Search

Background and Aims Plant–fungus symbioses may experience temporal turnover during the ontogenetic or phenological development of the host, which can influence the ecological requirements of the host plant. In this study, we investigate temporal turnover of Ceratobasidiaceae orchid mycorrhizal fungal (OMF) communities in Prasophyllum (Orchidaceae), asking whether OMF communities are subject to temporal change attributable to orchid phenology or ontogeny. Methods Roots of adult Prasophyllum frenchii , Prasophyllum lindleyanum and Prasophyllum sp. aff. validum from Australia were sampled between autumn and spring. Seed was sown in situ as 'baits' to explore the mycorrhizal associations of germinating protocorms, which were compared with OMF in roots of co-occurring adult plants. Culture-dependent and -independent sequencing methods were used to amplify the internal transcribed spacer and mitochondrial large subunit loci, with sequences assigned to operational taxonomic units (OTUs) in phylogenetic analyses. Germination trials were used to determine whether fungal OTUs were mycorrhizal. Key Results A persistent core of OMF was associated with Prasophyllum , with Ceratobasidiaceae OMF dominant in all three species. Phenological turnover occurred in P. lindleyanum and P. sp. aff. validum , but not in P. frenchii , which displayed specificity to a single OTU. Ontogenetic turnover occurred in all species. However, phenological and ontogenetic turnover was typically driven by the presence or absence of infrequently detected OTUs in populations that otherwise displayed specificity to one or two dominant OTUs. Ex situ germination trials showed that 13 of 14 tested OTUs supported seed germination in their host orchid, including eight OTUs that were not found in protocorms in situ. Conclusions An understanding of OMF turnover can have practical importance for the conservation of threatened orchids and their mycorrhizal partners. However, frameworks for classifying OMF turnover should focus on OTUs important to the life cycle of the host plant, which we suggest are likely to be those that are frequently detected or functionally significant. [ABSTRACT FROM AUTHOR]

Background and Aims The entangled relationship of chromosome number and genome size with species distribution has been the subject of study for almost a century, but remains an open question due to previous ecological and phylogenetic knowledge constraints. To better address this subject, we used the clade Maxillariinae, a widely distributed and karyotypically known orchid group, as a model system to infer such relationships in a robust methodological framework. Methods Based on the literature and new data, we gathered the chromosome number and genome size for 93 and 64 species, respectively. We built a phylogenetic hypothesis and assessed the best macroevolutionary model for both genomic traits. Additionally, we collected together ecological data (preferences for bioclimatic variables, elevation and habit) used as explanatory variables in multivariate phylogenetic models explaining genomic traits. Finally, the impact of polyploidy was estimated by running the analyses with and without polyploids in the sample. Key Results The association between genomic and ecological data varied depending on whether polyploids were considered or not. Without polyploids, chromosome number failed to present consistent associations with ecological variables. With polyploids, there was a tendency to waive epiphytism and colonize new habitats outside humid forests. The genome size showed association with ecological variables: without polyploids, genome increase was associated with flexible habits, with higher elevation and with drier summers; with polyploids, genome size increase was associated with colonizing drier environments. Conclusions The chromosome number and genome size variations, essential but neglected traits in the ecological niche, are shaped in the Maxillariinae by both neutral and adaptive evolution. Both genomic traits are partially correlated to bioclimatic variables and elevation, even when controlling for phylogenetic constraints. While polyploidy was associated with shifts in the environmental niche, the genome size emerges as a central trait in orchid evolution by the association between small genome size and epiphytism, a key innovation to Neotropical orchid diversification. [ABSTRACT FROM AUTHOR]

