Histopathology of black spot symptoms in sweet oranges

In Brazil, citrus black spot (CBS) caused by Guignardia citricarpa is a major disease that has different symptoms on fruit. In this study, fruit of Citrus sinensis infected by G. citricarpa and showing the symptoms false melanosis, freckle spot and hard spot were cross-sectioned and analysed anatomically and histochemically by light microscopy. Immuno-histological assays were performed. All symptoms were accompanied by a thickening of the cuticle. False melanosis lesions did not contain pycnidia and remained restricted to the epicarp or to the first layers of the mesocarp. The stomata in this type of lesion showed phenolic compounds in the guard cells and in the sub-stomatal chamber. In some samples, the guard cells and their surrounding cells lysed, and a wound meristem began to form underneath them. Freckle spot and hard spot lesions had very similar histological alterations to the epicarp and mesocarp, but in our samples only hard spot lesions contained pycnidia. Both of these symptoms were accompanied by protein inclusions. Epidermal and sub-epidermal cells located in the oil-gland region were obliterated, causing alterations in these structures. All symptoms had regions that stained strongly for lipids and phenols.

GENETIC VARIABILITY FOR JUVENILE CHARACTERS OF Cedrela fissilis VELL. PROGENIES: A SUBSIDY FOR DEFINITION OF SEED COLLECTION ZONE

The objective of this study was to evaluate the genetic differences among three matrix groups of Cedrela fissilis based on quantitative juvenile variables on a progeny test to define seed collecting zones and use of seeds of this species in the study region as well as to evaluate genetic variability of the sampled material. A progeny test was established in a nursery with seeds from 48 seed trees collected in the municipalities of Rio Negrinho, Mafra and Sao Bento do Sul, state of Santa Catarina, and in the municipalities of Lapa, Rio Negro, Campo do Tenente and Antonio Olinto, state of Parana. Of the collected seed trees, 33 sampled trees were distributed in three sites and 15 trees were dispersed in the studied region. It was used a complete random block design, with 8 replicates and 20 plants per plot. Evaluated data included: emergency rate; seedling base diameter and height (61, 102 and 145 days after the seeds were sowed); seedling survival; number of leaves per seedling; aerial section dry mass and root dry mass; and the foliar area of the third fully expanded leaf measured from the apical meristem. The Maximum Restricted Likelihood Method (REML) was used, using the software SELEGEN for analysis. It was found that the juvenile characters are strongly genetically controlled and they can be used to estimate genetic variability of population samples of Cedrela fissilis. The three groups of trees spatially limited did not significantly differ among each other, allowing to conclude that the three areas are part of the same tree seed transfer zone.

Functional characterization of sugarcane mustang domesticated transposases and comparative diversity in sugarcane, rice, maize and sorghum

Transposable elements (TEs) account for a large portion of plant genomes, particularly in grasses, in which they correspond to 50%-80% of the genomic content. TEs have recently been shown to be a source of new genes and new regulatory networks. The most striking contribution of TEs is referred as "molecular domestication", by which the element coding sequence loses its movement capacity and acquires cellular function. Recently, domesticated transposases known as mustang and derived from the Mutator element have been described in sugarcane. In order to improve our understanding of the function of these proteins, we identified mustang genes from Sorghum bicolor and Zea mays and performed a phenetic analysis to assess the diversity and evolutionary history of this gene family. This analysis identified orthologous groups and showed that mustang genes are highly conserved in grass genomes. We also explored the transcriptional activity of sugarcane mustang genes in heterologous and homologous systems. These genes were found to be ubiquitously transcribed, with shoot apical meristem having the highest expression levels, and were downregulated by phytohormones. Together, these findings suggest the possible involvement of mustang proteins in the maintenance of hormonal homeostasis.

Is Drosera meristocaulis a pygmy sundew? Evidence of a long-distance dispersal between Western Australia and northern South America

South America and Oceania possess numerous floristic similarities, often confirmed by morphological and molecular data. The carnivorous Drosera meristocaulis (Droseraceae), endemic to the Neblina highlands of northern South America, was known to share morphological characters with the pygmy sundews of Drosera sect. Bryastrum, which are endemic to Australia and New Zealand. The inclusion of D. meristocaulis in a molecular phylogenetic analysis may clarify its systematic position and offer an opportunity to investigate character evolution in Droseraceae and phylogeographic patterns between South America and Oceania. was included in a molecular phylogenetic analysis of Droseraceae, using nuclear internal transcribed spacer (ITS) and plastid rbcL and rps16 sequence data. Pollen of D. meristocaulis was studied using light microscopy and scanning electron microscopy techniques, and the karyotype was inferred from root tip meristem. The phylogenetic inferences (maximum parsimony, maximum likelihood and Bayesian approaches) substantiate with high statistical support the inclusion of sect. Meristocaulis and its single species, D. meristocaulis, within the Australian Drosera clade, sister to a group comprising species of sect. Bryastrum. A chromosome number of 2n approx. 3236 supports the phylogenetic position within the Australian clade. The undivided styles, conspicuous large setuous stipules, a cryptocotylar (hypogaeous) germination pattern and pollen tetrads with aperture of intermediate type 78 are key morphological traits shared between D. meristocaulis and pygmy sundews of sect. Bryastrum from Australia and New Zealand. The multidisciplinary approach adopted in this study (using morphological, palynological, cytotaxonomic and molecular phylogenetic data) enabled us to elucidate the relationships of the thus far unplaced taxon D. meristocaulis. Long-distance dispersal between southwestern Oceania and northern South America is the most likely scenario to explain the phylogeographic pattern revealed.

