Analysis of the Nicotiana tabacum Stigma/Style Transcriptome Reveals Gene Expression Differences between Wet and Dry Stigma Species

The success of plant reproduction depends on pollen-pistil interactions occurring at the stigma/style. These interactions vary depending on the stigma type: wet or dry. Tobacco (Nicotiana tabacum) represents a model of wet stigma, and its stigmas/styles express genes to accomplish the appropriate functions. For a large-scale study of gene expression during tobacco pistil development and preparation for pollination, we generated 11,216 high-quality expressed sequence tags (ESTs) from stigmas/styles and created the TOBEST database. These ESTs were assembled in 6,177 clusters, from which 52.1% are pistil transcripts/genes of unknown function. The 21 clusters with the highest number of ESTs (putative higher expression levels) correspond to genes associated with defense mechanisms or pollen-pistil interactions. The database analysis unraveled tobacco sequences homologous to the Arabidopsis (Arabidopsis thaliana) genes involved in specifying pistil identity or determining normal pistil morphology and function. Additionally, 782 independent clusters were examined by macroarray, revealing 46 stigma/style preferentially expressed genes. Real-time reverse transcription-polymerase chain reaction experiments validated the pistil-preferential expression for nine out of 10 genes tested. A search for these 46 genes in the Arabidopsis pistil data sets demonstrated that only 11 sequences, with putative equivalent molecular functions, are expressed in this dry stigma species. The reverse search for the Arabidopsis pistil genes in the TOBEST exposed a partial overlap between these dry and wet stigma transcriptomes. The TOBEST represents the most extensive survey of gene expression in the stigmas/styles of wet stigma plants, and our results indicate that wet and dry stigmas/styles express common as well as distinct genes in preparation for the pollination process.

The influence of erbium:yttrium-aluminum-garnet laser ablation with variable pulse width on morphology and microleakage of composite restorations

The objective of this study was to evaluate the influence of various pulse widths with different energy parameters of erbium:yttrium-aluminum-garnet (Er:YAG) laser (2.94 mu m) on the morphology and microleakage of cavities restored with composite resin. Identically sized class V cavities were prepared on the buccal surfaces of 54 bovine teeth by high-speed drill (n = 6, control, group 1) and prepared by Er:YAG laser (Fidelis 320A, Fotona, Slovenia) with irradiation parameters of 350 mJ/ 4 Hz or 400 mJ/2 Hz and pulse width: group 2, very short pulse (VSP); group 3, short pulse (SP); group 4, long pulse (LP); group 5, very long pulse (VLP). All cavities were filled with composite resin (Z-250-3 M), stored at 37A degrees C in distilled water, polished after 24 h, and thermally stressed (700 cycles/5-55A degrees C). The teeth were impermeabilized, immersed in 50% silver nitrate solution for 8 h, sectioned longitudinally, and exposed to Photoflood light for 10 min to reveal the stain. The leakage was evaluated under stereomicroscope by three different examiners, in a double-blind fashion, and scored (0-3). The results were analyzed by Kruskal-Wallis test (P > 0.05) and showed that there was no significant differences between the groups tested. Under scanning electron microscopy (SEM) the morphology of the cavities prepared by laser showed irregular enamel margins and dentin internal walls, and a more conservative pattern than that of conventional cavities. The different power settings and pulse widths of Er:YAG laser in cavity preparation had no influence on microleakage of composite resin restorations.

The Effect of Fluoride Therapies on the Morphology of Bleached Human Dental Enamel

The aim of this in vitro study was to evaluate qualitatively the surface morphology of enamel bleached with 35% hydrogen peroxide (HP) followed by application of fluoridated agents. Forty intact pre molars were randomly distributed into four groups (n = 10), treated as follows: Group I (control group) remained stored in artificial saliva at 37 degrees C, Group II - 35% HP; Group III - 35% HP + acidulated fluoride (1.23%) and Group IV - 35% HP + neutral fluoride (2%). The experimental groups received three applications of bleaching gel and after the last application all specimens were polished. This procedure was repeated after 7 and 14 days, and during the intervals of applications, the specimens were stored in artificial saliva at 37 degrees C. Scanning electron microscopy (SEM) analysis showed superficial irregularities and porosities to varying degrees in bleached enamel compared to control group. Sample evaluation was made by attributing scores, and data were statistically analyzed using Kruskal-Wallis and Dunn tests (P < 0.05). SEM qualitative investigation demonstrated that 35% hydrogen peroxide affected human dental enamel morphology, producing porosities, depressions, and superficial irregularities at various degrees. These morphological changes were higher after the application of 1.23% acidulated fluoride gel. Microsc. Res. Tech. 74:512-516, 2011. (C) 2010 Wiley-Liss, Inc.

