Biodiscoveries within the Australian plant genus Eremophila based on international and interdisciplinary collaboration: results and perspectives on outstanding ethical dilemmas.

In a cross-continental research initiative, including researchers working in Australia and Denmark, and based on joint external funding by a 3-year grant from the Novo Nordisk Foundation, we have used DNA sequencing, extensive chemical profiling and molecular networking analyses across the entire Eremophila genus to provide new knowledge on the presence of natural products and their bioactivities using polypharmocological screens. Sesquiterpenoids, diterpenoids and dimers of branched-chain fatty acids with previously unknown chemical structures were identified. The collection of plant material from the Eremophila genus was carried out according to a 'bioprospecting agreement' with the Government of Western Australia. We recognize that several Eremophila species hold immense cultural significance to Australia's First Peoples. In spite of our best intentions to ensure that new knowledge gained about the genus Eremophila and any potential future benefits are shared in an equitable manner, in accordance with the Nagoya Protocol, we encounter serious dilemmas and potential conflicts in making benefit sharing with Australia's First Peoples a reality.

Phylogenomics reveals extreme gene tree discordance in a lineage of dominant trees: hybridization, introgression, and incomplete lineage sorting blur deep evolutionary relationships despite clear species groupings in Eucalyptus subgenus Eudesmia.

Eucalypts are a large and ecologically important group of plants on the Australian continent, and understanding their evolution is important in understanding evolution of the unique Australian flora. Previous phylogenies using plastome DNA, nuclear-ribosomal DNA, or random genome-wide SNPs, have been confounded by limited genetic sampling or by idiosyncratic biological features of the eucalypts, including widespread plastome introgression. Here we present phylogenetic analyses of Eucalyptus subgenus Eudesmia (22 species from western, northern, central and eastern Australia), in the first study to apply a target-capture sequencing approach using custom, eucalypt-specific baits (of 568 genes) to a lineage of Eucalyptus. Multiple accessions of all species were included, and target-capture data were supplemented by separate analyses of plastome genes (average of 63 genes per sample). Analyses revealed a complex evolutionary history likely shaped by incomplete lineage sorting and hybridization. Gene tree discordance generally increased with phylogenetic depth. Species, or groups of species, toward the tips of the tree are mostly supported, and three major clades are identified, but the branching order of these clades cannot be confirmed with confidence. Multiple approaches to filtering the nuclear dataset, by removing genes or samples, could not reduce gene tree conflict or resolve these relationships. Despite inherent complexities in eucalypt evolution, the custom bait kit devised for this research will be a powerful tool for investigating the evolutionary history of eucalypts more broadly.

Navigating through chemical space and evolutionary time across the Australian continent in plant genus Eremophila.

Eremophila is the largest genus in the plant tribe Myoporeae (Scrophulariaceae) and exhibits incredible morphological diversity across the Australian continent. The Australian Aboriginal Peoples recognize many Eremophila species as important sources of traditional medicine, the most frequently used plant parts being the leaves. Recent phylogenetic studies have revealed complex evolutionary relationships between Eremophila and related genera in the tribe. Unique and structurally diverse metabolites, particularly diterpenoids, are also a feature of plants in this group. To assess the full dimension of the chemical space of the tribe Myoporeae, we investigated the metabolite diversity in a chemo-evolutionary framework applying a combination of molecular phylogenetic and state-of-the-art computational metabolomics tools to build a dataset involving leaf samples from a total of 291 specimens of Eremophila and allied genera. The chemo-evolutionary relationships are expounded into a systematic context by integration of information about leaf morphology (resin and hairiness), environmental factors (pollination and geographical distribution), and medicinal properties (traditional medicinal uses and antibacterial studies), augmenting our understanding of complex interactions in biological systems.

Unique and highly specific cyanogenic glycoside localization in stigmatic cells and pollen in the genus Lomatia (Proteaceae).

