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Background and Aims: The petaline operculum that covers the inner whorls until anthesis and the woody capsule that develops after fertilization are reproductive structures of eucalypts that protect the flower and seeds. Although they are distinct organs, they both develop from flower buds and this common ontogeny suggests shared genetic control. In Eucalyptus globulus their morphology is variable and we aimed to identify the quantitative trait loci (QTL) underlying this variation and determine whether there is common genetic control of these ecologically and taxonomically important reproductive structures., Methods: Samples of opercula and capsules were collected from 206 trees that belong to a large outcrossed F2E. globulus mapping population. The morphological variation in these structures was characterized by measuring six operculum and five capsule traits. QTL analysis was performed using these data and a linkage map consisting of 480 markers., Key Results: A total of 27 QTL were detected for operculum traits and 28 for capsule traits, with the logarithm of odds ranging from 2.8 to 11.8. There were many co-located QTL associated with operculum or capsule traits, generally reflecting allometric relationships. A key finding was five genomic regions where co-located QTL affected both operculum and capsule morphology, and the overall trend for these QTL was to affect elongation of both organs. Some of these QTL appear to have a significant effect on the phenotype, with the strongest QTL explaining 26.4 % of the variation in operculum shape and 16.4 % in capsule shape. Flower bud measurements suggest the expression of these QTL starts during bud development. Several candidate genes were found associated with the QTL and their putative function is discussed., Conclusions: Variation in both operculum and capsule traits in E. globulus is under strong genetic control. Our results suggest that these reproductive structures share a common genetic pathway during flower bud development., (© The Author(s) 2022. 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.)

Drought is a major stress impacting forest ecosystems worldwide. We utilized quantitative trait loci (QTL) analysis to study the genetic basis of variation in (a) drought resistance and recovery and (b) candidate traits that may be associated with this variation in the forest tree Eucalyptus globulus. QTL analysis was performed using a large outcrossed F 2 mapping population from which 300 trees were phenotyped based on the mean performance of their open-pollinated F 3 progeny. Progenies were grown in a glasshouse in a randomized complete block design. A subset of seedlings was subjected to a drought treatment after which they were rewatered and scored for damage and growth postdrought. Nondroughted seedlings were assessed for growth traits as well as lignotuber size and resprouting following severe damage to the main stem. QTL were detected for most traits. Importantly, independent QTL were detected for (a) drought damage and plant size, (b) drought damage and growth recovery, and (c) lignotuber size and resprouting capacity. Such independence argues that trade-offs are unlikely to be a major limitation to the response to selection and at the early life history stage studied; there are opportunities to improve resilience to drought without adverse effects on productivity., (© 2019 John Wiley & Sons Ltd.)

Background and Aims: Many plants exhibit a mixed mating system. Published models suggest that this might be an evolutionarily stable rather than a transitional state despite the presence of inbreeding depression, but there is little empirical evidence. Through field experimentation, we studied the role of inbreeding depression in eliminating inbred progeny from the reproductive cohort of the forest tree Eucalyptus regnans, and demonstrate a stable mixed primary mating system over two successive generations., Methods: Two field experiments were conducted using seed from natural populations. We sowed open-pollinated seeds to simulate a natural regeneration event and determined isozyme genotypes of dominant and suppressed individuals over 10 years. We also planted a mixture of open-pollinated, outcross and selfed families with common maternal parentage; monitored survival of cross types over 29 years; and determined the percentage of outcrosses in open-pollinated seed from a sample of reproductively mature trees using microsatellite analysis., Key Results: Both experiments demonstrated progressive competitive elimination of inbred plants. By 29 years, the reproductive cohort in the planted experiment consisted only of outcrosses which produced seed which averaged 66 % outcrosses, similar to the estimate for the parental natural population (74 %)., Conclusions: Selective elimination of inbred genotypes during the intense intra-specific competition characteristic of the pre-reproductive phase of the life cycle of E. regnans results in a fully outcrossed reproductive population, in which self-fertility is comparable with that of its parental generation. The mixed mating system may be viewed as an unavoidable consequence of the species' reproductive ecology, which includes the demonstrated effects of inbreeding depression, rather than a strategy which is actively favoured by natural selection., (© The Author(s) 2019. 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.)

Intumescence is a nonpathogenic physiological disorder characterized by leaf blistering. This disorder can affect growth and development in glasshouses and growth chambers and may be confused with pathogenic diseases. We used quantitative trait loci (QTL) analysis to examine the genetic basis of variation in intumescence severity in Eucalyptus globulus, and test for colocation with previously detected QTLs for pathogen susceptibility. QTL analysis used the phenotype means of open-pollinated (OP) families of an outcrossed F 2 mapping family (OP F 3 ; n = 300) of E. globulus and the linkage map constructed in the F 2 . We validate this phenotyping approach for QTL analysis by assessing a trait previously used for QTL discovery in the F 2 and showing the same major QTL was detected with the OP F 3 . For intumescence severity, five putative QTLs were detected across four linkage groups. Four of these did not colocate with previously reported QTLs for fungal pathogen susceptibility in Eucalyptus, suggesting the mechanisms underlying susceptibility to intumescence and to the two fungal pathogens are largely independent. This study demonstrates there is a genetic basis for variation in intumescence severity, reports the first QTL for intumescence severity in plants, and provides a robust framework for investigating the potential mechanisms involved.

Eucalyptus globulus subsp. globulus Labill. (Tasmanian Blue Gum), native to southeast Australia, is a benchmark species for the pulp and paper industry. We genotyped 397 trees from 16 populations of E. globulus representing the native diversity in Australia using 24 microsatellite loci. Eight genetically distinct groups were detected, consistent with genetic groupings detected in previous quantitative and molecular studies. A sample of 29 Portuguese individuals was added to help clarify the origin of the Portuguese landrace. The results suggest a southern and eastern Tasmania origin for the Portuguese landrace. This genetic framework will enable researchers to investigate the provenance of individuals of unknown pedigree and assess the levels of representation of E. globulus natural variation in the Portuguese landrace.

