces hydraulic constraints caused by tree height. Here, we tested this hypothesis using the tallest hardwood species, Eucalyptus regnans. *Vertical profiles of conduit dimensions and vessel density were measured for three mature trees of height 47, 51 and 63 m. *Mean hydraulic diameter (Dh) increased rapidly from the tree apex to the point of crown insertion, with the greatest degree of tapering yet reported (b > 0.33). Conduit tapering was such that most of the total resistance was found close to the apex (82-93% within the first 1 m of stem) and the path length effect was reduced by a factor of 2000. Vessel density (VD) declined from the apex to the base of each tree, with scaling parameters being similar for all trees (a = 4.6; b = -0.5). *Eucalyptus regnans has evolved a novel xylem design that ensures a high hydraulic efficiency. This feature enables the species to grow quickly to heights of 50-60 m, beyond the maximum height of most other hardwood trees.

This paper reports on variation in leaf area index (L) in five Eucalyptus globulus Labill. plantations in response to application of nitrogen, thinning at age 2 years and variation in climate wetness index (the ratio of rainfall to potential evaporation). Observed L is compared with: (i) L predicted to optimize net primary productivity for a given average annual temperature, annual water use and potential evaporation (L(opt)) and (ii) L calculated as a linear function of climate wetness index (L(eq)). L peaked in fertilized plots at between 4 and 5 years of age or immediately after canopy closure. The value of L from canopy closure to age 8 years was not strongly related to annual rainfall or climate wetness index. At two sites with total soil nitrogen <1.2 mg g(-)(1), L in fertilized plots was about two units greater than in unfertilized plots. This difference persisted until measurements ended in 2004 when the trees were 8 years old. The L of plots thinned to 300 and 600 stems ha(-)(1) at age 2 years recovered quickly and was not significantly different from L in unthinned plots when the trees were 8 years old. L(opt) was a good predictor of the leaf area index of 8-year-old plots of E. globulus when nitrogen and phosphorus were non-limiting (model efficiency (EF) was 0.5). For the same plots, L(eq) underestimated observed L by an average of two units, and the model efficiency was low (-3.25). Data from two nitrogen-limited sites demonstrated that for fertilized plots L(opt) (EF = 0.6) was a much better predictor of L than L(eq) (EF = -3.36). At the same sites, L(eq) (EF = 0.42) was a better model for predicting L of unfertilized plots than L(opt) (-3.59). These results provide evidence that comparing observed L with L(opt) can identify stands limited by factors other than growing climate.