Three new species of <Emphasis Type="Italic">Stelis</Emphasis> (Orchidaceae) from Mexico

Three new species of the genus Stelis from Mexico are described and illustrated; they were recently collected in the Tacana volcano region of Chiapas, and are named as S. annedamoniae, S. hagsateri, and S. soconuscana. For each species information is provided about habitat, phenology, and conservation status, and the relationships with similar taxa are discussed. Stelis andamoniae is similar to S. hagsateri and both are sympatric at the Tacana volcano; they are members of the Stelis argentata complex. The first has red-wine flowers, and glabrous sepals, while S. hagsateri has greenish-yellow flowers and glandular-papillose sepals. Stelis soconuscana is similar to S. bidentata, from which it is different by its broader leaves, raceme as long as the leaf, smaller flowers, and its distribution.     

Impatiens chashanensis (Balsaminaceae) sp. nov. from Sichuan, China

Impatiens chashanensis H. Y. Bi & S. X. Yu (Balsaminaceae), a new species from the Sichuan Province in China, is described and illustrated. In comparison with the morphologically close I. dicentra , both species have succulent stems and the lobes of the lateral united petals end in a long hair, but I. chashanensis differs in having white flowers, four lateral sepals, lacerate apex of the inner sepal, shorter spur with swollen, slightly bifid apices. Moreover, leaf epidermal characters also support the recognition of the new species.     

The production of a key floral volatile is dependent on UV light in a sexually deceptive orchid

Background and Aims

Plants use a diverse range of visual and olfactory cues to advertize to pollinators. Australian Chiloglottis orchids employ one to three related chemical variants, all 2,5-dialkylcyclohexane-1,3-diones or ‘chiloglottones’ to sexually attract their specific male pollinators. Here an investigation was made of the physiological aspects of chiloglottone synthesis and storage that have not previously been examined.

Methods

The location of chiloglottone production was determined and developmental and diurnal changes by GC-MS analysis of floral tissue extracts was monitored in two distantly related Chiloglottis species. Light treatment experiments were also performed using depleted flowers to evaluate if sunlight is required for chiloglottone production; which specific wavelengths of light are required was also determined.

Key Results

Chiloglottone production only occurs in specific floral tissues (the labellum calli and sepals) of open flowers. Upon flower opening chiloglottone production is rapid and levels remain more or less stable both day and night, and over the 2- to 3-week lifetime of the flower. Furthermore, it was determined that chiloglottone production requires continuous sunlight, and determined the optimal wavelengths of sunlight in the UV-B range (with peak of 300 nm).

Conclusions

UV-B light is required for the synthesis of chiloglottones – the semiochemicals used by Chiloglottis orchids to sexually lure their male pollinators. This discovery appears to be the first case to our knowledge where plant floral odour production depends on UV-B radiation at normal levels of sunlight. In the future, identification of the genes and enzymes involved, will allow us to understand better the role of UV-B light in the biosynthesis of chiloglottones.     

Phylogenetics of Stelis and closely related genera (Orchidaceae: Pleurothallidinae)

Stelis, one of the largest genera within Pleurothallidinae, was recently recircumscribed to include a few hundred more species, most of which had previously been assigned to Pleurothallis. Here, a new phylogenetic analysis of Stelis and closely related genera based on DNA sequences from nuclear ITS and chloroplast matK, based on a much larger sample, is presented; it includes more than 100 species assigned to Stelis and covers all proposed groupings within the genus, many of which have not previously been represented. Clades are proposed to enable easier discussion of groups of closely related species; each clade is characterized morphologically, ecologically, and geographically to explain the evidence found in the molecular analysis. Discussion of the evolutionary trends of character states found in the genus in its broad sense is given. The current taxonomy of the group is given and the possible taxonomical implications of the findings presented here are discussed.     

Dendrobium tamenglongense sp. nov. (Orchidaceae) from Manipur,India

Dendrobium tamenglongense R. Kishor, Y. N. Devi, H. B. Sharma, J. Tongbram & S. P. Vij sp. nov. from Manipur, India is described and illustrated as a new species. The new species differs from D. jaintianum in having larger and glabrous leaves with acute apices, green sepal apices, a 1–3‐flowered inflorescence, smaller and non‐fragrant flowers and a 3‐lobed labellum with fimbriate midlobe.     