Comparison of dentin root canal permeability and morphology after irradiation with Nd:YAG, Er:YAG, and diode lasers

The aim of this study was to compare the effects of Nd:YAG, Er:YAG, and diode lasers on the morphology and permeability of root canal walls. The three laser wavelengths mentioned interact differently with dentin and therefore it is possible that the permeability changes caused will determine different indications during endodontic treatment. Twenty-eight human single-rooted teeth were instrumented up to ISO 40 and divided into four groups: group C, control (GC), non-laser irradiated; group N (GN), irradiated with Nd:YAG laser; group E (GE), with Er:YAG laser and group D (GD) with diode laser. After that, the roots were filled with a 2% methylene blue dye, divided into two halves and then photographed. The images were analyzed using Image J software and the percentage of dye penetration in the cervical, middle, and apical root thirds were calculated. Additional scanning electron microscopy (SEM) analyses were also performed. The analysis of variance (ANOVA) showed significant permeability differences between all groups in the middle and cervical thirds (p < 0.05). The Tukey test showed that in the cervical third, GN presented means of dye penetration statistically significantly lower than all of the other groups. In the middle third, GE and GD showed statistically higher dye penetration means than GC and GN. SEM analysis showed melted surfaces for GN, clean wall surfaces with open dentinal tubules for GE, and mostly obliterated dentinal tubules for GD. Er:YAG (2,094 nm) laser and diode laser (808 nm) root canal irradiation increase dentinal permeability and Nd:YAG (1,064 nm) laser decreases dentin permeability, within the studied parameters.

Incidence, morphology and morphometry of the foramen of Vesalius: complementary study for a safer planning and execution of the trigeminal rhizotomy technique

The foramen of Vesalius (FV) is located in the greater wing of the sphenoid bone between the foramen ovale (FO) and the foramen rotundum in an intracranial view. The FO allows the passage of the mandibular branch of trigeminal nerve, which is the target of the trigeminal radiofrequency rhizotomy. We analyzed its location, morphology, morphometry and interrelation among other foramina. 400 macerated adult human skulls were examined. A digital microscope (Dino-Lite plus(A (R))) was used to capture images from the FV. A digital caliper was used to perform the measurements of the distance between the FV and other foramina (FO, foramen spinosum and the carotid canal) in an extracranial view of the skull base. In the 400 analyzed skulls, the FV was identified in 135 skulls (33.75%) and absent on both sides in 265 skulls (66.25%). The FV was observed present bilaterally in 15.5% of the skulls. The incidence of unilateral foramen was 18.25% of the skulls of which 7.75% on right side and 10.5% on left side. The diameter of the FV was measured and we found an average value of 0.65 mm, on right side 0.63 mm and on the left side 0.67 mm. We verified that positive correlations were statistically significant among the three analyzed distances. This study intends to offer specific anatomical data with morphological patterns (macroscopic and mesoscopic) to increase the understanding of the FV features as frequency, incidence and important distances among adjacent foramina.

Myxobolus cordeiroi n. sp., a parasite of Zungaro jahu (Siluriformes: Pimelodiade) from Brazilian Pantanal: Morphology, phylogeny and histopathology

This work is part of an ongoing investigation into the characteristics of Myxozoan parasites of freshwater fish in Brazil and was carried out using morphology, histopathology and molecular analysis. A new Myxosporea species (Myxobolus cordeiroi) is described infecting the jau catfish (Zungaro jahu). Fifty jau specimens were examined and 78% exhibited plasmodia of the parasite. The plasmodia were white and round, measuring 0.3-2.0 mm in diameter and the development occurred in the gill arch, skin, serosa of the body cavity, urinary bladder and eye. The spores had an oval body and the spore wall was smooth. Partial sequencing of the 18S rDNA gene resulted in a total of 505 bp and the alignment of the sequences obtained from samples in different organs revealed 100% identity. In the phylogenetic analysis, the Myxobolus species clustered into two clades-one primarily parasites of freshwater fish and the other primarily parasites of marine fish. M. cordeiroi n. sp. was clustered in a basal position in the freshwater fish species clade. The histological analysis revealed the parasite in the connective tissue of the different infected sites, thereby exhibiting affinity to this tissue. The plasmodium was surrounded by an outer collagen capsule of fibers with distinct orientation from the adjacent connective tissue and an inner layer composed of delicate collagen fibrils-more precisely reticular fibers. The development of the parasite in the cornea and urinary bladder caused considerable stretching of the epithelium. (C) 2009 Elsevier B.V. All rights reserved.

