Weak link or strong foundation? Vulnerability of fine root networks and stems to xylem embolism.

Resolving the position of roots in the whole-plant hierarchy of drought-induced xylem embolism resistance is fundamental for predicting when species become isolated from soil water resources. Published research generally suggests that roots are the most vulnerable organ of the plant vascular system, although estimates vary significantly. However, our knowledge of root embolism excludes the fine roots (< 2 mm diameter) that form the bulk of total absorptive surface area of the root network for water and nutrient uptake. We measured fine root and stem xylem vulnerability in 10 vascular plant species from the major land plant clades (five angiosperms, three conifers, a fern and lycophyte), using standardised in situ methods (Optical Methods and MicroCT). Mean fine root embolism resistance across the network matched or exceeded stems in all study species. In six of these species (one fern, one lycophyte, three conifers and one angiosperm), fine roots were significantly more embolism resistant than stems. No clear relationship was found between root xylem conduit diameter and vulnerability. These results provide insight into the resistance of the plant hydraulic pathway at the site of water and nutrient uptake, and challenge the long-standing assumption that fine roots are more vulnerable than stems.

Stomatal morphology and physiology explain varied sensitivity to abscisic acid across vascular plant lineages.

Abscisic acid (ABA) can induce rapid stomatal closure in seed plants, but the action of this hormone on the stomata of fern and lycophyte species remains equivocal. Here, ABA-induced stomatal closure, signaling components, guard cell K+ and Ca2+ fluxes, vacuolar and actin cytoskeleton dynamics, and the permeability coefficient of guard cell protoplasts (Pf) were analyzed in species spanning the diversity of vascular land plants including 11 seed plants, 6 ferns, and 1 lycophyte. We found that all 11 seed plants exhibited ABA-induced stomatal closure, but the fern and lycophyte species did not. ABA-induced hydrogen peroxide elevation was observed in all species, but the signaling pathway downstream of nitric oxide production, including ion channel activation, was only observed in seed plants. In the angiosperm faba bean (Vicia faba), ABA application caused large vacuolar compartments to disaggregate, actin filaments to disintegrate into short fragments and Pf to increase. None of these changes was observed in the guard cells of the fern Matteuccia struthiopteris and lycophyte Selaginella moellendorffii treated with ABA, but a hypertonic osmotic solution did induce stomatal closure in fern and the lycophyte. Our results suggest that there is a major difference in the regulation of stomata between the fern and lycophyte plants and the seed plants. Importantly, these findings have uncovered the physiological and biophysical mechanisms that may have been responsible for the evolution of a stomatal response to ABA in the earliest seed plants.

Functional diversity and convergence in the evolution of plant reproductive structures.

Background and Aims: Structures that simultaneously perform many functional roles are likely to show a variety of morphological solutions to these demands, and thus probably exhibit high morphological disparity. In contrast, specialization for a few simple functions should result in a more limited suite of morphologies. We explore this idea using lycopsid reproductive structures, which, throughout their history, have performed a limited set of functional roles compared with the reproductive structures of other plant groups such as seed plants. Methods: We scored living and fossil lycopsid taxa for 18 discrete character measurements and several continuous traits, including sporangium size, supporting axis diameter, and strobilus length and width. We used the discrete characters to construct a multivariate morphospace for lycopsid reproductive morphology through time, and the continuous characters to test whether fossil and extant lycopsids show similar patterns of tissue allocation within reproductive structures. Results: Lycopsids occupy similar areas of reproductive morphospace and show similar patterns of tissue allocation over most of their history, alternating between diffuse fertile zones with leaf-like sporophylls and compact strobili with specialized sporophylls that allow sporangia to be closely packed while also protected during their development. Growth habit also plays an important role in lycopsid reproductive evolution, broadly influencing the size and shape of reproductive structures. Conclusions: Lycopsid reproductive structures are primarily specialized for densely packaging sporangia, and are consistent with the idea that performing limited functional roles is associated with reduced morphological disparity. Morphologies similar to lycopsid strobili are also found in other groups with simple, wind-dispersed propagules, suggesting that the same processes occur across plant lineages.

Adaptive responses of mature giant chloroplasts in the deep-shade lycopod Selaginella erythropus to prolonged light and dark periods.