Background and Aims Many terrestrial orchids have an obligate dependence on their mycorrhizal associations for nutrient acquisition, particularly during germination and early seedling growth. Though important in plant growth and development, phosphorus (P) nutrition studies in mixotrophic orchids have been limited to only a few orchid species and their fungal symbionts. For the first time, we demonstrate the role of a range of fungi in the acquisition and transport of inorganic P to four phylogenetically distinct green-leaved terrestrial orchid species (Diuris magnifica , Disa bracteata , Pterostylis sanguinea and Microtis media subsp. media) that naturally grow in P-impoverished soils. Methods Mycorrhizal P uptake and transfer to orchids was determined and visualized using agar microcosms with a diffusion barrier between P source (33P orthophosphate) and orchid seedlings, allowing extramatrical hyphae to reach the source. Key Results Extramatrical hyphae of the studied orchid species were effective in capturing and transporting inorganic P into the plant. Following 7 d of exposure, between 0.5 % (D. bracteata) and 47 % (D. magnifica) of the P supplied was transported to the plants (at rates between 0.001 and 0.097 fmol h−1). This experimental approach was capable of distinguishing species based on their P-foraging efficiency, and highlighted the role that fungi play in P nutrition during early seedling development. Conclusions Our study shows that orchids occurring naturally on P-impoverished soils can obtain significant amounts of inorganic P from their mycorrhizal partners, and significantly more uptake of P supplied than previously shown in other green-leaved orchids. These results provide support for differences in mycorrhiza-mediated P acquisition between orchid species and fungal symbionts in green-leaved orchids at the seedling stage. The plant–fungus combinations of this study also provide evidence for plant-mediated niche differentiation occurring, with ecological implications in P-limited systems. [ABSTRACT FROM AUTHOR]

Background and Aims Mycorrhizal fungi are a critical component of the ecological niche of most plants and can potentially constrain their geographical range. Unlike other types of mycorrhizal fungi, the distributions of orchid mycorrhizal fungi (OMF) at large spatial scales are not well understood. Here, we investigate the distribution and diversity of Ceratobasidium OMF in orchids and soils across the Australian continent. Methods We sampled 217 Ceratobasidium isolates from 111 orchid species across southern Australia and combined these with 311 Ceratobasidium sequences from GenBank. To estimate the taxonomic diversity of Ceratobasidium associating with orchids, phylogenetic analysis of the ITS sequence locus was undertaken. Sequence data from the continent-wide Australian Microbiome Initiative were used to determine the geographical range of operational taxonomic units (OTUs) detected in orchids, with the distribution and climatic correlates of the two most frequently detected OTUs modelled using MaxEnt. Key Results We identified 23 Ceratobasidium OTUs associating with Australian orchids, primarily from the orchid genera Pterostylis , Prasophyllum , Rhizanthella and Sarcochilus. OTUs isolated from orchids were closely related to, but distinct from, known pathogenic fungi. Data from soils and orchids revealed that ten of these OTUs occur on both east and west sides of the continent, while 13 OTUs were recorded at three locations or fewer. MaxEnt models suggested that the distributions of two widespread OTUs are correlated with temperature and soil moisture of the wettest quarter and far exceeded the distributions of their host orchid species. Conclusions Ceratobasidium OMF with cross-continental distributions are common in Australian soils and frequently have geographical ranges that exceed that of their host orchid species, suggesting these fungi are not limiting the distributions of their host orchids at large spatial scales. Most OTUs were distributed within southern Australia, although several OTUs had distributions extending into central and northern parts of the continent, illustrating their tolerance of an extraordinarily wide range of environmental conditions. [ABSTRACT FROM AUTHOR]

Background Given the exceptional diversity of orchids (26 000+ species), improving strategies for the conservation of orchids will benefit a vast number of taxa. Furthermore, with rapidly increasing numbers of endangered orchids and low success rates in orchid conservation translocation programmes worldwide, it is evident that our progress in understanding the biology of orchids is not yet translating into widespread effective conservation. Scope We highlight unusual aspects of the reproductive biology of orchids that can have important consequences for conservation programmes, such as specialization of pollination systems, low fruit set but high seed production, and the potential for long-distance seed dispersal. Further, we discuss the importance of their reliance on mycorrhizal fungi for germination, including quantifying the incidence of specialized versus generalized mycorrhizal associations in orchids. In light of leading conservation theory and the biology of orchids, we provide recommendations for improving population management and translocation programmes. Conclusions Major gains in orchid conservation can be achieved by incorporating knowledge of ecological interactions, for both generalist and specialist species. For example, habitat management can be tailored to maintain pollinator populations and conservation translocation sites selected based on confirmed availability of pollinators. Similarly, use of efficacious mycorrhizal fungi in propagation will increase the value of ex situ collections and likely increase the success of conservation translocations. Given the low genetic differentiation between populations of many orchids, experimental genetic mixing is an option to increase fitness of small populations, although caution is needed where cytotypes or floral ecotypes are present. Combining demographic data and field experiments will provide knowledge to enhance management and translocation success. Finally, high per-fruit fecundity means that orchids offer powerful but overlooked opportunities to propagate plants for experiments aimed at improving conservation outcomes. Given the predictions of ongoing environmental change, experimental approaches also offer effective ways to build more resilient populations. [ABSTRACT FROM AUTHOR]