Bletia greenwoodiana (Orchidaceae), a new species from Durango,Mexico

Bletia greenwoodiana, a species found in oak-pine forests in Durango and Jalisco, Mexico, is described and illustrated. It is distinguished by its purplis flowers, with the dorsal sepal and petals narrowly elliptic and acuminate at the apex. Its trilobed lip has wide lateral lobes, forming short sinuses, with rounded apices. The midlobe is very short, one third of the total length of the lip, with an emarginate apex dividing it into two small lobes.     

Caught in the act: pollination of sexually deceptive trap-flowers by fungus gnats in Pterostylis (Orchidaceae)

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.     

A cell-type-specific defect in border cell formation in the Acacia mangium root cap developing an extraordinary sheath of sloughed-off cells

Background and Aims

Root caps release border cells, which play central roles in microbe interaction and root protection against soil stresses. However, the number and connectivity of border cells differ widely among plant species. Better understanding of key border-cell phenotype across species will help define the total function of border cells and associated genes.

Methods

The spatio-temporal detachment of border cells in the leguminous tree Acacia mangium was investigated by using light and fluorescent microscopy with fluorescein diacetate, and their number and structural connectivity compared with that in soybean (Glycine max).

Key Results

Border-like cells with a sheet structure peeled bilaterally from the lateral root cap of A. mangium. Hydroponic root elongation partially facilitated acropetal peeling of border-like cells, which accumulate as a sheath that covers the 0- to 4-mm tip within 1 week. Although root elongation under friction caused basipetal peeling, lateral root caps were minimally trimmed as compared with hydroponic roots. In the meantime, A. mangium columella caps simultaneously released single border cells with a number similar to those in soybean.

Conclusions

These results suggest that cell type-specific inhibitory factors induce a distinct defective phenotype in single border-cell formation in A. mangium lateral root caps.     

Correlation between number and position of floral organs in Arabidopsis

Background and Aims

The study of variation in number, position and type of floral organs may serve as a key to understanding the mechanisms underlying their variation, and will make it possible to improve the analysis of gene function in model plant species by means of a more accurate characterization of mutant phenotypes. The present analysis was carried out in order to understand the correlation between number and position of floral organs in Arabidopsis thaliana.

Methods

An analysis of number and position of organs in flowers of wild type as well as in a series of mutations with floral organ position alterations was carried out, using light and electron microscopy. Variation common to different genotypes was analysed by means of individual diagrams, upon which generalized diagrams depicting variation in number and position of organs, were built by superimposition.

Key Results and Conclusions

It is shown that in the Arabidopsis flower a correlation exists between positions of petals and sepals, as well as between positions of stamens and carpels, whereas the position of carpels does not seem to depend on number and position of petals and stamens. This suggests that the position of organs in the basal (sepals) and apical (carpels) parts of the flower are determined before that in the intermediate zone. This assumption is consistent with the results of mathematical modelling and is supposed to be the consequence of stem-cell activity in the flower.     

Floral elaiophores in Lockhartia Hook. (Orchidaceae: Oncidiinae): their distribution,diversity and anatomy

Background and Aims

A significant proportion of orchid species assigned to subtribe Oncidiinae produce floral oil as a food reward that attracts specialized bee pollinators. This oil is produced either by glabrous glands (epithelial elaiophores) or by tufts of secretory hairs (trichomal elaiophores). Although the structure of epithelial elaiophores in the Oncidiinae has been well documented, trichomal elaiophores are less common and have not received as much attention. Only trichomal elaiophores occur in the genus Lockhartia, and their distribution and structure are surveyed here for the first time.

Methods

Flowers of 16 species of Lockhartia were studied. The location of floral elaiophores was determined histochemically and their anatomical organization and mode of oil secretion was investigated by means of light microscopy, scanning electron microscopy and transmission electron microscopy.

Key Results and Conclusions –

All species of Lockhartia investigated have trichomal elaiophores on the adaxial surface of the labellum. Histochemical tests revealed the presence of lipoidal substances within the labellar trichomes. However, the degree of oil production and the distribution of trichomes differed between the three major groups of species found within the genus. All trichomes were unicellular and, in some species, of two distinct sizes, the larger being either capitate or apically branched. The trichomal cuticle was lamellate, and often appeared distended due to the subcuticular accumulation of oil. The labellar trichomes of the three species examined using transmission electron microscopy contained dense, intensely staining cytoplasm with apically located vacuoles. Oil-laden secretory vesicles fused with the plasmalemma and discharged their contents. Oil eventually accumulated between the cell wall and cuticle of the trichome and contained electron-transparent profiles or droplets. This condition is considered unique to Lockhartia among those species of elaiophore-bearing Oncidiinae studied to date.     

