Cortical microtubules comprise heterodimeric units of alpha- and beta-tubulin which have been shown to guide the deposition of cellulose microfibrils in plant cell walls where their arrangement is important in determining cell morphology and cell wall properties. Tubulin genes are highly expressed in woody tissues and a functional study has demonstrated a role for a beta-tubulin gene family member in affecting the orientation of cellulose microfibrils in wood fibre cells, an important trait in determining the mechanical properties of wood fibres. To further understand the role of tubulins in plant cell trait determination, this study identified and investigated the expression of the alpha- and beta-tubulin gene families in Eucalyptus and then, using transgenesis techniques, investigated the role of specific eucalypt tubulin isoforms in determining secondary cell wall traits of wood fibres in plant stems. This study found that the alpha- and beta-tubulin gene families in Eucalyptus are relatively small compared to other species and show higher expression in woody stem tissue when compared to leaf. Functional studies revealed that cambial cells transformed with alpha- and beta-tubulin overexpression and knockdown vectors, either on their own or in combination, lead to changes in the angle of microfibrils in the secondary cell wall of wood fibre cells with Class I- and Class I-like gene family members explicitly involved. This study demonstrates the importance of tubulins in determining the mechanical properties of wood fibres through a mechanism involving specific tubulin isoform expression during wood fibre formation.

Short-rotation plantation hardwoods are increasingly used to meet timber demand but are often harvested before optimal structural properties are achieved. This limits their suitability for high-value timber applications. Improving the mechanical properties of young trees through selective breeding is a viable strategy, particularly when supported by molecular tools. However, limited information exists on how genetic and phenotypic variation influence structural traits across the stem profile. This study assessed variation in the mechanical properties of Eucalyptus pellita and its hybrids to support breeding for structural timber. Modulus of elasticity (MOE), modulus of rupture (MOR), and compressive strength (CS) were measured in inner and outer wood across multiple stem heights. These traits were correlated with physical properties, growth, and chemical traits. A single nucleotide polymorphism (SNP)-based genetic association analysis was also conducted. Outer wood and middle stem positions showed the highest values, with MOE up to 17,700 MPa, MOR up to 147 MPa, and CS up to 70 MPa, particularly in the E. pellita × E. brassiana hybrid. Mechanical traits correlated positively with height and diameter at breast height, while microfibril angle was negatively associated with CS (r = − 0.30 inner; − 0.24 outer). Lignin content showed positive associations with MOR and CS. SNP-based genetic analysis identified loci linked to MOE and MOR, including SNP4096 near Eucgr.A00211, a candidate gene potentially involved in cell wall regulation and wood strength. These findings highlight the potential of combining phenotypic and genomic tools to improve the selection of fast-grown hardwoods for high-value structural applications.

Eucalyptus pellita is cultivated for pulpwood in tropical and subtropical regions but remains underutilised for structural applications, particularly in dry tropical environments. Compared to well-studied species such as Eucalyptus grandis and Eucalyptus globulus, its potential in engineered wood products is poorly understood. As plantation industries seek to diversify products and increase wood value, understanding the structural suitability of E. pellita hybrids is critical. This study evaluated the performance of E. pellita and its F1 and backcross hybrids across two dry tropical sites, spanning log quality, product recovery, structural properties, and genetic associations. Billets were assessed for form, diameter, and defects. Sawing and peeling trials quantified recovery rates and defects. Laminated veneer lumber (LVL) was fabricated from hybrid veneers and tested for density, modulus of elasticity (MOE), modulus of rupture (MOR), and bond quality. The E. pellita x Eucalyptus brassiana hybrid achieved highest recoveries among the materials tested (sawn board recovery up to 49.2

Dimensional stability is a key trait for structural wood applications such as flooring, yet its genetic basis in Eucalyptus pellita F.Muell. and its hybrids remain poorly understood. Addressing this gap is essential for improving processing efficiency and product quality through targeted breeding. This study assessed variation in shrinkage and density, their relationships with growth and chemical traits, and associated genetic markers. Wood samples from E. pellita, E. pellita × E. urophylla S.T.Blake, and E. pellita × E. brassiana S.T.Blake were collected from two plantation sites in northern Australia. Radial and tangential shrinkage and density were measured alongside growth and chemical traits. SNP genotyping was conducted to identify markers linked to these physical properties. Significant differences were observed among hybrid types. E. pellita × E. urophylla recorded the lowest tangential unit shrinkage (0.06%), while E. pellita × E. brassiana had the highest basic density (651 kg/m3). Shrinkage and density showed moderate to strong correlations with growth and chemical traits. Several SNPs were associated with these properties; all were located in the intergenic region near Eucgr.A00211. Among these, only one SNP exceeded the −log10(p) significance threshold. These results provide early genetic insights and potential candidate markers for improving wood quality in Eucalyptus breeding programs. This exploratory study, constrained by a small sample size (n = 58), identifies putative SNPs for future validation in broader, multi-environment trials.