Host-parasite-environment relationship, morphology and molecular analyses of Henneguya eirasi n. sp parasite of two wild Pseudoplatystoma spp. in Pantanal Wetland, Brazil

A new myxosporean species, Henneguya eirasi n. sp., is described parasitizing the gill filaments of Pseudoplatystoma corruscans and Pseudoplatystoma fasciatum (Siluriformes: Pimelodidae) caught in the Patanal Wetland of the state of Mato Grosso, Brazil. The parasite formed white, elongated plasmodia measuring up to 3 mm. Mature spores were ellipsoidal in the frontal view, measuring 37.1 +/- 1.8 mu m in total length, 12.9 +/- 0.8 mu m in body length, 3.4 +/- 0.3 mu m in width, 3.1 +/- 0.1 mu m in thickness and 24.6 +/- 2.2 mu m in the caudal process. Polar capsules were elongated and equal in size, measuring 5.4 +/- 0.5 mu m in length and 0.7 +/- 0.1 mu m in width. Polar filaments had 12-13 coils. Histopathological analysis revealed that the parasite developed in the sub-epithelial connective tissue of the gill filaments and the plasmodia were surrounded by a capsule of host connective tissue. The plasmodia caused slight compression of the adjacent tissues, but no inflammatory response was observed in the infection site. Ultrastructure analysis revealed a single plasmodial wall connected to the ectoplasmic zone through numerous pinocytotic canals. The plasmodial wall exhibited numerous projections and slightly electron-dense material was found in the ectoplasm next to the plasmodial wall, forming a line just below the wall. Partial sequencing of the 18S rDNA gene of H. eirasi n. sp. obtained from P. fasciatum resulted in a total of 1066 bp and this sequence did not match any of the Myxozoa available in the GenBank. Phylogenetic analysis revealed the Henneguya species clustering into clades following the order and family of the host fishes. H. eirasi n. sp. clustered alone in one clade, which was the basal unit for the clade composed of Henneguya species parasites of siluriform ictalurids. The prevalence of the parasite was 17.1% in both fish species examined. Parasite prevalence was not influenced by season, host sex or host size. (C) 2011 Elsevier B.V. All rights reserved.

Phylogenetic analysis of the Brazilian microendemic Paepalanthus subgenus Xeractis (Eriocaulaceae) inferred from morphology

Paepalanthus subgenus Xeractis (Eriocaulaceae) comprises 28 recognized species endemic to the Espinhaco Range, in Minas Gerais state, Brazil. Most species of the subgenus are restricted to small localities and critically endangered, but still in need of systematic study. The monophyly of the subgenus has already been tested, but only with a few species. Our study presents the first phylogenetic hypothesis within the group, based on morphology. A maximum parsimony analysis was conducted on a matrix of 30 characters for 30 terminal taxa, including all species of the subgenus and two outgroups. The biogeographical hypotheses for the subgenus were inferred based on dispersal-vicariance analysis (DIVA). The analysis provided one most-parsimonious hypothesis that supports most of the latest published subdivisions (sections and series). However, some conflicts remain concerning the position of a few species and the relationships between sections. The distribution and origin(s) of microendemism are also discussed, providing the ground for conservation strategies to be developed in the region. (C) 2011 The Linnean Society of London, Botanical Journal of the Linnean Society, 2011, 167, 137-152.

