A comparative study of cycad mucilages

The patterns of monosaccharide distribution of the mucilages of Cycadales are characteristic at the generic level. Arabinose, fucose, galactose, mannose, rhamnose and methylrhamnose were identified in the hydrolysed mucilage of Bowenia, Ceratozamia, Cycas, Dioon, Encephalartos, Lepidozamia, Macrozamia, Microcycas and Zamia. Stangeria contains no rhamnose and methylrhamnose, and Ceratozamia contains galactose only in traces. American genera may easily be distinguished from the others by means of their different monosaccharide composition. Lepidozamia appears to be well separated from Macrozamia.     

Phylogeny of the cycads based on multiple single-copy nuclear genes: congruence of concatenated parsimony,likelihood and species tree inference methods

Background and aims

Despite a recent new classification, a stable phylogeny for the cycads has been elusive, particularly regarding resolution of Bowenia, Stangeria and Dioon. In this study, five single-copy nuclear genes (SCNGs) are applied to the phylogeny of the order Cycadales. The specific aim is to evaluate several gene tree–species tree reconciliation approaches for developing an accurate phylogeny of the order, to contrast them with concatenated parsimony analysis and to resolve the erstwhile problematic phylogenetic position of these three genera.

Methods

DNA sequences of five SCNGs were obtained for 20 cycad species representing all ten genera of Cycadales. These were analysed with parsimony, maximum likelihood (ML) and three Bayesian methods of gene tree–species tree reconciliation, using Cycas as the outgroup. A calibrated date estimation was developed with Bayesian methods, and biogeographic analysis was also conducted.

Key Results

Concatenated parsimony, ML and three species tree inference methods resolve exactly the same tree topology with high support at most nodes. Dioon and Bowenia are the first and second branches of Cycadales after Cycas, respectively, followed by an encephalartoid clade (Macrozamia–Lepidozamia–Encephalartos), which is sister to a zamioid clade, of which Ceratozamia is the first branch, and in which Stangeria is sister to Microcycas and Zamia.

Conclusions

A single, well-supported phylogenetic hypothesis of the generic relationships of the Cycadales is presented. However, massive extinction events inferred from the fossil record that eliminated broader ancestral distributions within Zamiaceae compromise accurate optimization of ancestral biogeographical areas for that hypothesis. While major lineages of Cycadales are ancient, crown ages of all modern genera are no older than 12 million years, supporting a recent hypothesis of mostly Miocene radiations. This phylogeny can contribute to an accurate infrafamilial classification of Zamiaceae.     

KARYOTYPIC DATA ON NORTH AND CENTRAL AMERICAN ZAMIACEAE (CYCADALES) AND THEIR PHYLOGENETIC IMPLICATIONS

Chromosome numbers and karyotypes of species from four American Zamiaceae (Cycadales) are reported. Zamia shows interspecific and intraspecific chromosome variation, whereas Microcycas, Ceratozamia, and Dioon have constant karyotypes within each genus. In Zamia, all karyotypes have the same number of submetacentric and acrocentric chromosomes, but they differ in the number of metacentric and telocentric chromosomes. Centric fission of metacentric chromosomes is proposed to explain the karyotypic variation in this genus. Zamia shows karyological relationships with Microcycas and Ceratozamia, whereas Dioon appears very distinct from the other American cycad genera. Affinity among Zamia, Ceratozamia, and Microcycas karyotypes and distinctiveness of Dioon karyotypes are supported by comparative analysis of phenotypic characters in the four genera.     

Molecular systematic studies in cycads: Evidence fromtrnL Intron and ITS2 rDNA sequences

The results of a pilot DNA sequencing study of cycads conducted at the new molecular systematics laboratory at Fairchild Tropical Garden are presented and assessed with reference to previous phylogenetic analyses and classification schemes based on morphology and anatomy. Two DNA regions were sequenced and analyzed for variation, an intron in the trriL gene in the chloroplast genome (trriL intron) and the internal transcribed spacer region between the 5.8S and 26S ribosomal DNA subunits (ITS2). The trnL intron proved to be relatively conservative among cycad genera, while the ITS2 region contained higher levels of variation. Parsimony analysis of the sequences suggests a number of relationships, some of which were inferred by previous morphological studies, some of which are new. The sequences ofCycas are the most divergent among cycads, suggesting the longest isolation.Dioon is relatively isolated from the other genera and contains two major clades.Stangeria does not appear closely related toBowenia but does seem to have a weak affinity withZamia andMicrocycas. Lepidozamia is more closely related toEncephalartos than toMacrozamia. Sequence variation among the species ofCeratozamia is low.Microcycas andZamia are closely related.     