BACKGROUND AND AIMS: Floral chemical defence strategies remain understudied despite the significance of flowers to plant fitness, and the fact that many flowers contain secondary metabolites that confer resistance to herbivores. Optimal defence and apparency theories predict that the most apparent plant parts and/or those most important to fitness should be most defended. To test whether within-flower distributions of chemical defence are consistent with these theories we used cyanogenic glycosides (CNglycs), which are constitutive defence metabolites that deter herbivores by releasing hydrogen cyanide upon hydrolysis. METHODS: We used cyanogenic florets of the genus Lomatia to investigate at what scale there may be strategic allocation of CNglycs in flowers, what their localization reveals about function, and whether levels of floral CNglycs differ between eight congeneric species across a climatic gradient. Within-flower distributions of CNglycs during development were quantified, CNglycs were identified and their localization was visualized in cryosectioned florets using matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI). KEY RESULTS: Florets of all congeneric species studied were cyanogenic, and concentrations differed between species. Within florets there was substantial variation in CNglyc concentrations, with extremely high concentrations (up to 14.6 mg CN g-1 d. wt) in pollen and loose, specialized surface cells on the pollen presenter, among the highest concentrations reported in plant tissues. Two tyrosine-derived CNglycs, the monoglycoside dhurrin and diglycoside proteacin, were identified. MALDI-MSI revealed their varying ratios in different floral tissues; proteacin was primarily localized to anthers and ovules, and dhurrin to specialized cells on the pollen presenter. The mix of transient specialized cells and pollen of L. fraxinifolia was ~11 % dhurrin and ~1.1 % proteacin by mass. CONCLUSIONS: Tissue-specific distributions of two CNglycs and substantial variation in their concentrations within florets suggests their allocation is under strong selection. Localized, high CNglyc concentrations in transient cells challenge the predictions of defence theories, and highlight the importance of fine-scale metabolite visualization, and the need for further investigation into the ecological and metabolic roles of CNglycs in floral tissues.

The Roles of Ecological and Evolutionary Processes in Plant Community Assembly: The Environment, Hybridization, and Introgression Influence Co-occurrence of Eucalyptus.

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.

Chloroplast genome analysis of Australian eucalypts--Eucalyptus, Corymbia, Angophora, Allosyncarpia and Stockwellia (Myrtaceae).

We present a phylogenetic analysis and comparison of structural features of chloroplast genomes for 39 species of the eucalypt group (genera Eucalyptus, Corymbia, Angophora, and outgroups Allosyncarpia and Stockwellia). We use 41 complete chloroplast genome sequences, adding 39 finished-quality chloroplast genomes to two previously published genomes. Maximum parsimony and Bayesian analyses, based on >7000 variable nucleotide positions, produced one fully resolved phylogenetic tree (35 supported nodes, 27 with 100% bootstrap support). Eucalyptus and its sister lineage Angophora+Corymbia show a deep divergence. Within Eucalyptus, three lineages are resolved: the 'eudesmid', 'symphyomyrt' and 'monocalypt' groups. Corymbia is paraphyletic with respect to Angophora. Gene content and order do not vary among eucalypt chloroplasts; length mutations, especially frame shifts, are uncommon in protein-coding genes. Some non-synonymous mutations are highly incongruent with the overall phylogenetic signal, notably in rbcL, and may be adaptive. Application of custom informatics pipelines (GYDLE Inc.) enabled direct chloroplast genome assembly, resolving each genome to finished-quality with no need for PCR gap-filling or contig order resolution. Analysis of whole chloroplast genomes resolved major eucalypt clades and revealed variable regions of the genome that will be useful in lower-level genetic studies (including phylogeography and geneflow).

Testing the impact of calibration on molecular divergence times using a fossil-rich group: the case of Nothofagus (Fagales).