Deep-shade plants have adapted to low-light conditions by varying morphology and physiology of cells and chloroplasts, but it still remains unclear, if prolonged periods of high-light or darkness induce additional modifications in chloroplasts' anatomy and pigment patterns. We studied giant chloroplasts (bizonoplasts) of the deep-shade lycopod Selaginella erythropus in epidermal cells of mature fully developed microphylls and subjected them to prolonged darkness and high-light conditions. Chloroplast size and ultrastructure were investigated by light and electron microscopy. Physiological traits were studied by pigment analyses, photosynthetic performance of photosystem II, and formation of reactive oxygen species. Results show that (a) thylakoid patterns and shape of mature bizonoplasts vary in response to light and dark conditions. (b) Prolonged darkness induces transitory formation of prolamellar bodies, which so far have not been described in mature chloroplasts. (c) Photosynthetic activity is linked to structural responses of chloroplasts. (d) Photosystem II is less active in the upper zone of bizonoplasts and more efficient in the grana region. (e) Formation of reactive oxygen species reflects the stress level caused by high-light. We conclude that during prolonged darkness, chlorophyll persists and even increases; prolamellar bodies form de novo in mature chloroplasts; bizonoplasts have spatial heterogeneity of photosynthetic performance.

The enigma of sex allocation in Selaginella.

Background and Aims: The division of resource investment between male and female functions is poorly known for land plants other than angiosperms. The ancient lycophyte genus Selaginella is similar in some ways to angiosperms (in heterospory and in having sex allocation occur in the sporophyte generation, for example) but lacks the post-fertilization maternal investments that angiosperms make via fruit and seed tissues. One would therefore expect Selaginella to have sex allocation values less female-biased than in flowering plants and closer to the theoretical prediction of equal investment in male and female functions. Nothing is currently known of sex allocation in the genus, so even the simplest predictions have not been tested. Methods: Volumetric measurements of microsporangial and megasporangial investment were made in 14 species of Selaginella from four continents. In five of these species the length of the main above-ground axis of each plant was measured to determine whether sex allocation is related to plant size. Key Results: Of the 14 species, 13 showed male-biased allocations, often extreme, in population means and among the great majority of individual plants. There was some indication from the five species with axis length measurements that relative male allocation might be related to the release height of spores, but this evidence is preliminary. Conclusions: Sex allocation in Selaginella provides a phylogenetic touchstone showing how the innovations of fruit and seed investment in the angiosperm life cycle lead to typically female-biased allocations in that lineage. Moreover, the male bias we found in Selaginella requires an evolutionary explanation. The bias was often greater than what would occur from the mere absence of seed and fruit investments, and thus poses a challenge to sex allocation theory. It is possible that differences between microspores and megaspores in their dispersal ecology create selective effects that favour male-biased sexual allocation. This hypothesis remains tentative.

Vascularization of the Selaginella rhizophore: anatomical fingerprints of polar auxin transport with implications for the deep fossil record.

The Selaginella rhizophore is a unique and enigmatic organ whose homology with roots, shoots, or neither of the two remains unresolved. Nevertheless, rhizophore-like organs have been documented in several fossil lycophytes. Here we test the homology of these organs through comparisons with the architecture of rhizophore vascularization in Selaginella. We document rhizophore vascularization in nine Selaginella species using cleared whole-mounts and histological sectioning combined with three-dimensional reconstruction. Three patterns of rhizophore vascularization are present in Selaginella and each is comparable to those observed in rhizophore-like organs of fossil lycophytes. More compellingly, we found that all Selaginella species sampled exhibit tracheids that arc backward from the stem and side branch into the rhizophore base. This tracheid curvature is consistent with acropetal auxin transport previously documented in the rhizophore and is indicative of the redirection of basipetal auxin from the shoot into the rhizophore during development. The tracheid curvature observed in Selaginella rhizophores provides an anatomical fingerprint for the patterns of auxin flow that underpin rhizophore development. Similar tracheid geometry may be present and should be searched for in fossils to address rhizophore homology and the conservation of auxin-related developmental mechanisms from early stages of lycophyte evolution.

A subgeneric classification of Selaginella (Selaginellaceae).