Widespread species spanning strong environmental (e.g., climatic) gradients frequently display morphological and physiological adaptations to local conditions. Some adaptations are common to different species that occupy similar environments. However, the genomic architecture underlying such convergent traits may not be the same between species. Using genomic data from previous studies of three widespread eucalypt species that grow along rainfall gradients in southern Australia, our probabilistic approach provides evidence that adaptation to aridity is a genome-wide phenomenon, likely to involve multiple and diverse genes, gene families and regulatory regions that affect a multitude of complex genetic and biochemical processes., (© The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

We used genome-wide markers and an unprecedented scale of sampling to construct a phylogeny for a globally significant Eucalyptus lineage that has been impacted by hybridisation, recent radiation and morphological convergence. Our approach, using 3109 DArT markers distributed throughout the genome and 540 samples covering 185 terminal taxa in sections Maidenaria, Exsertaria, Latoangulatae and related smaller sections, with multiple geographically widespread samples per terminal taxon, produced a phylogeny that largely matched the morphological treatment of sections, though sections Exsertaria and Latoangulatae were polyphyletic. At lower levels there were numerous inconsistencies between the morphological treatment and the molecular phylogeny, and taxa within the three main sections were generally not monophyletic at the series (at least 62% polyphyly) or species (at least 52% polyphyly) level. Some of the discrepancies appear to be the result of morphological convergence or misclassifications, and we propose some taxonomic reassessments to address this. However, many inconsistencies appear to be the products of incomplete speciation and/or hybridisation. Our analysis represents a significant advance on previous phylogenies of these important eucalypt sections (which have mainly used single samples to represent each species), thus providing a robust phylogenetic framework for evolutionary and ecological studies., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: Meiotic recombination is a fundamental evolutionary process. It not only generates diversity, but influences the efficacy of natural selection and genome evolution. There can be significant heterogeneity in recombination rates within and between species, however this variation is not well understood outside of a few model taxa, particularly in forest trees. Eucalypts are forest trees of global economic importance, and dominate many Australian ecosystems. We studied recombination rate in Eucalyptus globulus using genetic linkage maps constructed in 10 unrelated individuals, and markers anchored to the Eucalyptus reference genome. This experimental design provided the replication to study whether recombination rate varied between individuals and chromosomes, and allowed us to study the genomic attributes and population genetic parameters correlated with this variation., Results: Recombination rate varied significantly between individuals (range = 2.71 to 3.51 centimorgans/megabase [cM/Mb]), but was not significantly influenced by sex or cross type (F1 vs. F2). Significant differences in recombination rate between chromosomes were also evident (range = 1.98 to 3.81 cM/Mb), beyond those which were due to variation in chromosome size. Variation in chromosomal recombination rate was significantly correlated with gene density (r = 0.94), GC content (r = 0.90), and the number of tandem duplicated genes (r = -0.72) per chromosome. Notably, chromosome level recombination rate was also negatively correlated with the average genetic diversity across six species from an independent set of samples (r = -0.75)., Conclusions: The correlations with genomic attributes are consistent with findings in other taxa, however, the direction of the correlation between diversity and recombination rate is opposite to that commonly observed. We argue this is likely to reflect the interaction of selection and specific genome architecture of Eucalyptus. Interestingly, the differences amongst chromosomes in recombination rates appear stable across Eucalyptus species. Together with the strong correlations between recombination rate and features of the Eucalyptus reference genome, we maintain these findings provide further evidence for a broad conservation of genome architecture across the globally significant lineages of Eucalyptus.

Plant cuticular wax compounds perform functions that are essential for the survival of terrestrial plants. Despite their importance, the genetic control of these compounds is poorly understood outside of model taxa. Here we investigate the genetic basis of variation in cuticular compounds in Eucalyptus globulus using quantitative genetic and quantitative trait loci (QTL) analyses. Quantitative genetic analysis was conducted using 246 open-pollinated progeny from 13 native sub-races throughout the geographic range. QTL analysis was conducted using 112 clonally replicated progeny from an outcross F2 population. Nine compounds exhibited significant genetic variation among sub-races with three exhibiting signals of diversifying selection. Fifty-two QTL were found with co-location of QTL for related compounds commonly observed. Notable among these was the QTL for five wax esters, which co-located with a gene from the KCS family, previously implicated in the biosynthesis of cuticular waxes in Arabidopsis. In combination, the QTL and quantitative genetic analyses suggest the variation and differentiation in cuticular wax compounds within E. globulus has a complex genetic origin. Sub-races exhibited independent latitudinal and longitudinal differentiation in cuticular wax compounds, likely reflecting processes such as historic gene flow and diversifying selection acting upon genes that have diverse functions in distinct biochemical pathways., (© 2015 University of Tasmania New Phytologist © 2015 New Phytologist Trust.)

We assess phylogenetic patterns of hybridization in the speciose, ecologically and economically important genus Eucalyptus, in order to better understand the evolution of reproductive isolation. Eucalyptus globulus pollen was applied to 99 eucalypt species, mainly from the large commercially important subgenus, Symphyomyrtus. In the 64 species that produce seeds, hybrid compatibility was assessed at two stages, hybrid-production (at approximately 1 month) and hybrid-survival (at 9 months), and compared with phylogenies based on 8,350 genome-wide DArT (diversity arrays technology) markers. Model fitting was used to assess the relationship between compatibility and genetic distance, and whether or not the strength of incompatibility "snowballs" with divergence. There was a decline in compatibility with increasing genetic distance between species. Hybridization was common within two closely related clades (one including E. globulus), but rare between E. globulus and species in two phylogenetically distant clades. Of three alternative models tested (linear, slowdown, and snowball), we found consistent support for a snowball model, indicating that the strength of incompatibility accelerates relative to genetic distance. Although we can only speculate about the genetic basis of this pattern, it is consistent with a Dobzhansky-Muller-model prediction that incompatibilities should snowball with divergence due to negative epistasis. Different rates of compatibility decline in the hybrid-production and hybrid-survival measures suggest that early-acting postmating barriers developed first and are stronger than later-acting barriers. We estimated that complete reproductive isolation can take up to 21-31 My in Eucalyptus. Practical implications for hybrid eucalypt breeding and genetic risk assessment in Australia are discussed., (© The Author 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