Although temporal calibration is widely recognized as critical for obtaining accurate divergence-time estimates using molecular dating methods, few studies have evaluated the variation resulting from different calibration strategies. Depending on the information available, researchers have often used primary calibrations from the fossil record or secondary calibrations from previous molecular dating studies. In analyses of flowering plants, primary calibration data can be obtained from macro- and mesofossils (e.g., leaves, flowers, and fruits) or microfossils (e.g., pollen). Fossil data can vary substantially in accuracy and precision, presenting a difficult choice when selecting appropriate calibrations. Here, we test the impact of eight plausible calibration scenarios for Nothofagus (Nothofagaceae, Fagales), a plant genus with a particularly rich and well-studied fossil record. To do so, we reviewed the phylogenetic placement and geochronology of 38 fossil taxa of Nothofagus and other Fagales, and we identified minimum age constraints for up to 18 nodes of the phylogeny of Fagales. Molecular dating analyses were conducted for each scenario using maximum likelihood (RAxML + r8s) and Bayesian (BEAST) approaches on sequence data from six regions of the chloroplast and nuclear genomes. Using either ingroup or outgroup constraints, or both, led to similar age estimates, except near strongly influential calibration nodes. Using "early but risky" fossil constraints in addition to "safe but late" constraints, or using assumptions of vicariance instead of fossil constraints, led to older age estimates. In contrast, using secondary calibration points yielded drastically younger age estimates. This empirical study highlights the critical influence of calibration on molecular dating analyses. Even in a best-case situation, with many thoroughly vetted fossils available, substantial uncertainties can remain in the estimates of divergence times. For example, our estimates for the crown group age of Nothofagus varied from 13 to 113 Ma across our full range of calibration scenarios. We suggest that increased background research should be made at all stages of the calibration process to reduce errors wherever possible, from verifying the geochronological data on the fossils to critical reassessment of their phylogenetic position.

Diverse organ-specific localisation of a chemical defence, cyanogenic glycosides, in flowers of eleven species of Proteaceae.

Floral chemical defence strategies remain under-investigated, despite the significance of flowers to plant fitness. We used cyanogenic glycosides (CNglycs)-constitutive secondary metabolites that deter herbivores by releasing hydrogen cyanide, but also play other metabolic roles-to ask whether more apparent floral tissues and those most important for fitness are more defended as predicted by optimal defence theories, and what fine-scale CNglyc localisation reveals about function(s)? Florets of eleven species from the Proteaceae family were dissected to quantitatively compare the distribution of CNglycs within flowers and investigate whether distributions vary with other floral/plant traits. CNglycs were identified and their localisation in florets was revealed by matrix-assisted laser desorption ionisation mass spectrometry imaging (MALDI-MSI). We identified extremely high CNglyc content in floral tissues of several species (>1% CN), highly tissue-specific CNglyc distributions within florets, and substantial interspecific differences in content distributions, not all consistent with optimal defence hypotheses. Four patterns of within-flower CNglyc allocation were identified: greater tissue-specific allocations to (1) anthers, (2) pedicel (and gynophore), (3) pollen presenter, and (4) a more even distribution among tissues with higher content in pistils. Allocation patterns were not correlated with other floral traits (e.g. colour) or taxonomic relatedness. MALDI-MSI identified differential localisation of two tyrosine-derived CNglycs, demonstrating the importance of visualising metabolite localisation, with the diglycoside proteacin in vascular tissues, and monoglycoside dhurrin across floral tissues. High CNglyc content, and diverse, specific within-flower localisations indicate allocations are adaptive, highlighting the importance of further research into the ecological and metabolic roles of floral CNglycs.

Phylogenomic analyses of Sapindales support new family relationships, rapid Mid-Cretaceous Hothouse diversification, and heterogeneous histories of gene duplication.