Background and Aims Pyroloids, forest sub-shrubs of the Ericaceae family, are an important model for their mixotrophic nutrition, which mixes carbon from photosynthesis and from their mycorrhizal fungi. They have medical uses but are difficult to cultivate ex situ ; in particular, their dust seeds contain undifferentiated, few-celled embryos, whose germination is normally fully supported by fungal partners. Their germination and early ontogenesis thus remain elusive. Methods An optimized in vitro cultivation system of five representatives from the subfamily Pyroloideae was developed to study the strength of seed dormancy and the effect of different media and conditions (including light, gibberellins and soluble saccharides) on germination. The obtained plants were analysed for morphological, anatomical and histochemical development. Key Results Thanks to this novel cultivation method, which breaks dormancy and achieved up to 100 % germination, leafy shoots were obtained in vitro for representatives of all pyroloid genera (Moneses, Orthilia, Pyrola and Chimaphila). In all cases, the first post-germination stage is an undifferentiated structure, from which a root meristem later emerges, well before formation of an adventive shoot. Conclusions This cultivation method can be used for further research or for ex situ conservation of pyroloid species. After strong seed dormancy is broken, the tiny globular embryo of pyroloids germinates into an intermediary zone, which is functionally convergent with the protocorm of other plants with dust seeds such as orchids. Like the orchid protocorm, this intermediary zone produces a single meristem: however, unlike orchids, which produce a shoot meristem, pyroloids first generate a root meristem. [ABSTRACT FROM AUTHOR]

* Background and Aims Recent tissue-level observations made indirectly via flow cytometry suggest that endoreplication (duplication of the nuclear genome within the nuclear envelope in the absence of subsequent cell division) is widespread within the plant kingdom. Here, we also directly observe ploidy variation among cells within individual petals, relating size of nucleus to cell micromorphology and (more speculatively) to function. * Methods We compared the labella (specialized pollinator-attracting petals) of two European orchid genera: Dactylorhiza has a known predisposition to organismal polyploidy, whereas Ophrys exhibits exceptionally complex epidermal patterning that aids pseudocopulatory pollination. Confocal microscopy using multiple staining techniques allowed us to observe directly both the sizes and the internal structures of individual nuclei across each labellum, while flow cytometry was used to test for progressively partial endoreplication. * Key Results In Dactylorhiza, endoreplication was comparatively infrequent, reached only low levels, and appeared randomly located across the labellum, whereas in Ophrys endoreplication was commonplace, being most frequent in large peripheral trichomes. Endoreplicated nuclei reflected both endomitosis and endocycling, the latter reaching the third round of genome doubling (16C) to generate polytene nuclei. All Ophrys individuals studied exhibited progressively partial endoreplication. * Conclusions Comparison of the two genera failed to demonstrate the hypothesized pattern of frequent polyploid speciation in genera showing extensive endoreplication. Endoreplication in Ophrys appears more strongly positively correlated with cell size/complexity than with cell location or secretory role. Epigenetic control of gene overexpression by localized induction of endoreplication within individual plant organs may represent a significant component of a plant's developmental programme, contributing substantially to organ plasticity. [ABSTRACT FROM AUTHOR]