We examined genome-wide patterns of DNA sequence diversity and divergence among six species of the important tree genus Eucalyptus and investigated their relationship with genomic architecture. Using c. 90 range-wide individuals of each Eucalyptus species (E. grandis, E. urophylla, E. globulus, E. nitens, E. dunnii and E. camaldulensis), genetic diversity and divergence were estimated from 2840 polymorphic diversity arrays technology markers covering the 11 chromosomes. Species differentiating markers (SDMs) identified in each of 15 pairwise species comparisons, along with species diversity (HHW ) and divergence (FST ), were projected onto the E. grandis reference genome. Across all species comparisons, SDMs totalled 1.1-5.3% of markers and were widely distributed throughout the genome. Marker divergence (FST and SDMs) and diversity differed among and within chromosomes. Patterns of diversity and divergence were broadly conserved across species and significantly associated with genomic features, including the proximity of markers to genes, the relative number of clusters of tandem duplications, and gene density within or among chromosomes. These results suggest that genomic architecture influences patterns of species diversity and divergence in the genus. This influence is evident across the six species, encompassing diverse phylogenetic lineages, geography and ecology., (© 2015 University of Tasmania New Phytologist © 2015 New Phytologist Trust.)

Eucalypts are the world's most widely planted hardwood trees. Their outstanding diversity, adaptability and growth have made them a global renewable resource of fibre and energy. We sequenced and assembled >94% of the 640-megabase genome of Eucalyptus grandis. Of 36,376 predicted protein-coding genes, 34% occur in tandem duplications, the largest proportion thus far in plant genomes. Eucalyptus also shows the highest diversity of genes for specialized metabolites such as terpenes that act as chemical defence and provide unique pharmaceutical oils. Genome sequencing of the E. grandis sister species E. globulus and a set of inbred E. grandis tree genomes reveals dynamic genome evolution and hotspots of inbreeding depression. The E. grandis genome is the first reference for the eudicot order Myrtales and is placed here sister to the eurosids. This resource expands our understanding of the unique biology of large woody perennials and provides a powerful tool to accelerate comparative biology, breeding and biotechnology.

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.)

A change in the timing or rate of developmental events throughout ontogeny is referred to as heterochrony, and it is a major evolutionary process in plants and animals. We investigated the genetic basis for natural variation in the timing of vegetative phase change in the tree Eucalyptus globulus, which undergoes a dramatic change in vegetative morphology during the juvenile-to-adult transition. Quantitative trait loci analysis in an outcross F2 family derived from crosses between individuals from a coastal population of E. globulus with precocious vegetative phase change and individuals from populations in which vegetative phase change occurs several years later implicated the microRNA EglMIR156.5 as a potential contributor to this heterochronic difference. Additional evidence for the involvement of EglMIR156.5 was provided by its differential expression in trees with early and late phase change. Our findings suggest that changes in the expression of miR156 underlie natural variation in vegetative phase change in E. globulus, and may also explain interspecific differences in the timing of this developmental transition., (Copyright © 2014 Hudson et al.)

Patterns of adaptive variation within plant species are best studied through common garden experiments, but these are costly and time-consuming, especially for trees that have long generation times. We explored whether genome-wide scanning technology combined with outlier marker detection could be used to detect adaptation to climate and provide an alternative to common garden experiments. As a case study, we sampled nine provenances of the widespread forest tree species, Eucalyptus tricarpa, across an aridity gradient in southeastern Australia. Using a Bayesian analysis, we identified a suite of 94 putatively adaptive (outlying) sequence-tagged markers across the genome. Population-level allele frequencies of these outlier markers were strongly correlated with temperature and moisture availability at the site of origin, and with population differences in functional traits measured in two common gardens. Using the output from a canonical analysis of principal coordinates, we devised a metric that provides a holistic measure of genomic adaptation to aridity that could be used to guide assisted migration or genetic augmentation., (© 2014 John Wiley & Sons Ltd.)

· Eucalypts are one of the most planted tree genera worldwide, and there is increasing interest in marker-assisted selection for tree improvement. Implementation of marker-assisted selection requires a knowledge of the stability of quantitative trait loci (QTLs). This study aims to investigate the stability of QTLs for wood properties and growth across contrasting sites and multiple pedigrees of Eucalyptus globulus. · Saturated linkage maps were constructed using 663 genotypes from four separate families, grown at three widely separated sites, and were employed to construct a consensus map. This map was used for QTL analysis of growth, wood density and wood chemical traits, including pulp yield. · Ninety-eight QTLs were identified across families and sites: 87 for wood properties and 11 for growth. These QTLs mapped to 38 discrete regions, some of which co-located with candidate genes. Although 16% of QTLs were verified across different families, 24% of wood property QTLs and 38% of growth QTLs exhibited significant genotype-by-environment interaction. · This study provides the most detailed assessment of the effect of environment and pedigree on QTL detection in the genus. Despite markedly different environments and pedigrees, many QTLs were stable, providing promising targets for the application of marker-assisted selection., (© 2013 The Authors. New Phytologist © 2013 New Phytologist Trust.)

Background: Genetic linkage maps are invaluable resources in plant research. They provide a key tool for many genetic applications including: mapping quantitative trait loci (QTL); comparative mapping; identifying unlinked (i.e. independent) DNA markers for fingerprinting, population genetics and phylogenetics; assisting genome sequence assembly; relating physical and recombination distances along the genome and map-based cloning of genes. Eucalypts are the dominant tree species in most Australian ecosystems and of economic importance globally as plantation trees. The genome sequence of E. grandis has recently been released providing unprecedented opportunities for genetic and genomic research in the genus. A robust reference linkage map containing sequence-based molecular markers is needed to capitalise on this resource. Several high density linkage maps have recently been constructed for the main commercial forestry species in the genus (E. grandis, E. urophylla and E. globulus) using sequenced Diversity Arrays Technology (DArT) and microsatellite markers. To provide a single reference linkage map for eucalypts a composite map was produced through the integration of data from seven independent mapping experiments (1950 individuals) using a marker-merging method., Results: The composite map totalled 1107 cM and contained 4101 markers; comprising 3880 DArT, 213 microsatellite and eight candidate genes. Eighty-one DArT markers were mapped to two or more linkage groups, resulting in the 4101 markers being mapped to 4191 map positions. Approximately 13% of DArT markers mapped to identical map positions, thus the composite map contained 3634 unique loci at an average interval of 0.31 cM., Conclusion: The composite map represents the most saturated linkage map yet produced in Eucalyptus. As the majority of DArT markers contained on the map have been sequenced, the map provides a direct link to the E. grandis genome sequence and will serve as an important reference for progressing eucalypt research.