Sapindales is an angiosperm order of high economic and ecological value comprising nine families, c. 479 genera, and c. 6570 species. However, family and subfamily relationships in Sapindales remain unclear, making reconstruction of the order's spatio-temporal and morphological evolution difficult. In this study, we used Angiosperms353 target capture data to generate the most densely sampled phylogenetic trees of Sapindales to date, with 448 samples and c. 85% of genera represented. The percentage of paralogous loci and allele divergence was characterized across the phylogeny, which was time-calibrated using 29 rigorously assessed fossil calibrations. All families were supported as monophyletic. Two core family clades subdivide the order, the first comprising Kirkiaceae, Burseraceae, and Anacardiaceae, the second comprising Simaroubaceae, Meliaceae, and Rutaceae. Kirkiaceae is sister to Burseraceae and Anacardiaceae, and, contrary to current understanding, Simaroubaceae is sister to Meliaceae and Rutaceae. Sapindaceae is placed with Nitrariaceae and Biebersteiniaceae as sister to the core Sapindales families, but the relationships between these families remain unclear, likely due to their rapid and ancient diversification. Sapindales families emerged in rapid succession, coincident with the climatic change of the Mid-Cretaceous Hothouse event. Subfamily and tribal relationships within the major families need revision, particularly in Sapindaceae, Rutaceae and Meliaceae. Much of the difficulty in reconstructing relationships at this level may be caused by the prevalence of paralogous loci, particularly in Meliaceae and Rutaceae, that are likely indicative of ancient gene duplication events such as hybridization and polyploidization playing a role in the evolutionary history of these families. This study provides key insights into factors that may affect phylogenetic reconstructions in Sapindales across multiple scales, and provides a state-of-the-art phylogenetic framework for further research.

A box on the river: The phylogenetics and phylogeography of Eucalyptus baueriana (Eucalyptus sect. Adnataria ser. Heterophloiae).

We present a phylogeographic study of the tree species Eucalyptus baueriana Schauer, which occurs in disjunct areas on the near coastal plains and ranges of the south-east Australian mainland. DArTseq data are used to build a phylogeny including E. baueriana and closely related taxa to test its monophyly, test the genetic distinctness of the three subspecies of E. baueriana, and investigate relationships between its disjunct populations. Additionally, we use population structure analysis to investigate the genetic distinctness of populations, and MaxEnt to investigate the environmental factors potentially influencing the species' distribution. We show E. baueriana is monophyletic and most closely related to three other Blue Box eucalypt species: E. conica H.Deane & Maiden, E. dalveenica T.L.Collins, R.L.Andrew & J.J.Bruhl and E. magnificata L.A.S.Johnson & K.D.Hill, with some evidence for genetic introgression between these taxa. Within E. baueriana, the deepest genetic breaks do not correspond with the subspecies classification as the two geographically restricted subspecies, together with samples of the more widespread E. baueriana subsp. baueriana from west of the Gippsland lowlands, form a south-western clade with that is sister to other populations of subsp. baueriana. The oldest genetic break in the species occurs in far eastern Gippsland (Victoria), corresponding to one of the shortest geographic disjunctions in the species' distribution. Genetic breaks in other species have been observed in this region which is broadly referred to as the southern transition zone. Both total annual rainfall and the seasonality of this rainfall are hypothesised to affect the species' distribution; gaps in its distribution are in areas of higher rainfall that support closed forest and in regions with more winter dominated rainfall.

Phylogeny and classification of the Australasian and Indomalayan mimosoid legumes Archidendron and Archidendropsis (Leguminosae, subfamily Caesalpinioideae, mimosoid clade).