COMPARATIVE POLLEN MORPHOLOGY AND TAXONOMIC AFFINITIES IN CYCADALES

Scanning and transmission electron microscopy of pollen grains of 29 species, representing the ten extant genera of Cycadales, has provided valuable insight into their relationships. Pollen grains of these taxa are boat-shaped, monosulcate, and bilaterally symmetrical. They range from narrowly to widely elliptical or subcircular when viewed distally, and have an exine surface of psilate, foveolate, or fossulate. Pollen wall ultrastructure of Cycadales is typically tectate with alveolate∗∗∗spongy exine. The nexine is laminated in all genera. Nexine 1 (footlayer) is present in most species as a thin and often discontinuous layer. There is consistent variation in thickness of the sporoderm layers among the genera but relative uniformity within them. Pollen characteristics are well correlated with macro- and micromorphological features, chromosome numbers, geographical distribution, and postulated pollination mode. A close affinity between Encephalartos, Lepidozamia, and Macrozamia is recognized. Pollen characteristics of the genus Bowenia show some similarity with those of the latter group. Except for two species of Macrozamia which are narrowly elliptic, all of the genera have widely elliptic pollen and share a psilate exine surface and the thinnest sexine with nearly identical arrangement of alveoli. Pollen grains of the species in the genus Dioon exhibit a unique morphology but are more similar to Stangeria than they are to those of taxa in Zamiaceae. The circular outline of the grains and the foveolate exine surface are characters shared by these two genera, but several morphological features distinguish Dioon from Stangeria. Ceratozamia and Zamia share a widely elliptic shape, foveolate exine surface and nearly identical sexine, as well as morphological features and chromosome numbers. They differ from Microcycas in sexine thickness, gross morphology and chromosome numbers. The pollen grains of Cycas circinalis and C. revoluta differ in size and structure of the sexine from all other genera and from each other, substantiating their distinct subgeneric delimitations.     

Seed coat carotenoids of the cycad genera Dioon,Encephelartos, Macrozamia and Zamia: Evolutionary significance

Seed coat carotenoids of Dioon, Encephelartos, Macrozamia and Zamia with ranges in Africa, the West Indies, Australia and Mexico are simple mixtures which conform to the theoretical primitive angiosperm “magnolian pattern”. Lycopene is the principal pigment of Zamia while Encephelartos, Dioon and Macrozamia coats contain a mixture of the unsubstituted, mono-and di-hydroxy-β-carotenes. The evolutionary significance of these results is discussed.     

Midribs of cycad pinnae

Comparisons of midribs of three cycad genera,Chigua, Cycas, andStangeria, the only three genera characterized by midribs in the pinnae, show that those ofChigua andStangeria are very similar to each other and quite unlike those ofCycas. Midribs ofCycas include a single, median vein, and the pinnae of these species lack lateral veins. Pinnae midribs ofChigua andStangeria include several (2–5 and 2–8, respectively) longitudinal parallel veins, and dichotomizing lateral veins arising from the midrib. Pinnae of other cycads, includingZamia, with whichChigua appears to be most closely allied, exhibit evenly spaced, longitudinally parallel, dichotomizing veins, a character considered to be primitive. All veins in cycad leaves have a single mesarch protoxylem pole. The midrib condition inChigua andStangeria represents an advanced state in comparison to that in the leaf of the Marattiales, for example, where there is a single veined midrib but with numerous mesarch protoxylem poles. It would appear that the midrib has been derived independently at least three times within the cycads, once in each major group.     