Key message Cortical microtubule arrays are the primary mechanism for guiding the re-orientation of cellulose microfibrils and determining MFA in secondary cell wall of wood fibre and tracheid cells in reaction wood. Abstract Microtubules are directly and indirectly involved in guiding cellulose synthase complexes (CSCs) through the plasma membrane. The angle of cellulose deposition is a critical response to environmental signals and/or stress conditions, and particularly crucial during reaction wood formation, a process in which woody plants deposit additional cell wall material to counteract gravitational forces. Tubulin genes are upregulated in response to gravitational stimulus during reaction wood formation, which can result in changes to microtubule assembly. In this study, microtubules were visualised in three woody tree species (two angiosperms: Eucalyptus globulus Labill., Populus alba L., and one gymnosperm: Pinus radiata D. Don.) using immunofluorescence to quantitatively evaluate microtubule organisation during reaction wood formation. Our results suggest that reorientation of the cortical microtubule array affects secondary cell wall deposition, even across different types of reaction wood, by ensuring context-appropriate orientation of cellulose microfibrils and determining MFA in wood cells. Pharmacological studies conducted on in vitro cultured stem segments or in vivo during reaction wood formation corroborated these important roles for microtubules during wood development. This study starts to unveil the role of tubulins during wood formation by exploring cortical microtubule array organisation in trees subjected to gravitational stimulus and it sheds light on cellular and molecular mechanisms behind cellulose deposition in tree species.

Trees play a pivotal role in terrestrial ecosystems as well as being an important natural resource. These attributes are primarily associated with the capacity of trees to continuously produce woody tissue from the vascular cambium, a ring of stem cells located just beneath the bark. Long-lived trees are exposed to a myriad of biological and environmental stresses that may result in wounding, leading to a loss of bark and the underlying vascular cambium. This affects both wood formation and the quality of timber arising from the tree. In addition, the exposed wound site is a potential entry point for pathogens that cause disease. In response to wounding, trees have the capacity to regenerate lost or damaged tissues at this site. Investigating gene expression changes associated with different stages of wound healing reveals complex and dynamic changes in the activity of transcription factors, signalling pathways and hormone responses. In this review we summarise these data and discuss how they relate to our current understanding of vascular cambium formation and xylem differentiation during secondary growth. Based on this analysis, a model for wound healing that provides the conceptual foundations for future studies aimed at understanding this intriguing process is proposed.

Trees play a pivotal role in terrestrial ecosystems as well as being an important natural resource. These attributes are primarily associated with the capacity of trees to continuously produce woody tissue from the vascular cambium, a ring of stem cells located just beneath the bark. Long-lived trees are exposed to a myriad of biological and environmental stresses that may result in wounding, leading to a loss of bark and the underlying vascular cambium. This affects both wood formation and the quality of timber arising from the tree. In addition, the exposed wound site is a potential entry point for pathogens that cause disease. In response to wounding, trees have the capacity to regenerate lost or damaged tissues at this site. Investigating gene expression changes associated with different stages of wound healing reveals complex and dynamic changes in the activity of transcription factors, signalling pathways and hormone responses. In this review we summarise these data and discuss how they relate to our current understanding of vascular cambium formation and xylem differentiation during secondary growth. Based on this analysis, a model for wound healing that provides the conceptual foundations for future studies aimed at understanding this intriguing process is proposed.