The morphologically variable genus Archidendron is the second largest mimosoid legume genus from the Indomalayan-Australasian region, yet it has not been well represented in phylogenetic studies. Phylogenies that have included multiple representatives of Archidendron suggest it may not be monophyletic, and the same applies to Archidendropsis, another understudied genus of the Archidendron clade. The most comprehensive phylogeny of Archidendron and Archidendropsis to date is presented, based on four nuclear markers (ITS, ETS, SHMT and RBPCO). Exemplars from all genera of the wider Archidendron clade are sampled, including representatives of all series within Archidendron and the two subgenera of Archidendropsis. Our results confirm that Archidendron and Archidendropsis are not monophyletic. Within Archidendron, only one series (ser. Ptenopae) is resolved as monophyletic and species of Archidendron are divided into two primarily geographic lineages. One clade is distributed in western Malesia and mainland Asia and includes most representatives of series Clypeariae, while the other is mostly restricted to eastern Malesia and Australia and includes representatives of the seven other series plus two samples of series Clypeariae. No taxonomic changes are made for Archidendron due to the high level of topological uncertainty and the lack of discrete macromorphological characters separating these two lineages. Each of the two subgenera of Archidendropsis is monophyletic but they are not closely related. A new genus endemic to Queensland (Australia), Heliodendron Gill.K. Br. & Bayly, gen. nov., is described for the former Archidendropsissubg.Basaltica, and combinations for its three species are proposed: Heliodendronbasalticum (F. Muell.) Gill.K. Br. & Bayly, comb. nov., Heliodendronthozetianum (F. Muell.) Gill.K. Br. & Bayly, comb. nov., and Heliodendronxanthoxylon (C.T. White & W.D. Francis) Gill.K. Br. & Bayly, comb. nov.

Divergent lineages in a semi-arid mallee species, Eucalyptus behriana, correspond to a major geographic break in southeastern Australia.

AIM: To infer relationships between populations of the semi-arid, mallee eucalypt, Eucalyptus behriana, to build hypotheses regarding evolution of major disjunctions in the species' distribution and to expand understanding of the biogeographical history of southeastern Australia. LOCATION: Southeastern Australia. TAXON: Eucalyptus behriana (Myrtaceae, Angiospermae). METHODS: We developed a large dataset of anonymous genomic loci for 97 samples from 11 populations of E. behriana using double digest restriction site-associated DNA sequencing (ddRAD-seq), to determine genetic relationships between the populations. These relationships, along with species distribution models, were used to construct hypotheses regarding environmental processes that have driven fragmentation of the species' distribution. RESULTS: Greatest genetic divergence was between populations on either side of the Lower Murray Basin. Populations west of the Basin showed greater genetic divergence between one another than the eastern populations. The most genetically distinct population in the east (Long Forest) was separated from others by the Great Dividing Range. A close relationship was found between the outlying northernmost population (near West Wyalong) and those in the Victorian Goldfields despite a large disjunction between them. CONCLUSIONS: Patterns of genetic variation are consistent with a history of vicariant differentiation of disjunct populations. We infer that an early disjunction to develop in the species distribution was that across the Lower Murray Basin, an important biogeographical barrier separating many dry sclerophyll plant taxa in southeastern Australia. Additionally, our results suggest that the western populations fragmented earlier than the eastern ones. Fragmentation, both west and east of the Murray Basin, is likely tied to climatic changes associated with glacial-interglacial cycles although it remains possible that major geological events including uplift of the Mount Lofty Ranges and basalt flows in the Newer Volcanics Province also played a role.

Divergent responses of above- and below-ground chemical defence to nitrogen and phosphorus supply in waratahs (Telopea speciosissima).

Plant nutrition can affect the allocation of resources to plant chemical defences, yet little is known about how phosphorus (P) supply, and relative nitrogen (N) and P supply, affect chemical defences, especially in species with intrinsically conservative nutrient use adapted to P-impoverished soils. Waratah (Telopea speciosissima (Sm.) R.Br.), like other Proteaceae, is adapted nutrient-poor soils. It was identified as having cyanogenic glycosides (CNglycs) throughout the plant. T. speciosissima seedlings were grown for 15 weeks under two N and P concentrations. CNglycs (N-based defence) and nutrients were quantified in above- and below-ground organs; foliar carbon (C)-based phenolics and tannins were also quantified. CNglyc concentrations in roots were on average 51-fold higher than in above-ground tissues and were affected by both N and P supply, whereas foliar CNglyc concentrations only responded to N supply. Leaves had high concentrations of C-based defences, which increased under low N, and were not correlated with N-based defences. Greater root chemical defence against herbivores and pathogens may be important in a non-mycorrhizal species that relies on basal resprouting following disturbance. The differing responses of secondary chemistry in above- and below-ground organs to P and N demonstrate the importance of broadening the predominantly foliar focus of plant defence studies.