PREMISE OF THE STUDY: The lycophyte family Selaginellaceae includes approximately 750 herbaceous species worldwide, with the main species richness in the tropics and subtropics. We recently presented a phylogenetic analysis of Selaginellaceae based on DNA sequence data and, with the phylogeny as a framework, the study discussed the character evolution of the group focusing on gross morphology. Here we translate these findings into a new classification. METHODS: To present a robust and useful classification, we identified well-supported monophyletic groups from our previous phylogenetic analysis of 223 species, which together represent the diversity of the family with respect to morphology, taxonomy, and geographical distribution. Care was taken to choose groups with supporting morphology. KEY RESULTS: In this classification, we recognize a single genus Selaginella and seven subgenera: Selaginella, Rupestrae, Lepidophyllae, Gymnogynum, Exaltatae, Ericetorum, and Stachygynandrum. The subgenera are all well supported based on analysis of DNA sequence data and morphology. A key to the subgenera is presented. CONCLUSIONS: Our new classification is based on a well-founded hypothesis of the evolutionary relationships of Selaginella, and each subgenus can be identified by a suite of morphological features, most of them possible to study in the field. Our intention is that the classification will be useful not only to experts in the field, but also to a broader audience.

Phylogeny of Selaginellaceae: There is value in morphology after all!

PREMISE OF THE STUDY: The cosmopolitan lycophyte family Selaginellaceae, dating back to the Late Devonian-Early Carboniferous, is notorious for its many species with a seemingly undifferentiated gross morphology. This morphological stasis has for a long time hampered our understanding of the evolutionary history of the single genus Selaginella. Here we present a large-scale phylogenetic analysis of Selaginella, and based on the resulting phylogeny, we discuss morphological evolution in the group. METHODS: We sampled about one-third of the approximately 750 recognized Selaginella species. Evolutionary relationships were inferred from both chloroplast (rbcL) and single-copy nuclear gene data (pgiC and SQD1) using a Bayesian inference approach. The morphology of the group was studied and important features mapped onto the phylogeny. KEY RESULTS: We present an overall well-supported phylogeny of Selaginella, and the phylogenetic positions of some previously problematic taxa (i.e., S. sinensis and allies) are now resolved with strong support. We show that even though the evolution of most morphological characters involves reversals and/or parallelisms, several characters are phylogenetically informative. Seven major clades are identified, which each can be uniquely diagnosed by a suite of morphological features. There is value in morphology after all! CONCLUSIONS: Our hypothesis of the evolutionary relationships of Selaginella is well founded based on DNA sequence data, as well as morphology, and is in line with previous findings. It will serve as a firm basis for further studies on Selaginella with respect to, e.g., the poorly known alpha taxonomy, as well as evolutionary questions such as historical biogeographic reconstructions.

Independent recruitment of an O-methyltransferase for syringyl lignin biosynthesis in Selaginella moellendorffii.

Syringyl lignin, an important component of the secondary cell wall, has traditionally been considered to be a hallmark of angiosperms because ferns and gymnosperms in general lack lignin of this type. Interestingly, syringyl lignin was also detected in Selaginella, a genus that represents an extant lineage of the most basal of the vascular plants, the lycophytes. In angiosperms, syringyl lignin biosynthesis requires the activity of ferulate 5-hydroxylase (F5H), a cytochrome P450-dependent monooxygenase, and caffeic acid/5-hydroxyferulic acid O-methyltransferase (COMT). Together, these two enzymes divert metabolic flux from the biosynthesis of guaiacyl lignin, a lignin type common to all vascular plants, toward syringyl lignin. Selaginella has independently evolved an alternative lignin biosynthetic pathway in which syringyl subunits are directly derived from the precursors of p-hydroxyphenyl lignin, through the action of a dual specificity phenylpropanoid meta-hydroxylase, Sm F5H. Here, we report the characterization of an O-methyltransferase from Selaginella moellendorffii, COMT, the coding sequence of which is clustered together with F5H at the adjacent genomic locus. COMT is a bifunctional phenylpropanoid O-methyltransferase that can methylate phenylpropanoid meta-hydroxyls at both the 3- and 5-position and function in concert with F5H in syringyl lignin biosynthesis in S. moellendorffii. Phylogenetic analysis reveals that Sm COMT, like F5H, evolved independently from its angiosperm counterparts.

Taxonomic significance of megaspores in some species of Selaginella from Arunachal Pradesh in India.