Diversity Arrays Technology (DArT) provides a robust, high throughput, cost-effective method to query thousands of sequence polymorphisms in a single assay. Despite the extensive use of this genotyping platform for numerous plant species, little is known regarding the sequence attributes and genome-wide distribution of DArT markers. We investigated the genomic properties of the 7,680 DArT marker probes of a Eucalyptus array, by sequencing them, constructing a high density linkage map and carrying out detailed physical mapping analyses to the Eucalyptus grandis reference genome. A consensus linkage map with 2,274 DArT markers anchored to 210 microsatellites and a framework map, with improved support for ordering, displayed extensive collinearity with the genome sequence. Only 1.4 Mbp of the 75 Mbp of still unplaced scaffold sequence was captured by 45 linkage mapped but physically unaligned markers to the 11 main Eucalyptus pseudochromosomes, providing compelling evidence for the quality and completeness of the current Eucalyptus genome assembly. A highly significant correspondence was found between the locations of DArT markers and predicted gene models, while most of the 89 DArT probes unaligned to the genome correspond to sequences likely absent in E. grandis, consistent with the pan-genomic feature of this multi-Eucalyptus species DArT array. These comprehensive linkage-to-physical mapping analyses provide novel data regarding the genomic attributes of DArT markers in plant genomes in general and for Eucalyptus in particular. DArT markers preferentially target the gene space and display a largely homogeneous distribution across the genome, thereby providing superb coverage for mapping and genome-wide applications in breeding and diversity studies. Data reported on these ubiquitous properties of DArT markers will be particularly valuable to researchers working on less-studied crop species who already count on DArT genotyping arrays but for which no reference genome is yet available to allow such detailed characterization.

A set of over 8000 Diversity Arrays Technology (DArT) markers was tested for its utility in high-resolution population and phylogenetic studies across a range of Eucalyptus taxa. Small-scale population studies of Eucalyptus camaldulensis, Eucalyptus cladocalyx, Eucalyptus globulus, Eucalyptus grandis, Eucalyptus nitens, Eucalyptus pilularis and Eucalyptus urophylla demonstrated the potential of genome-wide genotyping with DArT markers to differentiate species, to identify interspecific hybrids and to resolve biogeographic disjunctions within species. The population genetic studies resolved geographically partitioned clusters in E. camaldulensis, E. cladocalyx, E. globulus and E. urophylla that were congruent with previous molecular studies. A phylogenetic study of 94 eucalypt species provided results that were largely congruent with traditional taxonomy and ITS-based phylogenies, but provided more resolution within major clades than had been obtained previously. Ascertainment bias (the bias introduced in a phylogeny from using markers developed in a small sample of the taxa that are being studied) was not detected. DArT offers an unprecedented level of resolution for population genetic, phylogenetic and evolutionary studies across the full range of Eucalyptus species., (Copyright © 2011. Published by Elsevier Inc.)

Background and Aims: One of the major factors affecting the outcrossing rate in Eucalyptus globulus is thought to be the inherent self-incompatibility (SI) level of the female tree. SI in this species is mainly due to late-acting pre- and post-zygotic mechanisms operating in the ovary, and not S alleles. This study aimed to assess the phenotypic variation in SI levels within E. globulus and determine its genetic control and stability across pollination techniques, sites and seasons., Methods: SI levels were estimated for 105 genotypes originating from across the geographical range of E. globulus over multiple years of crossing. Separate grafted trees of some genotypes growing at the same and different sites allowed the genetic basis of the variation in SI to be tested and its stability across sites and seasons to be determined. The SI level of a tree was measured as the relative reduction in seeds obtained per flower pollinated following selfing compared with outcross pollinations. Thus, if seed set is the same, SI is 0 %, and if no self seed is set, SI is 100 %., Key Results: The average SI in E. globulus was 91 % and genotypes ranged from 8 to 100 % SI. Most genotypes (>75 %) had SI levels >90 %. There were highly significant differences between genotypes and the within-site broad-sense heritability of percentage SI was high (H(2) = 0.80 +/- 0.13). However, there was evidence that growing site, and to a lesser extent season, can affect the expression of SI levels. Trees with low reproductive loads produced relatively more seed from selfed flowers., Conclusions: There is a strong genetic basis to the phenotypic variation in SI in E. globulus within a site. However, the level of SI was affected, but to a lesser extent, by the environment, which in part may reflect the higher probability of selfed zygotes surviving on sites or in seasons where competition for resources is less.

While habitat fragmentation is a central issue in forest conservation studies in the face of broad-scale anthropogenic changes to the environment, its effects on contemporary mating patterns remain controversial. This is partly because of the inherent variation in mating patterns which may exist within species and the fact that few studies have replication at the landscape level. To study the effect of forest fragmentation on contemporary mating patterns, including effective pollen dispersal, we compared four native populations of the Australian forest tree, Eucalyptus globulus. We used six microsatellite markers to genotype 1289 open-pollinated offspring from paired fragmented and continuous populations on the island of Tasmania and in Victoria on mainland Australia. The mating patterns in the two continuous populations were similar, despite large differences in population density. In contrast, the two fragmented populations were variable and idiosyncratic in their mating patterns, particularly in their pollen dispersal kernels. The continuous populations showed relatively high outcrossing rates (86-89%) and low correlated paternity (0.03-0.06) compared with the fragmented populations (65-79% and 0.12-0.20 respectively). A greater proportion of trees contributed to reproduction in the fragmented (de/d>or= 0.5) compared with the continuous populations (de/d = 0.03-0.04). Despite significant inbreeding in the offspring of the fragmented populations, there was little evidence of loss of genetic diversity. It is argued that enhanced medium- and long-distance dispersal in fragmented landscapes may act to partly buffer the remnant populations from the negative effects of inbreeding and drift.