Ribosomal DNA distribution in somatic chromosomes of Stangeria eriopus (Stangeriaceae, Cycadales) and molecular-cytotaxonomic relationships to some other cycad genera

Somatic chromosomes ofStangeria eriopus (Stangeriaceae, Cycadales) were investigated by fluorescentin situ hybridization (FISH) using an 18S ribosomal DNA (rDNA) probe.Stangeria eriopus showed a chromosome number of 2n=16 with a karyotype of 12 median-, 2 subterminal-, and 2 terminal-centromeric chromosomes. FISH study ofS. eriopus revealed 16 signals made up of rDNA sites located on the terminal regions of the long arms of the 7 median- and 2 subterminal-centromeric chromosomes, on terminal region of the short arm of the 1 median-centromeric chromosome, on the terminal regions of the long and the short arms of 1 median- and 2 terminal-centromeric chromosomes. This result suggests that, not only karyomorphologically but also molecular-cytologically, the genusStangeria may be more closely related to the genusCeratozamia than the genusBowenia or the genusMicrocycas previously hypothesized.     

Structure and distribution of floral trichomes in Lycaste and Sudamerlycaste (Orchidaceae: Maxillariinae s.l.)

Phylogenetic analysis indicates that Lycastinae should be incorporated into a more broadly defined Maxillariinae. This is supported by several anatomical features, including the presence of sunken, glandular trichomes in both Lycastinae and Maxillariinae s.s. Until recently, these were known only from vegetative organs, but have since been reported from flowers of Maxillaria dichroma. One character currently used to distinguish between Lycaste and Sudamerlycaste is the distribution of floral trichomes. In this article, we test the reliability of this character, describe the floral micromorphology of Lycaste and Sudamerlycaste and investigate whether their flowers bear sunken hairs. Their floral micromorphology is compared with that of other genera currently assigned to Maxillariinae s.l. Flowers of Lycaste and Sudamerlycaste bear conical or obpyriform papillae and unbranched and unequally branched multicellular trichomes. Contrary to previous reports that trichomes are confined to the column in Sudamerlycaste, they also occur in the tepal axils. Labellar trichomes, although often present in Lycaste, are lacking in Sudamerlycaste. In Lycaste sections Lycaste and Aromaticae, floral trichomes tend to be unbranched, whereas section Intermediae has both unbranched and branched hairs. Branched hairs are more common in Sudamerlycaste. Some hairs are tracheoidal, pitted and lignified. These mainly occur in section Lycaste and, to a degree, in section Intermediae, but are absent from section Aromaticae and most species of Sudamerlycaste. Branched column hairs, present in Sudamerlycaste, are absent from all sections of Lycaste, and tracheoidal column hairs occur only in Sudamerlycaste. Sunken floral hairs are absent from both genera. Trichome structure and distribution may prove useful in distinguishing between these taxa and in elucidating the intergeneric relationships of Maxillariinae s.l.© 2010 The Linnean Society of London, Botanical Journal of the Linnean Society, 2010, 164 , 409–421.     

Biflavonoids from the seed testa of cycadales

Biflavonoid patterns from ethanolic extracts of the brightly coloured testa of eight species of Cycadales have been determined. Species from the genus Macrozamia are characterized by the occurrence of cupressuflavone- and amentoflavone-based patterns, while the Cycas species contain only amentoflavone-based patterns. The other species from the genera Encephalartos, Lepidozamia, and Zamia contain a very minor or no detectable biflavonoid component. This is the first report of the occurrence of cupressuflavone in the Cycadales.     