Interspecific gene flow in a multispecies oak hybrid zone in the Sierra Tarahumara of Mexico

Background and Aims

Interspecific gene flow can occur in many combinations among species within the genus Quercus, but simultaneous hybridization among more than two species has been rarely analysed. The present study addresses the genetic structure and morphological variation in a triple hybrid zone formed by Q. hypoleucoides, Q. scytophylla and Q. sideroxyla in north-western Mexico.

Methods

A total of 247 trees from ten reference and 13 presumed intermediate populations were characterized using leaf shape variation and geometric morphometrics, and seven nuclear microsatellites as genetic markers. Discriminant function analysis was performed for leaf shape variation, and estimates of genetic diversity and structure, and individual Bayesian genetic assignments were obtained.

Key Results

Reference populations formed three completely distinct groups according to discriminant function analysis based on the morphological data, and showed low, but significant, genetic differentiation. Populations from the zone of contact contained individuals morphologically intermediate between pairs of species in different combinations, or even among the three species. The Bayesian admixture analysis found that three main genetic clusters best fitted the data, with good correspondence of reference populations of each species to one of the genetic clusters, but various degrees of admixture evidenced in populations from the contact area.

Conclusions

The three oak species have formed a complex hybrid zone that is geographically structured as a mosaic, and comprising a wide range of genotypes, including hybrids between different species pairs, backcrosses and probable triple hybrids.     

Replacement of species along altitude gradients: the role of branch architecture

Background and aims

Plant species typical of cold and warm habitats differ in a suite of morpho-physio-phenological traits, although their evolutionary routes have been poorly explored. Here, it is advocated that traits typical of different climate regimes can be largely driven by contrasting branch architectures. This is explored within Saxifraga. First, an investigation was carried out to determine whether series Ceratophyllae (lateral inflorescences) is segregated to lowlands compared with Pentadactylis (terminal inflorescences). Then, two altitudinal vicariants, S. trifurcata (lowland, with lateral inflorescences) and S. canaliculata (highland, with apical inflorescences), were selected. It was hypothesized that apical flowering of S. canaliculata constrains its growth period, bringing with it traits typical of short growth season plants, and conversely for S. trifurcata.

Methods

The hypothesis was tested by measuring plant compactness and organ pre-formation in seven populations of these species along an altitude gradient.

Key Results

Most variables differed among species. Morphological variables at all scales support that the architecture of S. canaliculata generates a more compact habit. A higher number of primordia and earlier inflorescence pre-formation in S. canaliculata indicate that it begins organogenesis earlier. Data on organogenesis suggest that the different timing of inflorescence initiation may be the origin of the contrasting architectures. Within species, shoot compactness increased, and the length of lateral primordia decreased, as altitude increased. All other metrics were similar among locations of the same species at contrasting altitudes.

Conclusions

The hypotheses linking elevational segregation of species, architecture and pheno-morphological traits were validated at broad (gen. Saxifraga) and local (altitudinal vicariants) scales. This supports the initial idea that shoot architecture may to a large extent condition high altitude adaptive syndrome.Key words: Saxifraga, meristem activity, altitude, architecture, phenology, organogenesis     

The barley MATE gene,HvAACT1, increases citrate efflux and Al3+ tolerance when expressed in wheat and barley

Background and Aims

Aluminium is toxic in acid soils because the soluble Al³⁺ inhibits root growth. A mechanism of Al³⁺ tolerance discovered in many plant species involves the release of organic anions from root apices. The Al³⁺-activated release of citrate from the root apices of Al³⁺-tolerant genotypes of barley is controlled by a MATE gene named HvAACT1 that encodes a citrate transport protein located on the plasma membrane. The aim of this study was to investigate whether expressing HvAACT1 with a constitutive promoter in barley and wheat can increase citrate efflux and Al³⁺ tolerance of these important cereal species.

Methods HvAACT1

was over-expressed in wheat (Triticum aestivum) and barley (Hordeum vulgare) using the maize ubiquitin promoter. Root apices of transgenic and control lines were analysed for HvAACT1 expression and organic acid efflux. The Al³⁺ tolerance of transgenic and control lines was assessed in both hydroponic solution and acid soil.