The complete chloroplast genome sequence of the Australian Mirbelioid pea Platylobium obtusangulum Hook. (Leguminosae: subf. Papilionoideae, tribe Bossiaeeae).

We sequenced and assembled the whole chloroplast genome of the Australian-endemic shrub Platylobium obtusangulum. The total size of the genome is 150,090 base pairs (bp), including two inverted repeat regions of 25,511 bp each, one large single copy region of 80,567 bp and a small single copy region of 18,501 bp. The genome has a GC content of 36.7% and includes 127 annotated genes (83 protein coding, 36 tRNA genes and eight rRNA genes). Phylogenetic analysis of chloroplast genomes placed the Platylobium obtusangulum genome in the expected position of the Mirbelioid clade in the legume family (Leguminosae: Papilionoideae).

Annotated plastome of the temperate woody vine Muehlenbeckia australis (G.Forst.) Meisn. (Polygonaceae).

We assembled the plastome of the temperate, Southern Hemisphere liana Muehlenbeckia australis from high throughput sequencing data (paired-end Illumina reads) generated from total genomic DNA sequencing libraries. M. australis' chloroplast genome sequence (GenBank: MG604297) is 163,484 bp in length and composed of long single copy (LSC; 88,166 bp) and short single copy (SSC; 13,486 bp) regions flanked by inverted repeats (IR; 30,916 bp each) typical for angiosperms. The plastome includes 131 genes comprising 83 protein-coding genes, 37 transfer RNA genes, eight ribosomal RNA genes, two possible pseudogenes, psbL and rpl23 with internal stop codons, and truncated repeats of ndhF and rps19 at IR boundaries.

The complete chloroplast genome sequence of Spyridium parvifolium var. parvifolium (family Rhamnaceae; tribe Pomaderreae).

We assembled the complete chloroplast genome of the Australian shrub Spyridium parvifolium var. parvifolium. The genome was 161,012 bp in length, with a pair of inverted repeats (IRs) of 26,515 bp, separated by a large single copy (LSC) region of 88,814 bp and a small single copy region (SCC) of 19,168 bp. The GC content was 36.9%. In total, 130 genes were annotated, including 86 protein coding genes, 36 tRNA genes and 8 rRNA genes. Phylogenetic analysis of 56 chloroplast genes placed this genome of S. parvifolium var. parvifolium within the family Rhamnaceae.

Chloroplast variation is incongruent with classification of the Australian bloodwood eucalypts (genus Corymbia, family Myrtaceae).

Previous molecular phylogenetic analyses have resolved the Australian bloodwood eucalypt genus Corymbia (~100 species) as either monophyletic or paraphyletic with respect to Angophora (9-10 species). Here we assess relationships of Corymbia and Angophora using a large dataset of chloroplast DNA sequences (121,016 base pairs; from 90 accessions representing 55 Corymbia and 8 Angophora species, plus 33 accessions of related genera), skimmed from high throughput sequencing of genomic DNA, and compare results with new analyses of nuclear ITS sequences (119 accessions) from previous studies. Maximum likelihood and maximum parsimony analyses of cpDNA resolve well supported trees with most nodes having >95% bootstrap support. These trees strongly reject monophyly of Corymbia, its two subgenera (Corymbia and Blakella), most taxonomic sections (Abbreviatae, Maculatae, Naviculares, Septentrionales), and several species. ITS trees weakly indicate paraphyly of Corymbia (bootstrap support <50% for maximum likelihood, and 71% for parsimony), but are highly incongruent with the cpDNA analyses, in that they support monophyly of both subgenera and some taxonomic sections of Corymbia. The striking incongruence between cpDNA trees and both morphological taxonomy and ITS trees is attributed largely to chloroplast introgression between taxa, because of geographic sharing of chloroplast clades across taxonomic groups. Such introgression has been widely inferred in studies of the related genus Eucalyptus. This is the first report of its likely prevalence in Corymbia and Angophora, but this is consistent with previous morphological inferences of hybridisation between species. Our findings (based on continent-wide sampling) highlight a need for more focussed studies to assess the extent of hybridisation and introgression in the evolutionary history of these genera, and that critical testing of the classification of Corymbia and Angophora requires additional sequence data from nuclear genomes.