Stomatal development in the cycad family Zamiaceae

Background and AimsThe gymnosperm order Cycadales is pivotal to our understanding of seed-plant phylogeny because of its phylogenetic placement close to the root node of extant spermatophytes and its combination of both derived and plesiomorphic character states. Although widely considered a ‘living fossil’ group, extant cycads display a high degree of morphological and anatomical variation. We investigate stomatal development in Zamiaceae to evaluate variation within the order and homologies between cycads and other seed plants.MethodsLeaflets of seven species across five genera representing all major clades of Zamiaceae were examined at various stages of development using light microscopy and confocal microscopy.Key ResultsAll genera examined have lateral subsidiary cells of perigenous origin that differ from other pavement cells in mature leaflets and could have a role in stomatal physiology. Early epidermal patterning in a ‘quartet’ arrangement occurs in Ceratozamia, Zamia and Stangeria. Distal encircling cells, which are sclerified at maturity, are present in all genera except Bowenia, which shows relatively rapid elongation and differentiation of the pavement cells during leaflet development.ConclusionsStomatal structure and development in Zamiaceae highlights some traits that are plesiomorphic in seed plants, including the presence of perigenous encircling subsidiary cells, and reveals a clear difference between the developmental trajectories of cycads and Bennettitales. Our study also shows an unexpected degree of variation among subclades in the family, potentially linked to differences in leaflet development and suggesting convergent evolution in cycads.     

SYSTEMATIC STUDIES ON THE MEXICAN ZAMIACEAE. I. CHROMOSOME NUMBERS AND KARYOTYPES

Chromosome numbers and karyotypes are reported for seven species in three genera, of Mexican cycads. Karyotypic analysis of Cycads is made difficult by differential contraction, of their large chromosomes but idiograms are presented. Satellite variation is reported for three species of Ceratozamia. No satellites are reported for the three species of Zamia.     

Ultrastructural ontogeny of leaf cavity trichomes inAzolla implies a functional role in metabolite exchange

Summary Anabaena azollae is associated with two types of multicellular epidermal trichomes inAzolla leaf cavities, the simple and branched hairs. The observation of transfer cell ultrastructure in some hair cells led to speculation that the cavity hairs might participate in metabolite exchange between the symbionts. The developmental ontogeny of cavity trichomes is described here, using transmission electron microscopy, with a goal of improving our understanding of possible functions of these structures in the symbiosis. The observations have established that all cells of simple and branched hairs develop the structural characteristics of transfer cells, but not simultaneously. Rather, there is an acropetal succession of transfer cell ultrastructure beginning in terminal cells, moving to body cells where present, and ending in stalk cells. The transfer cell stage is followed immediately by senescence in all hair cells. The timing of transfer cell differentiation, considered together with information from other studies, suggests that branched hairs may be involved in exchange of fixed nitrogen between the symbionts, while simple hairs may participate in exchange of fixed carbon fromAzolla toAnabaena. Contribution no. 869 from the Battelle-C. F. Kettering Research Laboratory.     

黑龙江悬钩子属植物叶表皮形态结构的研究

利用扫描电子显微镜对黑龙江悬钩子属植物的叶表皮形态结构进行比较研究。结果显示:(1)悬钩子属植物叶的上表皮细胞呈多边形,垂周壁平直,或无规则形,垂周壁浅波纹;下表皮细胞无规则形,垂周壁浅波纹或深波纹。(2)表皮毛类型有单细胞直立不分支、卷曲不分支,头状腺毛和盾状腺毛四种类型。(3)气孔器均分布于下表皮,且气孔器类型为无规则形;气孔外拱盖单层、内缘平滑或不规则波状。研究表明,黑龙江悬钩子属植物的叶表皮微形态学特征表现出一定差异性,对种间的划分和鉴定具有一定的分类学意义。     

A chemotaxonomic study of mucilages in American Zamiaceae

Abstract

Methylrhamnose, fucose, rhamnose, mannose, arabinose, galactose, and glucuronolactone are the monosaccharides identified in hydrolysed mucilages collected from 37 species belonging to the five American cycad genera (Ceratozamia, Chigua, Dioon, Microcycas and Zamia). The patterns of monosaccharide distribution in the mucilages resulted to be typical at the generic level, with the exception of Chigua and Zamia the patterns of which were similar. No differences appeared among species within single genera, with the exception of Zamia in which minor differences were observed at the intrageneric level. Differences and similarities among patterns at both intergeneric and intrageneric levels are discussed for their systematic implications.     