* Background and Aims Wildfires are common in seasonally dry parts of the world with a Mediterranean climate. Prescribed burning is used to reduce fuel load and fire risk, but often without reliable information on its effects. This study investigated the effects of prescribed burns in different seasons on Pterostylis revoluta, an autumn-flowering Australian terrestrial orchid, and its orchid mycorrhizal fungi (OMFs) to find the least damaging season for a prescribed burn. * Methods Burns were conducted mid-season in spring and summer 2011 and autumn and winter 2012. Orchids were enumerated and measured during their flowering season in autumn 2011-2014 and mycorrhizal fungi were isolated before and after the burns in autumn 2011, 2012 and 2014. Micro-organisms isolated were characterized. DNA was extracted from the OMFs, and the internal transcribed spacer region was amplified by PCR. Amplicons were clustered by restriction fragment length polymorphism (RFLP), and representative amplicons were sequenced. OMF were tested for sensitivity to smoke water. * Key Results The number of plants increased up to 4-fold and 90 % of plants became vegetative during this study. Isolation of mycorrhizal fungi increased and isolation of bacteria decreased. Before the burns, the main OMF isolated was unexpectedly Tulasnella calospora (Boud.) Juel. By 2014, after the burns, the expected Ceratobasidium sp. D.P. Rogers was the only OMF isolated in most burnt quadrats, whereas T. calospora was confined to a minority of unburnt 'control' and the 'spring burn' quadrats, which were also the only ones with flowering plants. * Conclusions The decline in rainfall during 2010--2012 probably caused the switch from mainly flowering to mainly vegetative plants and the change in OMFs. Burning in spring to summer was less damaging to this orchid than burning in autumn to winter, which should be noted by authorities in fire management plans for fire-prone areas in which this orchid occurs. [ABSTRACT FROM AUTHOR]

Background and Aims The endangered leafless ghost orchid, Dendrophylax lindenii, one of the most renowned orchids in the world, is difficult to grow under artificial conditions. Published information on asymbiotic and symbiotic (co-culture with a mycobiont) seed germination, seedling anatomy and developmental morphology of this leafless orchid is completely lacking. This information is critical for the development of efficient procedures for ghost orchid production for successful reintroduction.Methods Ghost orchid seedling early development stages were morphologically and anatomically defined to compare germination, embryo and protocorm maturation and seedling development during asymbiotic and symbiotic culture with one of two mycorrhizal strains (Dlin-379 and Dlin-394) isolated from ghost orchid roots in situ.Key Results Seeds symbiotically germinated at higher rates when cultured with fungal strain Dlin-394 than with strain Dlin-379 or asymbiotically on P723 medium during a 10-week culture period. Fungal pelotons were observed in protocorm cells co-cultured with strain Dlin-394 but not Dlin-379. Some 2-year-old seedlings produced multinode inflorescences in vitro. Production of keikis from inflorescence nodes indicated the capacity for clonal production in the ghost orchid.Conclusions Ghost orchid embryo and seedling development were characterized into seven stages. Fungal strain Dlin-394 was confirmed as a possible ghost orchid germination mycobiont, which significantly promoted seed germination and seedling development. Internal transcribed spacer sequencing data confirmed that Dlin-394 belongs within the genus Ceratobasidium. These results offer the opportunity to examine the benefits of using a mycobiont to enhance in vitro germination and possibly ex vitro acclimatization and sustainability following outplanting. [ABSTRACT FROM AUTHOR]

• Background The origin of new species and of new forms is one of the fundamental characteristics of evolution. However, the mechanisms that govern the diversity and disparity of lineages remain poorly understood. Particularly unclear are the reasons why some taxa are vastly more species-rich than others and the manner in which species diversity and morphological disparity are interrelated. • Scope and Conclusions Evolutionary innovations and ecological opportunities are usually cited as among the major factors promoting the evolution of species diversity. In many cases it is likely that these factors are positively reinforcing, with evolutionary innovations creating ecological opportunities that in turn foster the origin of new innovations. However, we propose that a third factor, developmental robustness, is very often essential for this reinforcement to be effective. Evolutionary innovations need to be stably and robustly integrated into the developmental genetic programme of an organism to be a suitable substrate for selection to 'explore' ecological opportunities and morphological 'design' space (morphospace). In particular, we propose that developmental robustness of the bauplan is often a prerequisite for the exploration of morphospace and to enable the evolution of further novelties built upon this bauplan. Thus, while robustness may reduce the morphological disparity at one level, it may be the basis for increased morphological disparity and for evolutionary innovations at another level, thus fostering species diversity. [ABSTRACT FROM AUTHOR]