Major clades of Australasian Rutoideae (Rutaceae) based on rbcL and atpB sequences.

BACKGROUND: Rutaceae subfamily Rutoideae (46 genera, c. 660 species) is diverse in both rainforests and sclerophyll vegetation of Australasia. Australia and New Caledonia are centres of endemism with a number of genera and species distributed disjunctly between the two regions. Our aim was to generate a high-level molecular phylogeny for the Australasian Rutoideae and identify major clades as a framework for assessing morphological and biogeographic patterns and taxonomy. METHODOLOGY/PRINCIPAL FINDINGS: Phylogenetic analyses were based on chloroplast genes, rbcL and atpB, for 108 samples (78 new here), including 38 of 46 Australasian genera. Results were integrated with those from other molecular studies to produce a supertree for Rutaceae worldwide, including 115 of 154 genera. Australasian clades are poorly matched with existing tribal classifications, and genera Philotheca and Boronia are not monophyletic. Major sclerophyll lineages in Australia belong to two separate clades, each with an early divergence between rainforest and sclerophyll taxa. Dehiscent fruits with seeds ejected at maturity (often associated with myrmecochory) are inferred as ancestral; derived states include woody capsules with winged seeds, samaras, fleshy drupes, and retention and display of seeds in dehisced fruits (the last two states adaptations to bird dispersal, with multiple origins among rainforest genera). Patterns of relationship and levels of sequence divergence in some taxa, mostly species, with bird-dispersed (Acronychia, Sarcomelicope, Halfordia and Melicope) or winged (Flindersia) seeds are consistent with recent long-distance dispersal between Australia and New Caledonia. Other deeper Australian/New Caledonian divergences, some involving ant-dispersed taxa (e.g., Neoschmidia), suggest older vicariance. CONCLUSIONS/SIGNIFICANCE: This comprehensive molecular phylogeny of the Australasian Rutoideae gives a broad overview of the group's evolutionary and biogeographic history. Deficiencies of infrafamilial classifications of Rutoideae have long been recognised, and our results provide a basis for taxonomic revision and a necessary framework for more focused studies of genera and species.

Phylogeographic patterns in the Australasian genus Chionohebe (Veronica s.l., Plantaginaceae) based on AFLP and chloroplast DNA sequences.

The alpine genus Chionohebe is one of seven genera in the southern hemisphere Hebe complex. The main aims of this study were to infer the evolutionary relationships and assess phylogeographic patterns among the six species of Chionohebe, determine the origin of the two species with trans-Tasman distributions, and test species delimitations and specimen identifications based on morphology. Analyses of AFLP data recovered five major lineages within Chionohebe, some of which corresponded to species and varieties as currently circumscribed. Although the cushion chionohebes were strongly supported as monophyletic, the sole non-cushion species, C. densifolia, was sister to Parahebe trifida, and thus the AFLP data do not support a monophyletic Chionohebe as usually circumscribed. Strong north/south and west/east phylogeographic patterns were found among and within the main AFLP lineages in New Zealand. Analyses of chloroplast DNA (cpDNA) revealed eight haplotypes in Chionohebe, but these did not correspond to current taxonomy or geography due to widespread interspecific haplotype sharing. Based on both AFLP and cpDNA results, the two trans-Tasman species are shown to have originated in New Zealand and dispersed to Australia independently.