Foliar indumentum in central‐European Rubus species (Rosaceae) and its contribution to the systematics of the group

The foliar indumentum of 94 taxa of Rubus has been surveyed by scanning electron microscopy. The analysed species belonged to five subgenera. Rubus subgen. Rubus with its three sections and 21 series was the most numerously represented. In all studied species, indumentum was present, but the species differed more or less from each other with respect to the density of the indumentum on the abaxial leaf surface. Three main hair types were present: simple eglandular (unbranched) trichomes, branched eglandular trichomes, and very short secretory uni‐seriate trichomes; subtypes were distinguished according to trichome size. Apart from investigating separate traits, we described character patterns for the best represented and homogeneous series: Discolores, Subthyrsoidei, Rhamnifolii and Rubus. Other large groups (Sylvatici, Glandulosi and Micantes) were more diverse.     

Variable responses of tuber moth to the leaf trichomes of wild potatoes

This study examines the response of tuber moth, Phthorimaea operculella (Zeller) (Lepidoptera: Gelechiidae), during the initial stages of attack, to variability in trichome density and composition on foliage of Solanum berthaultii (Hawkes) and Solanum tarijense (Hawkes) (Solanaceae). Solanum berthaultii bears two types of glandular trichome (type A and type B) that together reduced oviposition by the moth. Females were often completely deterred from ovipositing on foliage with >300 trichomes per cm². In contrast, neonate establishment on S. berthaultii was generally positively related to trichome densities, indicating that trichomes may be a poor defense against P. operculella when the moth oviposits in soil and neonate larvae select the host plant. Solanum tarijense has only one type of glandular trichome (type A) and eglandular hairs. Most eggs were deposited on the adaxial leaf surfaces that had lower trichome densities. Although the density of type A trichomes was negatively related to oviposition, high densities of hairs on the abaxial and adaxial leaf surfaces appeared to stimulate oviposition, leading to stronger positive relations between hair densities and oviposition. Larvae generally established on the abaxial surface where hair densities were greatest. Relationships between the abaxial densities of leaf hairs and neonate establishment on S. tarijense were positive. The results indicate that the responses by P. operculella to the types and density of trichomes are complex. Whereas type A and type B trichomes may act synergistically to reduce oviposition by the moth, leaf hairs do not defend against oviposition and neither leaf hairs nor type A and B trichomes reduce neonate establishment by this herbivore species.     

Nutlet micro‐morphology of the genus Microula (Boraginaceae) from the Qinghai–Tibetan Plateau,and its systematic implications

The micro‐morphology of nutlets in 30 taxa (28 species and 2 varieties) of Microula Benth. (Boraginaceae) was surveyed by light and scanning electron microscopy. A principal coordinate‐based ordination analysis (PCO) (8 morphological and 14 micro‐morphological characters) was applied to explore whether taxa clustered according to the current sectional taxonomy. The nutlet epidermis showed three major features: trichomes, papillae and lamellae. Two basic trichome types could be identified: branched and unbranched. Stellate trichomes (STT) and stipitate stellate trichomes (SST) can be considered as two subtypes of branched trichomes based on shape and branching location. Three kinds of unbranched trichomes were recognized by their shapes: slender trichomes (SLT), spear‐like trichomes (SPT) and conical trichomes (CT). Four types of papillae were revealed in this genus: central apiculate papillae (CAP), debris‐covered convex papillae (DCP), rugous papillae (RP) and tabular papillae (TP). The lamellae was classified in five types: glossy lamellae (GL), thread‐like lamellae (TL), rugous lamellae (RL), interrupted lamellae (IL) and debris‐covered lamellae (DCL). The variation in the nature of surface sculpturing, nutlet shape and size, dorsal aperture shape, and attachment scar positions proved useful diagnostic characters. Taxa belonging to the same section generally clustered together in the PCO analysis. A key based on the morphology and micro‐morphology of nutlets is provided to distinguish the sections of Microula. The presence of SLT, STT, DCP and DCL is consistent with the macro‐morphological classification and provides additional evidence for the subgeneric delimitations. The SST, SLT, CAP, DCP, RP, DCL, GL and RL are useful in discriminating species in Microula. Additionally, the distribution of some trichomes, papillae and lamellae types is correlated with the altitudinal distribution of the species.