The present study was carried out to record the color, size, and ornamentation of megaspores in 18 species of the family Selaginellaceae from Arunachal Pradesh using scanning electron microscope (SEM) and light microscope (LM). Electron microscopic study of the spore features revealed the type of ornamentation and microsculptural detail. SEM study on megaspore of Selaginella pentagona (S. pentagona), Selaginella tenuifolia, Selaginella semicordata, and Selaginella chrysorrhizos is presented for the first time. Variation in the megaspore ornamentation is noted at the interspecific level and intraspecific level in some cases. Examination of the megaspores under study found all the megaspores as trilete, with a size ranging from 116 to 560 µm in diameter. Taxonomic key is prepared to differentiate the species. Short descriptions of megaspores are provided and supported by photo plates. New features are recorded for seven species. The study contributes to the separation of species within the genus Selaginella based on the spore feature and brings forward the use of spore as a diagnostic tool in the taxonomy of the genus. These works contribute to the systematic of the family Selaginellaceae and provide useful information in the field of palynology. RESEARCH HIGHLIGHTS: Study on megaspore features of Selaginella species collected from Arunachal Pradesh, using SEM and LM. Taxonomic key is provided for each species based on megaspores features. New megaspore features are recorded for the seven species. The study brings forward the use of spore as a diagnostic tool in the taxonomy of the genus. These works provide useful information in the field of systematic and palynology.

Molecular evolution and expression divergence of the Populus polygalacturonase supergene family shed light on the evolution of increasingly complex organs in plants.

Plant polygalacturonases (PGs) are involved in cell separation processes during many stages of plant development. Investigation into the diversification of this large gene family in land plants could shed light on the evolution of structural development. We conducted whole-genome annotation, molecular evolution and gene expression analyses of PG genes in five species of land plant: Populus, Arabidopsis, rice, Selaginella and Physcomitrella. We identified 75, 44, 16 and 11 PG genes from Populus, rice, Selaginella and Physcomitrella genomes, respectively, which were divided into three classes. We inferred rapid expansion of class I PG genes in Populus, Arabidopsis and rice, while copy numbers of classes II and III PG genes were relatively conserved in all five species. Populus, Arabidopsis and rice class I PG genes were under more relaxed selection constraints than class II PG genes, while this selective pressure divergence was not observed in Selaginella and Physcomitrella PG families. In addition, class I PG genes underwent marked expression divergence in Populus, rice and Selaginella. Our results suggest that PG gene expansion occurred after the divergence of the lycophytes and euphyllophytes, and this expansion was likely paralleled by the evolution of increasingly complex organs in land plants.

[Study on quality standard of Selaginella moellendorffii].

OBJECTIVE: To study the quality standard of Selaginella moellendorffii. METHODS: The macroscopic and microscopic characteristics were observed, amentoflavone was used as reference substance in the TLC identification and HPLC method was used to determine the content of amentoflavone. RESULTS: The TLC method used GF254 taking toluene-ethyl formate-formic acid (5: 4: 0. 5) as the developer and ethanol solution of ferric chloride as the chromogenic reagent with the results that it appeared four clear spots. The HPLC method took amentoflavone as the reference substance, and acetonitrile-water (containing 3% tetrahydrofuran and 3% trifluoroacetic acid) as the mobile phase; Detection wavelength was 330 nm. The contents of samples from 3 different batches were determined, and the lowest content of amentoflavone was 0.4%. CONCLUSION: This method is reliable and accurate, which can be used for the quality control of Selaginella moellendorfii.

Distinct developmental mechanisms reflect the independent origins of leaves in vascular plants.

Vascular plants diverged more than 400 million years ago into two lineages, the lycophytes and the euphyllophytes . Leaf-like organs evolved independently in these two groups . Microphylls in lycophytes are hypothesized to have originated as lateral outgrowths of tissue that later became vascularized (the enation theory) or through the sterilization of sporangia (the sterilization hypothesis) . Megaphylls in euphyllophytes are thought to represent modified lateral branches . The fossil record also indicates that the seed plant megaphyll evolved uniquely in the ancestor of seed plants, independent of megaphylls in ferns, because seed plants evolved from leafless progymnosperm ancestors . Surprisingly, a recent study of KNOX and ARP gene expression in a lycophyte was reported to indicate recruitment of a similar mechanism for determinacy in both types of leaves . We examined the expression of Class III HD-Zip genes in the lycophyte Selaginella kraussiana and in two gymnosperms, Ginkgo and Pseudotsuga. Our data indicate that mechanisms promoting leaf initiation, vascularization, and polarity are quite different in lycophytes and seed plants, consistent with the hypotheses that megaphylls originated as lateral branches whereas microphylls originated as tissue outgrowths.

[RAPD analysis of 8 medicinal species of Selaginella].