Comparative leaf anatomy and morphology of some neotropical Rutaceae: Pilocarpus Vahl and related genera

Previous anatomical studies have been restricted to the foliar aspects of Pilocarpus. However, no anatomical studies analyzing the foliar aspects of Pilocarpus in relation to related genera have been carried out. Therefore, the aim of this study was to identify characters for future taxonomic and phylogenetic studies in Rutaceae, particularly in Pilocarpus, and to discuss the characteristics associated with the simple or compound leaf condition for the group. The petiole and the leaf blade of 14 neotropical Rutaceae species were analyzed, and the following characteristics were observed in all leaves studied: stomata on both surfaces; secretory cavities, including mesophyll type; camptodromous-brochidodromous venation pattern; and free vascular cylinder in the basal region of the petiole. Additional promising characters were identified for future taxonomic and phylogenetic studies in the Rutaceae family, especially for the Pilocarpus genera.

Morphology and anatomy of inflorescence and inflorescence axis in Paepalanthus sect. Diphyomene Ruhland (Eriocaulaceae, Poales) and its taxonomic implications

Paepalanthus sect. Diphyomene has inflorescences arranged in umbels. The underlying bauplan seems however to be more complex and composed of several distinct subunits. Despite appearing superficially very similar, the morphology and anatomy of the inflorescences can supply useful information for the understanding of the phylogeny and taxonomy of the group. Inflorescences of Paepalanthus erectifolius, Paepalanthus flaccidus, Paepalanthus giganteus, and Paepalanthus polycladus were analyzed in regard to branching pattern and anatomy. In P. erectifolius, P. giganteus and P. polycladus the structure is a tribotryum, with terminal dibotryum, and with pherophylls bearing lateral dibotrya. In P. flaccidus, the inflorescence is a pleiobotryum, with terminal subunit, and without pherophylls. Secondary inflorescences may occur in all species without regular pattern. Especially when grown in sites without a pronounced seasonality, the distinction between enrichment zone (part of the same inflorescence) and new inflorescences may be obscured. The main anatomical features supplying diagnostic and phylogenetic information are as follows: (a) in the elongated axis, the thickness of the epidermal cell walls and the cortex size; (b) in the bracts, the quantity of parenchyma cells (c) in the scapes, the shape and the presence of a pith tissue. Therefore, P. sect. Diphyomene can be divided in two groups; group A is represented by P. erectifolius, P. giganteus and P. polycladus, and group B is represented by P. flaccidus. The differentiation is based in both, inflorescence structure and anatomy. Group A presents a life cycle and anatomical features similar to species of Actinocephalus. Molecular trees also point that these two groups are closely related. However, inflorescence morphology and blooming sequence are different. Species of group B present an inflorescence structure and anatomical features shared with many genera and species in Eriocaulaceae. The available molecular and morphology based phylogenies still do not allow a precise allocation of the group in the bulk of basal species of Paepalanthus collocated in P. sect. Variabiles. The characters described and used here supply however important information towards this goal. (C) 2009 Elsevier GmbH. All rights reserved.

Stigmatic surface in the Vochysiaceae: reproductive and taxonomic implications

(Stigmatic surface, reproductive biology and taxonomy of the Vochysiaceae). The Vochysiaceae are Neotropical trees and shrubs, common in the savanna areas in Central Brazil (Cerrados). The family has been traditionally divided into two tribes: Erismeae, with three genera, and Vochysieae, with five genera. We investigated the stigmatic surface of six Vochysiaceae species, belonging to four genera of Vochysieae: Vochysia, Salvertia, Callisthene and Qualea. Flowers and buds at different developmental stages were collected. Morphological features were observed on fresh material and stigmatic receptivity was inferred based on esterasic activity. Pistils were fixed and embedded in paraplast and sectioned on a rotary microtome; the sections were stained before histological analysis. Stigmas of open flowers were also observed by scanning electron microscopy. Stigmas of all species were wet and showed esterasic activity at pre-anthesis and anthesis stages. Stigmatic surface was continuous with transmitting tissue of glandular nature. Vochysia and Salvertia stigmatic surfaces were formed by multicelular uniseriate hairs, and species of the remaining genera showed papillate surface. The exudate over mature stigmas in all species flowed without rupture of stigmatic Surface and pollen tubes grew down between hairs or papillae. Differences on the stigmatic surface agreed with a phylogenetic reconstruction that separated two clades and indicated that Vochysieae is not monophyletic. Stigmatic features could not be associated with pollination and breeding systems.