Abstract Plant chimaeras and mosaics are not only found in our gardens as variegated ornamentals. They also constitute valuable research tools with a range of applications in developmental and molecular biology. For example, much of our knowledge about the development of the shoot and root apical meristems is derived from studying patterns of variegation, and where patterns persist throughout the plant, deductions can be made as to the fate of individual meristematic cells and the number and arrangement of initial cells giving rise to a variety of plant organs or structures. At a molecular level, chimaeras and mosaics can shed light on the cell autonomy of a certain trait, inform us about the propagation and movement of molecular signals and indeed allow us to directly compare genetically distinct tissues in a single organism to determine specific gene function. Key Concepts Meristematic tissue contains undifferentiated cells which divide and differentiate to produce the plant body. Apical meristems are found at the tips of shoots and roots and comprise of meristematic tissue. By grafting, plant parts are manually brought together to physically connect to each other and continue to grow. Fate mapping is a methodology used in developmental biology to understand how cell/s divide to contribute to an organism's body. Cell signalling involves obtaining, interpreting and/or transferring information between cells and their environment. Genetic transformation involves the transfer of foreign DNA and its integration into the genome of a host cell from where it is then passed onto daughter cells. Bud sports are lateral shoots or inflorescences which are visibly different to the rest of the plant.

Microtubules are directly and indirectly involved in guiding cellulose synthase complexes (CSCs) through the plasma membrane. The angle of cellulose deposition is a critical response to environmental signals and/or stress conditions, and particularly crucial during reaction wood formation, a process in which woody plants deposit additional cell wall material to counteract gravitational forces. Tubulin genes are upregulated in response to gravitational stimulus during reaction wood formation, which can result in changes to microtubule assembly. In this study, microtubules were visualised in three woody tree species (two angiosperms: Eucalyptus globulus Labill., Populus alba L., and one gymnosperm: Pinus radiata D. Don.) using immunofluorescence to quantitatively evaluate microtubule organisation during reaction wood formation. Our results suggest that reorientation of the cortical microtubule array affects secondary cell wall deposition, even across different types of reaction wood, by ensuring context-appropriate orientation of cellulose microfibrils and determining MFA in wood cells. Pharmacological studies conducted on in vitro cultured stem segments or in vivo during reaction wood formation corroborated these important roles for microtubules during wood development. This study starts to unveil the role of tubulins during wood formation by exploring cortical microtubule array organisation in trees subjected to gravitational stimulus and it sheds light on cellular and molecular mechanisms behind cellulose deposition in tree species.

Key message Our Induced Somatic Sector Analysis and protein–protein interaction experiments demonstrate that Eucalyptus grandis IAA13 regulates xylem fibre and vessel development, potentially via EgrIAA13 modules involving ARF2, ARF5, ARF6 and ARF19. Abstract Auxin is a crucial phytohormone regulating multiple aspects of plant growth and differentiation, including regulation of vascular cambium activity, xylogenesis and its responsiveness towards gravitropic stress. Although the regulation of these biological processes greatly depends on auxin and regulators of the auxin signalling pathway, many of their specific functions remain unclear. Therefore, the present study aims to functionally characterise Eucalyptus grandis AUX/INDOLE-3-ACETIC ACID 13 ( EgrIAA13 ), a member of the auxin signalling pathway. In Eucalyptus and Populus , Egr IAA13 and its orthologs are preferentially expressed in the xylogenic tissues and downregulated in tension wood. Therefore, to further investigate EgrIAA13 and its function during xylogenesis, we conducted subcellular localisation and Induced Somatic Sector Analysis experiments using overexpression and RNAi knockdown constructs of EgrIAA13 to create transgenic tissue sectors on growing stems of Eucalyptus and Populus . Since Aux/IAAs interact with Auxin Responsive Factors (ARFs), in silico predictions of IAA13-ARF interactions were explored and experimentally validated via yeast-2-hybrid experiments. Our results demonstrate that EgrIAA13 localises to the nucleus and that downregulation of EgrIAA13 impedes Eucalyptus xylem fibre and vessel development. We also observed that EgrIAA13 interacts with Eucalyptus ARF2, ARF5, ARF6 and ARF19A. Based on these results, we conclude that EgrIAA13 is a regulator of Eucalyptus xylogenesis and postulate that the observed phenotypes are likely to result from alterations in the auxin-responsive transcriptome via IAA13-ARF modules such as EgrIAA13-EgrARF5. Our results provide the first insights into the regulatory role of EgrIAA13 during xylogenesis.