Foliar fungal pathogens from the genus Mycosphaerella affect eucalyptus in natural forests and plantations worldwide. QTL analysis was conducted to dissect the genetic control of resistance in Eucalyptus globulus to a natural infection by Mycosphaerella leaf disease, using a clonally replicated outbred F2 family (112 genotypes) planted in a field trial. Two major QTL, with high LOD support (20.2 and 10.9) and high genomewide significance, explained a large proportion (52%) of the phenotypic variance in the severity of damage by Mycosphaerella cryptica, which may be indicative of oligogenic control. Both QTL were validated in a second F2 family and one was validated in a third F2 family. The mean values of different genotype classes at both major QTL argue for Mendelian inheritance with resistance dominant over susceptibility. There were strong correlations between the levels of Mycosphaerella damage in related genetic material planted in three widely separated locations in Tasmania. These findings together provide evidence that the genes controlling resistance to Mycosphaerella damage are stable in different genetic backgrounds and across different environments.

Visualizing the pattern of variation using microsatellites within a Eucalyptus globulus forest on the island of Tasmania provided surprising insights into the complex nature of the fine-scale spatial genetic structure that resides in these forests. We used spatial autocorrelation and principal coordinate analysis to compare fine-scale genetic structure between juvenile and mature cohorts in a study area, 140 m in diameter, located within a typical, continuous E. globulus forest. In total, 115 juvenile and 168 mature individuals were genotyped with eight highly polymorphic microsatellite loci. There was no significant difference in the level of genetic diversity between cohorts. However, there were differences in the spatial distribution of the genetic variation. Autocorrelation analysis provided clear evidence for significant spatial genetic structure in the mature cohort and significant, but weaker, structure in the juvenile cohort. The spatial interpolation of principal coordinate axes, derived from ordination of the genetic distance matrix between individuals, revealed a spatially coherent family group which was evident in both cohorts. Direct comparison of the genetic structure within each cohort allowed visualization of a shift in the spatial distribution of genetic variation within the population of approximately 10 m. As the shift coincided with the direction of prevailing winds, it is hypothesized that this phenomenon is due to downwind dispersal of seeds and is indicative of the important role of prevailing winds in forcing eastward gene flow in these high-latitude forests.

Three small populations of a dwarf ecotype of the forest tree Eucalyptus globulus are found on exposed granite headlands in south-eastern Australia. These populations are separated by at least 100 km. Here, we used 12 nuclear microsatellites and a chloroplast DNA marker to investigate the genetic affinities of the dwarf populations to one another and to their nearest populations of tall E. globulus. Cape Tourville was studied in greater detail to assess the processes enabling the maintenance of distinct ecotypes in close geographical proximity. The three dwarf populations were not related to one another and were more closely related to adjacent tall trees than to one another. At Cape Tourville the dwarf and tall ecotypes were significantly differentiated in microsatellites and in chloroplast DNA. The dwarf and tall populations differed in flowering time and no evidence of pollen dispersal from the more extensive tall to the dwarf population was found. The three dwarf populations have evolved in parallel from the local tall ecotypes. This study shows that small marginal populations of eucalypts are capable of developing reproductive isolation from nearby larger populations through differences in flowering time and/or minor spatial separation, making parapatric speciation possible.

Low copy number nuclear genes have been found to be useful for phylogenetic reconstruction at different taxonomic levels. This study investigated the utility of a single copy gene, cinnamoyl CoA reductase (CCR), for resolving phylogenetic relationships at the sectional level within Eucalyptus. The monophyly of sections Exsertaria and Latoangulatae was explored, using section Maidenaria as an outgroup, and the impact of intragenic recombination on phylogenetic reconstruction examined. Phylogenetic analysis did not resolve monophyletic groups. Latoangulatae and Maidenaria were polyphyletic or paraphyletic. Exsertaria species formed a clade but included a single Latoangulatae species (E. major). Recombination analysis identified two intragenic recombination events that involved species from different sections, which have probably been facilitated by inter-sectional hybridisation. One of the events most likely occurred prior to speciation, with several Latoangulatae species having the recombinant allele. The other event may have occurred after speciation, since only one of two E. globulus samples possessed the recombinant allele. This is the first detailed report of intragenic recombination in both CCR and Eucalyptus, and between species of different sections of a plant genus. The occurrence of intragenic recombination may explain the anomalous positions of some species within the phylogenetic tree, and indicates that phylogenetic analysis of Eucalyptus using nuclear genes will be problematic unless recombination is taken into account.

Eucalyptus L'Hérit. is a genus comprised of more than 700 species that is of vital importance ecologically to Australia and to the forestry industry world-wide, being grown in plantations for the production of solid wood products as well as pulp for paper. With the sequencing of the genomes of Arabidopsis thaliana and Oryza sativa and the recent completion of the first tree genome sequence, Populus trichocarpa, attention has turned to the current status of genomic research in Eucalyptus. For several eucalypt species, large segregating families have been established, high-resolution genetic maps constructed and large EST databases generated. Collaborative efforts have been initiated for the integration of diverse genomic projects and will provide the framework for future research including exploiting the sequence of the entire eucalypt genome which is currently being sequenced. This review summarises the current position of genomic research in Eucalyptus and discusses the direction of future research.