Background and Aims The green orchid Goodyera repens has been shown to transfer carbon to its mycorrhizal partner, and this flux may therefore be affected by light availability. This study aimed to test whether the C and N exchange between plant and fungus is dependent on light availability, and in addition addressed the question of whether flowering and/or fruiting individuals of G. repens compensate for changes in leaf chlorophyll concentration with changes in C and N flows from fungus to plant. Methods The natural abundances of stable isotopes of plant C and N were used to infer changes in fluxes between orchid and fungus across natural gradients of irradiance at five sites. Mycorrhizal fungi in the roots of G. repens were identified by molecular analyses. Chlorophyll concentrations in the leaves of the orchid and of reference plants were measured directly in the field. Key Results Leaf δ13C values of G. repens responded to changes in light availability in a similar manner to autotrophic reference plants, and different mycorrhizal fungal associations also did not affect the isotope abundance patterns of the orchid. Flowering/fruiting individuals had lower leaf total N and chlorophyll concentrations, which is most probably explained by N investments to form flowers, seeds and shoot. Conclusions The results indicate that mycorrhizal physiology is relatively fixed in G. repens, and changes in the amount and direction of C flow between plant and fungus were not observed to depend on light availability. The orchid may instead react to low-light sites through increased clonal growth. The orchid does not compensate for low leaf total N and chlorophyll concentrations by using a 13C- and 15N-enriched fungal source [ABSTRACT FROM AUTHOR]

Background and Aims Polyploidization, the doubling of chromosome sets, is common in angiosperms and has a range of evolutionary consequences. Newly formed polyploid lineages are reproductively isolated from their diploid progenitors due to triploid sterility, but also prone to extinction because compatible mating partners are rare. Models have suggested that assortative mating and increased reproductive fitness play a key role in the successful establishment and persistence of polyploids. However, little is known about these factors in natural mixed-ploidy populations. This study investigated floral traits that can affect pollinator attraction and efficiency, as well as reproductive success in diploid and tetraploid Gymnadenia conopsea (Orchidaceae) plants in two natural, mixed-ploidy populations.Methods Ploidy levels were determined using flow cytometry, and flowering phenology and herbivory were also assessed. Reproductive success was determined by counting fruits and viable seeds of marked plants. Pollinator-mediated floral isolation was measured using experimental arrays, with pollen flow tracked by means of staining pollinia with histological dye.Key Results Tetraploids had larger floral displays and different floral scent bouquets than diploids, but cytotypes differed only slightly in floral colour. Significant floral isolation was found between the two cytotypes. Flowering phenology of the two cytotypes greatly overlapped, and herbivory did not differ between cytotypes or was lower in tetraploids. In addition, tetraploids had higher reproductive success compared with diploids.Conclusions The results suggest that floral isolation and increased reproductive success of polyploids may help to explain their successful persistence in mixed-ploidy populations. These factors might even initiate transformation of populations from pure diploid to pure tetraploid. [ABSTRACT FROM AUTHOR]

Background and Aims Lizard orchids of the genus Himantoglossum include many of Eurasia's most spectacular orchids, producing substantial spikes of showy flowers. However, until recently the genus had received only limited, and entirely traditional, systematic study. The aim of the current work was to provide a more robust molecular phylogeny in order to better understand the evolutionary relationships among species of particular conservation concern. Methods All putative species of Himantoglossum s.l. were sampled across its geographical range. A large subsample of the 153 populations studied contributed to an initial survey of nuclear ribosomal internal transcribed spacer (nrITS) ribotypes. Smaller subsets were then sequenced for four plastid regions and the first intron of the low-copy-number nuclear gene LEAFY. Rooted using Steveniella as outgroup, phylogenetic trees were generated using parsimony and Bayesian methods from each of the three datasets, supplemented with a ribotype network. Key Results The resulting trees collectively determined the order of branching of the early divergent taxa as Himantoglossum comperianum > H. robertianum group > H. formosum, events that also involved significant morphological divergence. Relaxed molecular clock dating suggested that these divergences preceded the Pleistocene glaciations (the origin of the H. robertianum group may have coincided with the Messinian salinity crisis) and occurred in Asia Minor and/or the Caucasus. Among more controversial taxa of the H. hircinum-jankae clade, which are only subtly morphologically divergent, topological resolution was poorer and topological incongruence between datasets was consequently greater. Conclusions Plastid sequence divergence is broadly consistent with prior, morphologically circumscribed taxa and indicates a division between H. hircinum–adriaticum to the west of the Carpathians and H. jankae–caprinum (plus local endemics) to the east, a distinction also suggested by nrITS ribotypes. LEAFY phylogenies are less congruent with prior taxonomic arrangements and include one likely example of paralogy. Himantoglossum metlesicsianum fully merits its IUCN Endangered status. Potentially significant genetic variation was detected within Steveniella satyrioides, H. robertianum and H. hircinum. However, confident circumscription of the more derived species of Himantoglossum s.s., including local endemics of hybrid origin, must await future morphometric and population-genetic analyses. [ABSTRACT FROM PUBLISHER]