Pollination biology of Syngonanthus elegans (Eriocaulaceae - Poales)

Studies on the pollination biology of Eriocaulaceae are scarce although particularly interesting because of its inclusion in the Poales, a predominantly wind-pollinated order. The pollination biology of Syngonanthus elegans (Bong.) Ruhland was studied during two annual flowering periods to test the hypothesis that insect pollination was its primary pollination system. A field study was carried out, including observations of the morphology and biology of the flowers, insect visits and pollinator behaviour. We also evaluated seed set, seed germination and seedling development for different pollination modes. Although seeds were produced by self-pollination, pollination by small insects contributed most effectively to the reproductive success of S. elegans, resulting in the greatest seed set, with the highest germination percentage and optimum seedling vigour. The. oral resources used by flower visitors were pollen and nectar that was produced by staminate and pistillate flowers. Self-pollination played a minor role and its consequence was inbreeding depression.

Reproductive biology of six Brazilian Myrtaceae: is there a syndrome associated with buzz-pollination?

The floral phenology and reproductive biology of six sympatric arboreal Myrtaceae species were studied in the coastal plain forest (Ubatuba, Brazil, 44 degrees 48`W 23 degrees 22`S), from September 1999 to April 2002. Flowering started in the transition from the driest to the most humid season (Sep/Oct) and lasted until March. The sequence with which the species flowered each year was consistently the same. However, the timing of flowering onset, peak, end, and overlap differed from one year to another. Myrtaceae species were classified as xenogamic according to the pollen:ovule ratios, but two of them seem to present some degree of self-compatibility. Flowers of all species opened at sunrise and lasted for I day. Bombus morio (Apidae: Bombini) was the most common visitor followed by Melipona rufiventris (Apidae: Meliponini). Buzz pollination in Myrtaceae was common at the study area and seems to be related to bees` behaviour and to some aspects of flowers` morphology.

Paleoamerican Morphology in the Context of European and East Asian Late Pleistocene Variation: Implications for Human Dispersion Into the New World

Early American crania show a different morphological pattern from the one shared by late Native Americans. Although the origin of the diachronic morphological diversity seen on the continents is still debated, the distinct morphology of early Americans is well documented and widely dispersed. This morphology has been described extensively for South America, where larger samples are available. Here we test the hypotheses that the morphology of Early Americans results from retention of the morphological pattern of Late Pleistocene modern humans and that the occupation of the New World precedes the morphological differentiation that gave rise to recent Eurasian and American morphology. We compare Early American samples with European Upper Paleolithic skulls, the East Asian Zhoukoudian Upper Cave specimens and a series of 20 modern human reference crania. Canonical Analysis and Minimum Spanning Tree were used to assess the morphological affinities among the series, while Mantel and Dow-Cheverud tests based on Mahalanobis Squared Distances were used to test different evolutionary scenarios. Our results show strong morphological affinities among the early series irrespective of geographical origin, which together with the matrix analyses results favor the scenario of a late morphological differentiation of modern humans. We conclude that the geographic differentiation of modern human morphology is a late phenomenon that occurred after the initial settlement of the Americas. Am J Phys Anthropol 144:442-453, 2011. (c) 2010 Wiley-Liss, Inc.

Morphology, molecular phylogeny, and taxonomic inconsistencies in the study of Bradypus sloths (Pilosa: Bradypodidae)

This study focuses on morphological and molecular data analyses, misidentifications, and phylogenetic inconsistencies regarding Bradypus variegatus (the brown-throated sloth) and B. tridactylus (the pale-throated sloth). Misidentifications were recorded on 75 of 313 museum specimens of Bradypus. Almost 90% of the misidentified specimens were B. variegatus from north-central Brazil, erroneously attributed to B. tridactylus. These misidentified specimens are reported in taxonomic reviews as the southernmost records of B. tridactylus. A history of confusing nomenclature regarding sloth species exists, and these particular misidentifications could be attributable to the similarity in face and throat color between B. variegatus from north-central Brazil and B. tridactylus. The molecular phylogeny of morphologically confirmed sloth specimens exhibits 2 monophyletic lineages representing B. variegatus and B. tridactylus. The split time between these 2 lineages was estimated at 6 million years ago (mya), contradicting previous studies that estimated this divergence to be 0.4 mya. Taxonomic inconsistencies were detected when comparing the molecular phylogeny to previously published DNA sequences ascribed to B. tridactylus. Misidentification or introgression could underlie such phylogenetic incongruities. Regardless of their causes, these discrepancies lead to misstatements regarding geographic distribution, phylogeny, and taxonomy of B. variegatus and B. tridactylus.