The process of genetic assimilation of rare species by hybridizing congeners has been documented in a number of plant genera. This raises the possibility that some of the genetic diversity found in phylogeographical studies of widespread species has been acquired through hybridization with species that are now rare or extinct. In this fine-scale phylogeographical analysis, we show that a rare eucalypt species is leaving its trace in the chloroplast genome of a more abundant congener. The heart-leafed silver gum, Eucalyptus cordata, is a rare endemic of south-eastern Tasmania. Its populations are scattered amidst populations of more abundant related species, including the Tasmanian blue gum, Eucalyptus globulus. Using 339 samples from across the full range of both species, we compared chloroplast (cp) DNA haplotype phylogeography in E. globulus and E. cordata. The genealogy and distribution of chloroplast haplotypes suggest that E. globulus has acquired cpDNA from E. cordata in at least four different mixed populations. Shared haplotypes are highest in E. globulus sampled within 2 km of known E. cordata populations and drop to zero at a distance of 25 km from the nearest known E. cordata population. Localized haplotype sharing occurs in the absence of obvious hybrid zones or locally shared nuclear ribosomal DNA sequences. Given that the future loss of E. cordata from some mixed populations is likely, these findings indicate that phylogeographical analyses of organellar DNA should consider the possibility of introgression, even from species that have been eliminated from the sites of interest.

Tasmania is a natural laboratory for investigating the evolutionary processes of the Quaternary. It is a large island lying 40-44 degrees S, which was repeatedly glaciated and linked to southeastern continental Australia during the Quaternary. Climate change promoted both the isolation of species in glacial refugia, and an exchange between Tasmanian and mainland floras. Eucalyptus is a complex and diverse genus, which has increased in abundance in Australia over the past 100 kyr, probably in response to higher fire frequency. Morphological evidence suggests that gene flow may have occurred between many eucalypt species after changes in their distribution during the Quaternary. This paper summarizes recent genetic evidence for migration and introgressive hybridization in Tasmanian Eucalyptus. Maternally inherited chloroplast DNA reveals a long-term persistence of eucalypts in southeastern Tasmanian refugia, coupled with introgressive hybridization involving many species. Detailed analysis of the widespread species Eucalyptus globulus suggests that migration from mainland Australia was followed by introgression involving a rare Tasmanian endemic. The data support the hypothesis that changes in distribution of interfertile species during the Quaternary have promoted reticulate evolution in Eucalyptus.

There is increasing interest in the phenotypic plasticity of tree species to variation in climate as it may affect the economic value of current and future plantations. This study applied ensemble learning methods to explore the plastic response of Eucalyptus nitens pulpwood selection traits of growth (DBH, diameter at 1.3 m height), wood density and Kraft pulp yield to variation in elevation, geography and associated climate variables. To help explain the pulp yield response, we also modelled underlying biological traits—cellulose, lignin and extractives. The study was based on data from 84 harvest-age plots of common genetic origin across the pulpwood plantation estate in north-western Tasmania, Australia. DBH and wood density were obtained from resistance profile traces, collected on standing trees using a drilling resistance tool. In addition, outerwood cores were taken at 1.3 m stem height for (i) calibrating the resistance measures for wood density, and (ii) for assessment of pulp yield and wood chemistry based on near infrared spectroscopy. Modelling of the variation in plot means using the random forest algorithms showed growth and wood properties were influenced by the growing period climate of the plot. Of the climate variables studied, growth was mainly influenced by temperature, while wood density was mainly affected by rainfall-related variables. Wood density varied independently of growth and decreased with increasing annual rainfall and elevation of the plots. Pulp yield had the poorest fit statistics, it was influenced by a mix of climatic and geographic variables, and appeared independent of variation in growth and wood density. The plot variation in pulp yield was best explained by the modelled trends in the underlying biological traits of cellulose and lignin. Using these models, the plastic response of the key pulpwood traits to climate was mapped across the E. nitens plantation estate to help predict current plantation attributes and guide future choices of sites for plantation establishment. [ABSTRACT FROM AUTHOR]

Climatic changes during the Pleistocene were responsible for dramatic redistributions of plant species worldwide. On the rugged southern hemisphere island of Tasmania, temperature increases following the last glaciation saw upslope migration of climatically suitable species from lowland refugia and the expansion of eucalypt-dominated forests and woodlands in the Central Highlands. We integrate multiple lines of evidence (chloroplast and nuclear DNA markers, seedling morphology, and survival in common garden experiments) from a group of closely related endemic eucalypts (the alpine white gums) to argue that (i) the Central Highlands of the island were colonised by multiple glacial refugia with hybridisation among species and previously separated populations, and (ii) natural selection has filtered the admixed populations, resulting in local adaptation to the harsh sub-alpine environment. Chloroplast haplotype diversity decreased and nuclear microsatellite diversity increased with altitude, chloroplast sharing among taxa was common, and nuclear DNA differentiation of morphologically distinct taxa was lower in the Central Highlands compared with lowland regions. Local adaptation in the highlands was signalled by evidence from (i) a glasshouse trial in which directional selection (QST > FST) had shaped seedling morphological trait variation and (ii) population survival differences in 35-year-old reciprocal plantings along the major environmental gradients. We conclude that the evolutionary response of these island endemic trees to past climate change has involved the interplay of both hybridisation and natural selection, highlighting the importance of maintaining species interactions under future climate change. [ABSTRACT FROM AUTHOR]

In plants where seed dispersal is limited compared with pollen dispersal, hybridisation may enhance gene exchange and species dispersal. We provide genetic evidence of hybridisation contributing to the expansion of the rare Eucalyptus risdonii into the range of the widespread Eucalyptus amygdalina. These closely related tree species are morphologically distinct, and observations suggest that natural hybrids occur along their distribution boundaries and as isolated trees or in small patches within the range of E. amygdalina. Hybrid phenotypes occur outside the range of normal dispersal for E. risdonii seed, yet in some hybrid patches small individuals resembling E. risdonii occur and are hypothesised to be a result of backcrossing. Using 3362 genome‐wide SNPs assessed from 97 individuals of E. risdonii and E. amygdalina and 171 hybrid trees, we show that (i) isolated hybrids match the genotypes expected of F1/F2 hybrids, (ii) there is a continuum in the genetic composition among the isolated hybrid patches from patches dominated by F1/F2‐like genotypes to those dominated by E. risdonii‐backcross genotypes, and (iii) the E. risdonii‐like phenotypes in the isolated hybrid patches are most‐closely related to proximal larger hybrids. These results suggest that the E. risdonii phenotype has been resurrected in isolated hybrid patches established from pollen dispersal, providing the first steps in its invasion of suitable habitat by long‐distance pollen dispersal and complete introgressive displacement of E. amygdalina. Such expansion accords with the population demographics, common garden performance data, and climate modelling which favours E. risdonii and highlights a role of interspecific hybridisation in climate change adaptation and species expansion. [ABSTRACT FROM AUTHOR]