Background and Aims Floral secretions are common in Bulbophyllum Thouars, and the labella of a number of Asian species are said to produce secretions rich in lipids that act as food rewards for insect pollinators. Although some of these reports are based on simple histochemical tests, a much greater number are anecdotal and, hitherto, neither the ultrastructure of the labellum nor the secretory process has been investigated in detail. Furthermore, sophisticated histochemical approaches have generally not been applied. Here, both the labellar structure and the secretory process are investigated for four species of Asian Bulbophyllum sect. Racemosae Benth. & Hook. f., namely Bulbophyllum careyanum (Hook.) Spreng., B. morphologorum Kraenzl., B. orientale Seidenf. and B. wangkaense Seidenf., and compared with those of unequivocal lipid-secreting orchids. Methods Labellar, secretory tissue was investigated using light microscopy, scanning electron microscopy, transmission electron microscopy and histochemistry. Key Results The adaxial median longitudinal groove of the labellum contained secretory tissue comprising palisade-like epidermal cells, similar to those of certain lipid-secreting Oncidiinae Benth. However, these cells and their secretions gave positive results mainly for protein and mucilage, and their organelle complement was consistent with that of cells involved in protein and mucilage synthesis. Sub-cuticular accumulation of secretion resulted in cuticular distension and blistering. The sub-epidermal layer of isodiametric parenchyma contained starch and, like the epidermal cells, ultrastructure consistent with mucilage synthesis. Lipids were mainly confined to the cuticle, and hardly any intracellular lipid droplets were observed. Conclusions It is proposed that mucilage is produced by dictyosomes present in the palisade-like epidermal cells. Mucilage precursors may also be produced by these same organelles in sub-epidermal cells and are thought to pass along the symplast via plasmodesmata into the adjoining palisade-like secretory cells, which contain abundant arrays of rough endoplasmic reticulum. Here, they become chemically modified and form a protein-rich, mucilaginous secretion that, following vesicle-mediated transport across the cytoplasm, traverses the cell wall and accumulates in blisters formed from the distended cuticle. Rupture of these blisters releases the secretion onto the labellar surface. However, in certain species, there is some evidence that the secretion may traverse the cuticle via cuticular pores, and micro-channels may permit the passage of fragrance. Hydrolysis of sub-epidermal starch probably generates the carbohydrate and, together with mitochondria, much of the energy required for the secretory process. This anatomical organization resembles that found in certain lipid-secreting, Neotropical species of Bulbophyllum and Oncidiinae, but since the chemical composition of their secretions is different, and these taxa occur on a separate continent and have different insect pollinators, parallelism of floral anatomy is likely. [ABSTRACT FROM PUBLISHER]

Background and Aims Pterostylis is an Australasian terrestrial orchid genus of more than 400 species, most of which use a motile, touch-sensitive labellum to trap dipteran pollinators. Despite studies dating back to 1872, the mechanism of pollinator attraction has remained elusive. This study tested whether the fungus gnat-pollinated Pterostylis sanguinea secures pollination by sexual deception. Methods The literature was used to establish criteria for confirming sexual deception as a pollination strategy. Observations and video recordings allowed quantification of each step of the pollination process. Each floral visitor was sexed and DNA barcoding was used to evaluate the degree of pollinator specificity. Following observations that attraction to the flowers is by chemical cues, experimental dissection of flowers was used to determine the source of the sexual attractant and the effect of labellum orientation on sexual attraction. Fruit set was quantified for 19 populations to test for a relationship with plant density and population size. Key Results A single species of male gnat (Mycetophilidae) visited and pollinated the rewardless flowers. The gnats often showed probing copulatory behaviour on the labellum, leading to its triggering and the temporary entrapment of the gnat in the flower. Pollen deposition and removal occurred as the gnat escaped from the flower via the reproductive structures. The labellum was the sole source of the chemical attractant. Gnats always alighted on the labellum facing upwards, but when it was rotated 180 ° they attempted copulation less frequently. Pollination rate showed no relationship with orchid population size or plant density. Conclusions This study confirms for the first time that highly specific pollination by fungus gnats is achieved by sexual deception in Pterostylis. It is predicted that sexual deception will be widespread in the genus, although the diversity of floral forms suggests that other mechanisms may also operate. [ABSTRACT FROM PUBLISHER]