Myrtle rust (caused by Austropuccinia psidii Beenken) is exotic to Australia, yet specific resistance is present in a range of naïve myrtaceous host plants. Resistance to myrtle rust is primarily quantitative in nature and controlled by multiple interacting loci. We undertook a genome-wide association study (GWAS) to discover relationships between nuclear genomic sequence variation and multiple aspects of resistance to A. psidii in Eucalyptus obliqua, a representative species from subgenus Eucalyptus. Sequence variation was assessed with respect to numeric severity, binary symptomatic, hypersensitive, and pustulation responses to inoculation of seedlings with A. psidii. A total of 1.13 million, single-nucleotide polymorphisms (SNPs) were tested for association with the rust resistance responses of 637 phenotyped E. obliqua seedlings, each from different, single mother tree seedlots. Thirty-three highly significant SNP-trait associations were detected, of which 26 associated with the binary symptomatic/asymptomatic response to A. psidii. Comparison of the genomic position of these SNPs with rust resistance loci (Ppr1-5), previously reported in species from subgenus Symphyomyrtus, suggested that several were positioned near the major Ppr1 locus and other Ppr loci. This study provides the first integrated genomic view of A. psidii resistance across the Eucalyptus subgenera and provides the foundation for discovering key resistance genes for use in marker-based resistance breeding.

The Editorial Office of Plants wants to make the following corrections to the paper by Tobias, L.M., et al. (2020)[...]

Recent advances in our understanding of the molecular control of secondary cell wall (SCW) formation have shed light on molecular mechanisms that underpin domestication traits related to wood formation. One such trait is the cellulose microfibril angle (MFA), an important wood quality determinant that varies along tree developmental phases and in response to gravitational stimulus. The cytoskeleton, mainly composed of microtubules and actin filaments, collectively contribute to plant growth and development by participating in several cellular processes, including cellulose deposition. Studies in Arabidopsis have significantly aided our understanding of the roles of microtubules in xylem cell development during which correct SCW deposition and patterning are essential to provide structural support and allow for water transport. In contrast, studies relating to SCW formation in xylary elements performed in woody trees remain elusive. In combination, the data reviewed here suggest that the cytoskeleton plays important roles in determining the exact sites of cellulose deposition, overall SCW patterning and more specifically, the alignment and orientation of cellulose microfibrils. By relating the reviewed evidence to the process of wood formation, we present a model of microtubule participation in determining MFA in woody trees forming reaction wood (RW).

The products of secondary xylem are of significant biological and commercial importance, and as a result, the biology of secondary growth and how intrinsic and extrinsic factors influence this process have been the subject of intense investigation. Studies into secondary xylem range in scale from the cellular to the forest stand level, with phenotypic analyses often involving the assessment of traits relating to cell morphology and cell wall chemical composition. While numerous techniques are currently available for phenotypic analyses of samples containing abundant amounts of secondary tissue, only a few of them (microanalytical techniques) are suitable when working with limiting amounts of secondary tissue or where a fine-scale resolution of morphological features or cell wall chemical composition is required. While polarised light microscopy, scanning electron microscopy, field emission-scanning electron microscopy and X-ray scattering and micro-tomography techniques serve as the most frequently used microanalytical techniques in morphotyping, techniques such as scanning ultraviolet microspectrophotometry, X-ray photoelectron spectroscopy, gas chromatography, Fouriertransform infrared spectroscopy and matrix-assisted laser desorption ionisation mass spectrometry serve as the most commonly used microanalytical techniques in chemotyping. Light microscopy, fluorescence microscopy, confocal laser scanning microscopy, transmission electron microscopy and Raman spectroscopy serve as dual micro morphotyping and chemotyping techniques. In this review, we summarise and discuss these techniques in the light of their applicability as microanalytical techniques to study secondary xylem.

Eucalyptus foliar terpenes have been reported widely to function as chemical defence agents against mammalian herbivores and defoliating insects both via direct toxicity and/or indirect priming of systemic defence mechanisms. Less frequently they have been reported as disease resistance biomarkers. The present study compares composition of foliar essential oils in Eucalyptus globulus Labill. and Eucalyptus obliqua L’Hér. between distinct myrtle rust (causal pathogen Austropuccinia psidii Beenken) response phenotypes i.e. completely resistant, hypersensitive and highly susceptible, using canonical discriminant analysis. Stepwise forward variable selection identified six terpene compounds (bicyclogermacrene, globulol, geraniol, β-pinene, cis-p-menth-2-en-1-ol and δ-terpinene) in E. globulus and four terpenes (δ-cadinene, caryophyllene oxide, longifolenaldehyde and α-humulene) in E. obliqua, which in combination, significantly discriminated between rust response phenotypes. The discriminating compounds varied between E. globulus and E. obliqua and were different from those previously identified as rust resistance biomarkers in Eucalyptus grandis W.Hill ex Maiden × Eucalyptus urophylla S.T.Blake hybrids (1,8-cineole and α-terpinyl acetate). While the discriminant functions were statistically significant within each species, cross validation of the marker sets between Eucalyptus species was not significant. These findings indicate that either (a) terpene composition or amount is not a driver of resistance to myrtle rust or (b) that terpene contribution to resistance is through combined synergistic effects of variable terpenes, likely in combination with other plant metabolites or resistance mechanisms.