Key Results and Conclusions

Increased HvAACT1 expression in both cereal species was associated with increased citrate efflux from root apices and enhanced Al³⁺ tolerance, thus demonstrating that biotechnology can complement traditional breeding practices to increase the Al³⁺ tolerance of important crop plants.     

Whole Genome Sequence of Treponema pallidum ssp. pallidum,Strain Mexico A,Suggests Recombination between Yaws and Syphilis Strains

Background

Treponema pallidum ssp. pallidum (TPA), the causative agent of syphilis, and Treponema pallidum ssp. pertenue (TPE), the causative agent of yaws, are closely related spirochetes causing diseases with distinct clinical manifestations. The TPA Mexico A strain was isolated in 1953 from male, with primary syphilis, living in Mexico. Attempts to cultivate TPA Mexico A strain under in vitro conditions have revealed lower growth potential compared to other tested TPA strains.

Methodology/Principal Findings

The complete genome sequence of the TPA Mexico A strain was determined using the Illumina sequencing technique. The genome sequence assembly was verified using the whole genome fingerprinting technique and the final sequence was annotated. The genome size of the Mexico A strain was determined to be 1,140,038 bp with 1,035 predicted ORFs. The Mexico A genome sequence was compared to the whole genome sequences of three TPA (Nichols, SS14 and Chicago) and three TPE (CDC-2, Samoa D and Gauthier) strains. No large rearrangements in the Mexico A genome were found and the identified nucleotide changes occurred most frequently in genes encoding putative virulence factors. Nevertheless, the genome of the Mexico A strain, revealed two genes (TPAMA_0326 (tp92) and TPAMA_0488 (mcp2-1)) which combine TPA- and TPE- specific nucleotide sequences. Both genes were found to be under positive selection within TPA strains and also between TPA and TPE strains.

Conclusions/Significance

The observed mosaic character of the TPAMA_0326 and TPAMA_0488 loci is likely a result of inter-strain recombination between TPA and TPE strains during simultaneous infection of a single host suggesting horizontal gene transfer between treponemal subspecies.     

Correlated responses of root growth and sugar concentrations to various defoliation treatments and rhythmic shoot growth in oak tree seedlings (Quercus pubescens)

Background and Aims

To understand whether root responses to aerial rhythmic growth and contrasted defoliation treatments can be interpreted under the common frame of carbohydrate availability; root growth was studied in parallel with carbohydrate concentrations in different parts of the root system on oak tree seedlings.

Methods

Quercus pubescens seedlings were submitted to selective defoliation (removal of mature leaves, cotyledons or young developing leaves) at appearance of the second flush and collected 1, 5 or 10 d later for morphological and biochemical measurements. Soluble sugar and starch concentrations were measured in cotyledons and apical and basal root parts.

Key Results

Soluble sugar concentration in the root apices diminished during the expansion of the second aerial flush and increased after the end of aerial growth in control seedlings. Starch concentration in cotyledons regularly decreased. Continuous removal of young leaves did not alter either root growth or apical sugar concentration. Starch storage in basal root segments was increased. After removal of mature leaves (and cotyledons), root growth strongly decreased. Soluble sugar concentration in the root apices drastically decreased and starch reserves in the root basal segments were emptied 5 d after defoliation, illustrating a considerable shortage in carbohydrates. Soluble sugar concentrations recovered 10 d after defoliation, after the end of aerial growth, suggesting a recirculation of sugar. No supplementary recourse to starch in cotyledons was observed.

Conclusions

The parallel between apical sugar concentration and root growth patterns, and the correlations between hexose concentration in root apices and their growth rate, support the hypothesis that the response of root growth to aerial periodic growth and defoliation treatments is largely controlled by carbohydrate availability.     

A New Secondary Pollen Presentation Mechanism from a Wild Ginger (Zingiber densissimum) and Its Functional Roles in Pollination Process

Background and Aims

Secondary pollen presentation (SPP), a floral mechanism of reproductive adaptation, has been described for more than 200 years, with nine types SPP recorded. However, few studies have been done experimentally to link the floral mechanism of SPP to its functional roles in pollination process. This study aims to describe a new SPP mechanism from a wild ginger (Zingiber densissimum, Zingiberaceae) and explore how the pollen arrangement of SPP affects pollen removal during the interaction with different pollinators.