Background and Aims The events leading to speciation are best investigated in systems where speciation is ongoing or incomplete, such as incipient species. By examining reproductive barriers among incipient sister taxa and their congeners we can gain valuable insights into the relative timing and importance of the various barriers involved in the speciation process. The aim of this study was to identify the reproductive barriers among sexually deceptive orchid taxa in the genus Chiloglottis. Methods The study targeted four closely related taxa with varying degrees of geographic overlap. Chemical, morphological and genetic evidence was combined to explore the basis of reproductive isolation. Of primary interest was the degree of genetic differentiation among taxa at both nuclear and chloroplast DNA markers. To objectively test whether or not species boundaries are defined by the chemistry that controls pollinator specificity, genetic analysis was restricted to samples of known odour chemistry. Key Results Floral odour chemical analysis was performed for 600+ flowers. The three sympatric taxa were defined by their specific chiloglottones, the semiochemicals responsible for pollinator attraction, and were found to be fully cross-compatible. Multivariate morphometric analysis could not reliably distinguish among the four taxa. Although varying from very low to moderate, significant levels of genetic differentiation were detected among all pairwise combinations of taxa at both nuclear and chloroplast loci. However, the levels of genetic differentiation were lower than expected for mature species. Critically, a lack of chloroplast DNA haplotype sharing among the morphologically indistinguishable and most closely related taxon pair confirmed that chemistry alone can define taxon boundaries. Conclusions The results confirmed that pollinator isolation, mediated by specific pollinator attraction, underpins strong reproductive isolation in these taxa. A combination of large effective population sizes, initial neutral mutations in the genes controlling floral scent, and a pool of available pollinators likely drives diversity in this system. [ABSTRACT FROM PUBLISHER]

Background and Aims The Orchidaceae have a history of recurring convergent evolution in floral function as nectar production has evolved repeatedly from an ancestral nectarless state. However, orchids exhibit considerable diversity in nectary type, position and morphology, indicating that this convergence arose from alternative adaptive solutions. Using the genus Disa, this study asks whether repeated evolution of floral nectaries involved recapitulation of the same nectary type or diversifying innovation. Epidermis morphology of closely related nectar-producing and nectarless species is also compared in order to identify histological changes that accompanied the gain or loss of nectar production. Methods The micromorphology of nectaries and positionally equivalent tissues in nectarless species was examined with light and scanning electron microscopy. This information was subjected to phylogenetic analyses to reconstruct nectary evolution and compare characteristics of nectar-producing and nectarless species. Key Results Two nectary types evolved in Disa. Nectar exudation by modified stomata in floral spurs evolved twice, whereas exudation by a secretory epidermis evolved six times in different perianth segments. The spur epidermis of nectarless species exhibited considerable micromorphological variation, including strongly textured surfaces and non-secreting stomata in some species. Epidermis morphology of nectar-producing species did not differ consistently from that of rewardless species at the magnifications used in this study, suggesting that transitions from rewardlessness to nectar production are not necessarily accompanied by visible morphological changes but only require sub-cellular modification. Conclusions Independent nectary evolution in Disa involved both repeated recapitulation of secretory epidermis, which is present in the sister genus Brownleea, and innovation of stomatal nectaries. These contrasting nectary types and positional diversity within types imply weak genetic, developmental or physiological constraints in ancestral, nectarless Disa. Such functional convergence generated by morphologically diverse solutions probably also underlies the extensive diversity of nectary types and positions in the Orchidaceae. [ABSTRACT FROM PUBLISHER]

Background and Aims: The Platanthera clade dominates the North American orchid flora and is well represented in eastern Asia. It has also generated some classic studies of speciation in Platanthera sections Platanthera and Limnorchis. However, it has proved rich in taxonomic controversy and near-monotypic genera. The clade is reviewed via a new molecular phylogenetic analysis and those results are combined with brief reconsideration of morphology in the group, aiming to rationalize the species into a smaller number of larger monophyletic genera and sections. [ABSTRACT FROM PUBLISHER]