To understand the potential of forests to adapt to wildfire, we studied the genetic architecture of fire-related structural, damage and recovery traits in a globally important Australian forest tree species, Eucalyptus globulus. Fourteen traits were evaluated in an outcrossed F2 population in a field trial in Tasmania, Australia, which was burnt by a wildfire 14 years after planting. The trial also included open-pollinated families of the grandparental dwarf and tall ecotypes used to produce the F2 population. We studied the phenotypic correlations within the F2 population and performed quantitative trait loci (QTL) analyses using a linkage map comprised of 472 markers. Ecotype comparisons revealed that almost all traits were under genetic control, with trees of the dwarf ecotype significantly more damaged and mainly recovering from lignotubers, whereas tall ecotype trees mainly recovered from epicormic resprouts extending for a variable height up the stem. Within the F2, tree size was negatively correlated with fire damage and positively correlated with recovery. Genetic control of fire-related traits was confirmed by the detection of 38 QTL in the F2 population. These QTL accounted for 4 to 43% of the phenotypic variation in these traits. Several QTL co-located and likely reflect pleiotropic effects. However, many independent QTL were detected, including QTL for crown consumption and trunk scorch, epicormic resprouting, resprout herbivory, and seedling establishment. The QTL detected argue that many genetically controlled mechanisms are responsible for variation in fire damage and recovery. [ABSTRACT FROM AUTHOR]

With climate change impacting trees worldwide, enhancing adaptation capacity has become an important goal of provenance translocation strategies for forestry, ecological renovation, and biodiversity conservation. Given that not every species can be studied in detail, it is important to understand the extent to which climate adaptation patterns can be generalised across species, in terms of the selective agents and traits involved. We here compare patterns of genetic-based population (co)variation in leaf economic and hydraulic traits, climate–trait associations, and genomic differentiation of two widespread tree species (Eucalyptus pauciflora and E. ovata). We studied 2-year-old trees growing in a common-garden trial established with progeny from populations of both species, pair-sampled from 22 localities across their overlapping native distribution in Tasmania, Australia. Despite originating from the same climatic gradients, the species differed in their levels of population variance and trait covariance, patterns of population variation within each species were uncorrelated, and the species had different climate–trait associations. Further, the pattern of genomic differentiation among populations was uncorrelated between species, and population differentiation in leaf traits was mostly uncorrelated with genomic differentiation. We discuss hypotheses to explain this decoupling of patterns and propose that the choice of seed provenances for climate-based plantings needs to account for multiple dimensions of climate change unless species-specific information is available. [ABSTRACT FROM AUTHOR]

Understanding the capacity of forest tree species to adapt to climate change is of increasing importance for managing forest genetic resources. Through a genomics approach, we modelled spatial variation in climate adaptation within the Australian temperate forest tree Eucalyptus globulus, identified putative climate drivers of this genomic variation, and predicted locations of future climate refugia and populations at-risk of future maladaptation. Using 812,158 SNPs across 130 individuals from 30 populations (i.e., localities) spanning the species' natural range, a gradientForest algorithm found 1177 SNPs associated with locality variation in home-site climate (climate-SNPs), putatively linking them to climate adaptation. Very few climate-SNPs were associated with population-level variation in drought susceptibility, signalling the multi-faceted nature and complexity of climate adaptation. Redundancy analysis (RDA) showed 24% of the climate-SNP variation could be explained by annual precipitation, isothermality, and maximum temperature of the warmest month. Spatial predictions of the RDA climate vectors associated with climate-SNPs allowed mapping of genomically informed climate selective surfaces across the species' range under contemporary and projected future climates. These surfaces suggest over 50% of the current distribution of E. globulus will be outside the modelled adaptive range by 2070 and at risk of climate maladaptation. Such surfaces present a new integrated approach for natural resource managers to capture adaptive genetic variation and plan translocations in the face of climate change. [ABSTRACT FROM AUTHOR]

Summary: We overview an array of common garden genetics trials established across multiple sites and embedded in restoration plantings in the harsh, modified Midlands region of the island of Tasmania, Australia. The steps in establishing the common gardens, the diversity of experimental designs employed and their objectives are described. These trials provide a long‐term framework to test species choices and provenancing strategies, as well as enhance our understanding of adaptive genetic variation within key eucalypt species used for ecological restoration. Early results signal genetic‐based differences in early‐age performance, and we showcase the multiple benefits of integrating research infrastructure into restoration plantings. [ABSTRACT FROM AUTHOR]

Microfibril angle (MFA) is a key biological trait contributing to wood stiffness, which is a common breeding objective for solid wood products in many tree species. To explore its genetic architecture, area-weighted MFA was measured in two Eucalyptus nitens progeny trials in Tasmania, Australia, with common open-pollinated families. Radial strips were extracted from 823 trees in 131 families and MFA assessed using SilviScan-2®. Heritability, genotype-by-environment interaction and inter-trait genetic correlations were evaluated to examine the genetic variability and stability of MFA and its relationships with other solid wood and pulpwood selection traits. Significant family variation was found for MFA in both trials. There was no significant genotype-by-environment interaction and the across-site narrow-sense heritability was 0.27. MFA was genetically independent of basic density, growth, and tree form. However, MFA was strongly and favourable genetically correlated to acoustic wave velocity in standing trees, modulus of elasticity and kraft pulp yield (KPY). The present study has shown that genetic improvement of E. nitens for pulpwood selection traits is unlikely to have adversely affected MFA, and thus timber stiffness. Rather these results suggest the possibility that selection for increased KPY may have indirectly improved MFA favourably for solid wood products. [ABSTRACT FROM AUTHOR]