Methodology/Principal Findings

Field observations and experiments revealed that flowers lasted for less than one day. The breeding system was partially self-incompatible. Two bee species, Macropis hedini (which carried pollen dorsally) and Amegilla zonata (which carried pollen ventrally) were the primary pollinators. About a third of pollen grains were relocated from the anther to the labellum staminode of flowers through the adherence of aggregated pollen chains, while other grains were presented on the anther. In a single visit, each bee species removed pollen grains from both the labellum staminode and the anther. Macropis hedini was more effective than Amegilla zonata.

Conclusions/Significance

Our study describes a new SPP mechanism in angiosperms. The new SPP mode enables pollen grains presented on the anther and the labellum staminode simultaneously via the adherence of aggregated pollen chains, thus promoting pollen to be taken away by different pollinators. This SPP mechanism plays a key role during pollen removal and may have evolved under the pressure to improve male fitness.     

Pollinator convergence and the nature of species' boundaries in sympatric Sardinian Ophrys (Orchidaceae)

Background and Aims

In the sexually deceptive Ophrys genus, species isolation is generally considered ethological and occurs via different, specific pollinators, but there are cases in which Ophrys species can share a common pollinator and differ in pollen placement on the body of the insect. In that condition, species are expected to be reproductively isolated through a pre-mating mechanical barrier. Here, the relative contribution of pre- vs. post-mating barriers to gene flow among two Ophrys species that share a common pollinator and can occur in sympatry is studied.

Methods

A natural hybrid zone on Sardinia between O. iricolor and O. incubacea, sharing Andrena morio as pollinator, was investigated by analysing floral traits involved in pollinator attraction as odour extracts both for non-active and active compounds and for labellum morphology. The genetic architecture of the hybrid zone was also estimated with amplified fragment length polymorphism (AFLP) markers, and pollination fitness and seed set of both parental species and their hybrids in the sympatric zone were estimated by controlled crosses.

Key Results

Although hybrids were intermediate between parental species in labellum morphology and non-active odour compounds, both parental species and hybrids produced a similar odour bouquet for active compounds. However, hybrids produced significantly lower fruit and seed set than parental species, and the genetic architecture of the hybrid zone suggests that they were mostly first-generation hybrids.

Conclusions

The two parental species hybridize in sympatry as a consequence of pollinator overlap and weak mechanical isolation, but post-zygotic barriers reduce hybrid frequency and fitness, and prevent extensive introgression. These results highlight a significant contribution of late post-mating barriers, such as chromosomal divergence, for maintaining reproductive isolation, in an orchid group for which pre-mating barriers are often considered predominant.Key words: AFLP markers, floral scent variation, hybrid zone, hybrid fitness, Ophrys iricolor, Ophrys incubacea, reproductive isolation, sexual deception     

Chromosomal evolution in Pleurothallidinae (Orchidaceae: Epidendroideae) with an emphasis on the genus Acianthera: chromosome numbers and heterochromatin

In this study, we analysed chromosome number variation and chromomycin A₃/4′,6‐diamidino‐2‐phenylindole (CMA/DAPI) banding patterns in 48 species belonging to 12 genera of subtribe Pleurothallidinae (Orchidaceae) in order to understand the chromosome evolution based on recent phylogenetic hypotheses and taxonomic treatments. All species had small chromosomes, with numbers ranging from 2n = 20 in two Specklinia spp. to 2n = 80 in an unidentified Octomeria sp. In Acianthera, the most highly represented genus in this study, a great diversity of chromosome number and pattern of fluorescent bands was observed, showing heterochromatin accumulation in Acianthera section Sicariae subsection Pectinatae. Interspecific ascending and, mainly, descending dysploidy were the main mechanisms of chromosome number evolution in subtribe Pleurothallidinae. For Pleurothallidinae, x = 20 is suggested as the basic chromosome number, the same suggested for the related subtribe Laeliinae and for the whole tribe Epidendreae. The Brazilian species of the mega‐genus Stelis had chromosomes with small amounts of heterochromatin and chromosome numbers based on x₂ = 16. These are generally divergent from those reported for Andean and Meso‐American species, but in agreement with the monophyletic hypothesis proposed for Stelis spp. with a Brazilian Atlantic distribution. © 2015 The Linnean Society of London, Botanical Journal of the Linnean Society, 2015, 178 , 102–120.