Myrtle rust poses a significant biosecurity threat to Australia with potential for long‐term damaging impacts on native flora and plant industries. This study describes the disease cycle of Austropuccinia psidii, the myrtle rust pathogen, in Eucalyptus globulus and Eucalyptus obliqua, two commercially and ecologically important species from different subgenera of Eucalyptus. Ontogeny and morphology of infection structures of A. psidii on plants of both Eucalyptus species with different rust response phenotypes, i.e. completely resistant, hypersensitive and highly susceptible, were investigated. Plants were inoculated with single‐uredinium‐derived urediniospores and examined by scanning electron microscopy. No differences between rust response phenotypes were observed in germination of urediniospores, formation of appressoria or length of germ tubes. The growth of germ tubes had no affinity towards stomata of either species. Histological observations indicated direct penetration by infection pegs through the leaf cuticle and no penetration beyond the epidermis on rust‐resistant E. obliqua. Eucalyptus obliqua plants that were identified as susceptible to A. psidii at 3‐ and 6‐months‐old showed no disease when reinoculated with A. psidii at 12‐months‐old; this indicated possible early acquisition of adult plant resistance to A. psidii in this species. In the susceptible phenotype of E. globulus rust inoculation led to rapid colonization of leaf parenchyma cells with the disease cycle completed within 10 days. These findings provide valuable insights into host–pathogen interactions in the Eucalyptus–A. psidii pathosystem, which might be useful for the development of effective rust control strategies across Eucalyptus subgenera.

Myrtle rust, caused by the pathogen Austropuccinia psidii, is a disease affecting numerous species of Myrtaceae around the globe. Many Australian ecosystems are dominated by Myrtaceae, making them, along with the industries that rely on them, particularly vulnerable to this disease. With over 800 endemic species, Eucalyptus is a major genus within the Myrtaceae in Australia. Wide variation in response to A. psidii infection, from extreme susceptibility to resistance, has been reported among Eucalyptus species in which any pre-formed resistance to this invasive pathogen is unexpected. This study aims to define and contrast geographical patterns of variation in rust susceptibility within the overlapping, natural ranges of Eucalyptus globulus and Eucalyptus obliqua, two commercially and ecologically important species from different Eucalyptus subgenera. Phenotypic disease screening of seedlings of E. globulus races and E. obliqua forest districts (defined geographically) showed E. obliqua to be more susceptible than E. globulus with significant differences in disease susceptibility and symptomatic trait expression. Eucalyptus globulus showed a trend for decreased susceptibility to A. psidii from south- to north-eastern Tasmania, eastwards along the Otway Ranges and southward from the Strzelecki Ranges to the Wilson Promontory Lighthouse in Victoria, but no such geographical patterns were observed within E. obliqua. No significant correlations were found between climatic conditions (i.e. rainfall, temperature and elevation) and rust susceptibility at provenance levels in either species. Taken together, these results support a hypothesis that population divergence in resistance to A. psidii has not been driven by climate.

We used sector analysis to study cambium development and dynamics and to test whether fundamental developmental and functional differences exist between cambial initials as true 'stem cells' and more differentiated mother cells. In many higher plants, a cylindrical lateral meristem, the vascular cambium, forms along the plant axis. Most notably in stems of perennial tree species, this meristem gives rise to xylem (wood) towards the inside of the trunk and phloem (bark) towards the outside. As such, the vascular cambium is responsible for the production of most of the planet's forest biomass, significantly contributing to the global carbon cycle. Using the bacterial uidA reporter gene in Agrobacterium-based in vivo stem transformation experiments in poplar trees, we created 379 cambium sectors that originated from the transformation of individual cells. Results from our analysis of sector frequency and patterns are consistent with the poplar cambium featuring a single layer of true cambial initials (being able to divide both anti- and periclinally). We show that initials are frequently lost from the cambium, that such cell loss rarely occurs at mother cell level, that phloem and xylem differentiation are controlled independently, and that the frequency of mother cell replenishment is not pre-determined.