Eucalyptus globulus is widely grown for pulpwood production in temperate regions of the world. However, there is increasing interest in using it for solid-wood products. We studied the genetic architecture of key pulpwood and solid-wood selection traits using two E. globulus progeny trials in a high rainfall area (wet) of Tasmania and a previously studied trial in a low rainfall area (dry). These trials were established using open-pollinated families from native trees sampled from 13 subraces. We assessed traits in common [diameter at breast height (DBH) and wood basic density (BD)], and specific to pulpwood [Kraft pulp yield (KPY)] and solid-wood [stem straightness and acoustic wave velocity (AWV)] breeding. Significant genetic variation was found for all traits. Strong GxE was detected for DBH across the wet and dry sites, but little for BD and KPY. At the wet sites we show a positive genetic correlation between DBH and KPY, but not between DBH and BD. Subrace and family within subrace correlations between KPY and BD were significant but in opposite directions. We confirm previous reports of significant positive genetic and phenotypic correlations between KPY and AWV. A positive genetic correlation between stem straightness and DBH was detected, and subraces with straighter stems tended to have higher KPY. In general, correlations between most solid-wood and pulpwood traits were favourable, suggesting that past selection for pulpwood traits had neutral or favourable effects on many key solid-wood traits. We conclude that breeding for solid-wood and pulpwood are relatively compatible. [ABSTRACT FROM AUTHOR]

Anthropogenic change (climate change and habitat fragmentation) is driving a growing view that local seed collections may need to be supplemented with nonlocal seed as a strategy to bolster genetic diversity and thus increase evolutionary potential of plantings. While this strategy is becoming widely promoted, empirical support is limited, and there is a lack of accessible research tools to assist in its experimental testing. We therefore provide the Provenancing Using Climate Analogues ( PUCA) framework that integrates the principles of the climate-adjusted provenancing strategy with concepts from population genetics (i.e. potential inbreeding in small fragmented populations) as both a research and operational-ready tool to guide the collection of nonlocal seed. We demonstrate the application of PUCA using the Midlands of Tasmania, Australia, a region that is currently undergoing large-scale ecological restoration. We highlight multiple nonlocal seed sources for testing by identifying actual species distribution records that currently occupy environments similar to that projected to occur at the restoration site in the future. We discuss the assumptions of PUCA and the ecological considerations that need to be tested when moving nonlocal genotypes across the landscape. [ABSTRACT FROM AUTHOR]

As climates change, restoration programmes need to maximize the capacity of regenerating ecosystems to adapt to new environments, particularly when planting long-lived trees. By using a combined phenotypic and genomic approach, we assessed evidence of adaptation capacity in Eucalyptus loxophleba ssp. lissophloia, a mallee eucalypt important for restoration and oil production in Western Australia. Assessment of leaf traits in nine wild populations across a rainfall gradient showed that two traits were correlated with long-term moisture availability. Populations in more arid environments had lower specific leaf area and lower stomatal conductance, consistent with a stable plastic or adaptive response. Other leaf size and shape traits were correlated with short-term climate variables, suggesting a dynamic plastic response. Genome-wide scans with 4851 DArTseq markers and outlier analysis detected 50 markers showing signals of disruptive selection, consistent with local adaptation. Sixteen markers showed allele frequencies correlated with aridity; three were also associated with differences in stomatal conductance. Multivariate analysis and markerenvironment associations signalled secondary directions of adaptation associated with maximum temperatures and soil phosphorus. We suggest that adaptation and acclimation are both likely determinants of functional phenotype in E. loxophleba, and argue that aridity is a critical driver of adaptation in this widespread species. [ABSTRACT FROM AUTHOR]

We studied the evolutionary processes shaping the genetic diversity in the naturally fragmented Eucalyptus cordata, a rare homoblastic tree endemic to the island of Tasmania. A genome-wide scan showed that E. cordata and the endangered heteroblastic E. morrisbyi were closely related, suggesting a neotenous origin of E. cordata from an endemic heteroblastic ancestor. Bayesian cluster analysis based on nuclear microsatellites assayed in 567 E. cordata and E. morrisbyi individuals revealed five genetic clusters. Two clusters comprised populations that correspond to putative ancestral gene pools linking E. cordata and E. morrisbyi. Another cluster included populations that transgressed the drowned Derwent River valley, suggestive of a wider glacial distribution. However, the majority of individuals occurred in the two genetic clusters distributed in the south-west and north-east of the range of E. cordata. The elevated genetic diversity in populations comprising these clusters suggests that they represent two recently fragmented cores of the distribution. Genetic evidence suggests that the newly described, localised E. cordata subspecies quadrangulosa has been recently selected from within the morphologically diverse, south-western cluster. We argue that multiple phases of isolation and drift have led to the contemporary pattern of molecular variation and the scattering of relictual and more recently derived populations across the species distribution. [ABSTRACT FROM AUTHOR]

Abstract: Hybridisation through pollen dispersal from exotic plants is increasingly recognised as a threat to the genetic integrity of native plant populations. Its genetic impact can be greater in rare taxa, due to their vulnerability to pollen swamping by more abundant congeners. We assessed the likelihood of pollen dispersal from exotic eucalypt plantations into all of Australia''s rare native eucalypts, and conducted a case study of Eucalyptus perriniana, which is rare in Tasmania. The Australia-wide study involved spatial analyses of the locations for each rare species superimposed on distributions of eucalypt plantations, which were combined with known taxonomically based reproductive barriers. Of the 74 nationally listed rare eucalypt taxa, 22 had locations within 10km of plantations of the same genus, and eight were within 1km. These eight proximal taxa are considered priorities for monitoring. In the most extreme case, 30% of point locations originating from herbarium records and field surveys for Eucalyptus conglomerata were within 1km of exotic plantations. In the case study, E. perriniana revealed considerable reproductive compatibility with adjacent recently established Eucalyptus nitens plantations. However, F1 hybridisation between these species was limited, with 0.2% of the 18,625 seedlings grown from 100 single-tree open-pollinated seedlots being hybrids. For now, the probability of exotic gene flow into E. perriniana appears to be low, however this probability is likely to increase as more E. nitens flowers in the surrounding landscape. These studies suggest that understanding the breeding system and biology of these populations may reveal surprising resistance to such exotic hybridisation as well as identifying high risk situations to focus conservation management. [Copyright &y& Elsevier]