OBJECTIVE: To identify 8 medicinal species of Selaginella with method of molecular marker and to explore their genetic relationship. METHODS: 8 primers were selected from 60 random primers to amplify the total DNA extracts of 17 samples with RAPD method and then the amplified were analyzed. RESULTS: A total of amplified 58 bands and RAPD map were obtained. Based on them, a Jaccard's genetic similarity matrix and a dendrogram for the 8 plants were established using SPSS 11.5 software. CONCLUSION: The method revealed distinct differences of different species and the variation of the same species of Selaginella from different habitats. It can provide with genetic proof for authentication of some species and units below species.

[Investigations and studies on medicinal plants from Selaginellaceae in Hubei Province].

OBJECTIVE: To investigate the species, the distribution and the utilization of the medicinal plants from Selaginellaceae in Hubei Province. METHOD: Through field investigations and comparing the collected specimens and literatures, the classification and identification of the species in Hubei Province were studied. RESULT: The results indicated that 15 species of plants from Selaginellaceae in Hubei, including 14 medicinal and 1 newly recorded species existed. The distribution and use in folk medicine were investigated. And the morphological description of several species was appended. CONCLUSION: The results provided a basis for the exploitation and utilization of the medicinal plant resources of Selagingellaceae.

Analysing photonic structures in plants.

The outer layers of a range of plant tissues, including flower petals, leaves and fruits, exhibit an intriguing variation of microscopic structures. Some of these structures include ordered periodic multilayers and diffraction gratings that give rise to interesting optical appearances. The colour arising from such structures is generally brighter than pigment-based colour. Here, we describe the main types of photonic structures found in plants and discuss the experimental approaches that can be used to analyse them. These experimental approaches allow identification of the physical mechanisms producing structural colours with a high degree of confidence.

The glycosyltransferase repertoire of the spikemoss Selaginella moellendorffii and a comparative study of its cell wall.

Spike mosses are among the most basal vascular plants, and one species, Selaginella moellendorffii, was recently selected for full genome sequencing by the Joint Genome Institute (JGI). Glycosyltransferases (GTs) are involved in many aspects of a plant life, including cell wall biosynthesis, protein glycosylation, primary and secondary metabolism. Here, we present a comparative study of the S. moellendorffii genome across 92 GT families and an additional family (DUF266) likely to include GTs. The study encompasses the moss Physcomitrella patens, a non-vascular land plant, while rice and Arabidopsis represent commelinid and non-commelinid seed plants. Analysis of the subset of GT-families particularly relevant to cell wall polysaccharide biosynthesis was complemented by a detailed analysis of S. moellendorffii cell walls. The S. moellendorffii cell wall contains many of the same components as seed plant cell walls, but appears to differ somewhat in its detailed architecture. The S. moellendorffii genome encodes fewer GTs (287 GTs including DUF266s) than the reference genomes. In a few families, notably GT51 and GT78, S. moellendorffii GTs have no higher plant orthologs, but in most families S. moellendorffii GTs have clear orthologies with Arabidopsis and rice. A gene naming convention of GTs is proposed which takes orthologies and GT-family membership into account. The evolutionary significance of apparently modern and ancient traits in S. moellendorffii is discussed, as is its use as a reference organism for functional annotation of GTs.

Plant genomics: homoplasy heaven in a lycophyte genome.

The recent genomic sequencing of Selaginella, a member of the lycophyte lineage of vascular plants, opens up all kinds of new opportunities to examine the patterns of evolutionary innovation and the creation of the basic bauplan of plants.

Function of blue iridescence in tropical understorey plants.

The blue colouration seen in the leaves of Selaginella willdenowii is shown to be iridescent. Transmission electron microscopy studies confirm the presence of a layered lamellar structure of the upper cuticle of iridescent leaves. Modelling of these multi-layer structures suggests that they are responsible for the blue iridescence, confirming the link between the observed lamellae and the recorded optical properties. Comparison of blue and green leaves from the same plant indicates that the loss of the blue iridescence corresponds to a loss of the multi-layer structure. The results reported here do not support the idea that iridescence in plants acts to enhance light capture of photosynthetically important wavelengths. The reflectance of light in the range 600-700 nm is very similar for both iridescent and non-iridescent leaves. However, owing to the occurrence of blue colouration in a wide variety of shade dwelling plants it is probable that this iridescence has some adaptive benefit. Possible adaptive advantages of the blue iridescence in these plants are discussed.