Gene expression during early somatic embryogenesis in Brazilian pine (Araucaria angustifolia (Bert) O. Ktze)

Brazilian pine (Araucaria angustifolia (Bert) O. Ktze) is the only native conifer species with economic importance in Brazil. Recently, due to intensive exploitation Brazilian pine was included in the official list of endangered Brazilian plants, under the "vulnerable" category. Biotechnology tools like somatic embryogenesis (SE) are potentially useful for mass clonal propagation and ex situ conservation strategies of commercial and endangered plant species. In spite of that, numerous obstacles still hamper the full application of SE technology for a wider range of species, including Brazilian pine. To enhance somatic embryogenesis in Brazilian pine and to gain a better understanding of the molecular events associated with somatic embryo development, we analyzed the steady-state transcript levels of genes known to regulate somatic embryogenesis using semiquantitative reverse transcription polymerase chain reaction (sqRT-PCR). These genes included Argonaute (AaAGO), Cup-shaped cotyledon1 (AaCUC), wushel-related WOX (AaWOX), a S-locus lectin protein kinase (AaLecK), Scarecrow- like (AaSCR), Vicilin 7S (AaVIC), Leafy Cotyledon 1 (AaLEC), and a Reversible glycosylated polypeptide (AaRGP). Expression patterns of these selected genes were investigated in embryogenic cultures undergoing different stages of embryogenesis, and all the way to maturation. Up-regulation of AaAGO, AaCUC, AaWOX, AaLecK, and AaVIC was observed during transition of somatic embryos from stage I to stage II. During the maintenance phase of somatic embryogenesis, expression of AaAGO and AaSCR, but not AaRPG and AaLEC genes was influenced by presence/ absence of plant growth regulators, both auxins and cytokinins. The results presented here provide new insights on the molecular mechanisms responsible for somatic embryo formation, and how selected genes may be used as molecular markers for Brazilian pine embryogenesis.

Oleoresin glands in copaiba (Copaifera trapezifolia Hayne: Leguminosae), a Brazilian rainforest tree

Although studies have addressed the chemical analysis and the biological activity of oleoresin in species of Copaifera, the cellular mechanisms of oleoresin production, storage, and release have rarely been investigated. This study detailed the distribution, ontogeny, and ultrastructure of secretory cavities and canals distributed in leaf and stem, respectively, of Copaifera trapezifolia, a Brazilian species included in a plant group of great economic interest. Axillary vegetative buds, leaflets, and portions of stem in primary and secondary growth were collected and processed in order to study the anatomy, histolocalization of substances, and ultrastructure. Secretory cavities are observed in the foliar blade and secretory canals in the petiolule and stem. They are made up of a uniseriate epithelium delimiting an isodiametric or elongated lumen. Biseriate epithelium is rarely observed and is a novelty for Leguminosae. Cavities and canals originate from ground meristem cells and the lumen is formed by schizogenesis. The content of the cavities and canals of both stem and leaf is oily and resinous, which suggests that the oleoresin could be extracted from the leaf instead of the stem. Phenolic compounds are also detected in the epithelial cell cytoplasm. Cavities and canals in the beginning of developmental stages have polarized epithelial cells. The cytoplasm is rich in smooth and rough endoplasmic reticula connected to vesicles or plastids. Smooth and rough endoplasmic reticulum and plastids were found to be predominant in the epithelial cells of the secretory cavities and canals of C. trapezifolia. Such features testify the quantities of oleoresin found in the lumen and phenolic compounds in the epithelial cell cytoplasm of these glands. Other studies employing techniques such as correlative light electron microscopy could show the vesicle traffic and the compartmentalization of the produced substances in such glands.

WUSCHEL-related genes are expressed during somatic embryogenesis of the basal angiosperm Ocotea catharinensis Mez. (Lauraceae)

Ocotea catharinensis is a basal angiosperm and an endangered tree species from the Brazilian Atlantic Rain Forest. Despite its economical and ecological importance, mass-propagation of this species is hampered by seldom-produced short-lived seeds, and in vitro propagation is challenged by frequently malformed somatic embryos. Therefore, O. catharinensis somatic embryos are also a good experimental material to study the physiological and molecular mechanisms underlying in vitro morphogenesis. In an ongoing effort to characterize genes expressed during somatic embryogenesis of O. catharinensis we have cloned two Ocotea WUSCHEL-related genes. According to our RT-PCR data, both genes were preferentially expressed in embryogenic cell aggregates. One of them, OcWUS, is a possible ortholog of the Arabidopsis WUSCHEL (WUS) gene, which codes for a homeodomain-containing protein involved in the specification and maintenance of the shoot apical meristem. We analyzed the expression patterns of OcWUS and OcWOX4 by RT-PCR, and OcWUS expression was also assessed by in situ hybridization. The expression patterns of OcWUS were very similar to those described for the Arabidopsis WUS. OcWUS transcripts were generally restricted to a small group of cells in the center of the putative shoot apical meristem of O. catharinensis somatic embryos. Perturbed expression of OcWUS might be related to abnormally formed somatic embryos of O. catharinensis obtained through tissue culture.

The meristematic activity of the endodermis and the pericycle and its role in the primary thickening of stems in monocotyledonous plants

Background: It had long been thought that a lateral meristem, the so-called primary thickening meristem (PTM) was responsible for stem thickening in monocotyledons. Recent work has shown that primary thickening in the stems of monocotyledons is due to the meristematic activity of both the endodermis and the pericycle. Aims: The aim of this work is to answer a set of questions about the developmental anatomy of monocotyledonous plants: (1) Do the stem apices of monocots have a special meristematic tissue, the PTM? (2) Are the primary tissues of the stem the same as those of the root? (3) Is there good evidence for the formation of both the cortex and the vascular tissue from a single meristem, the PTM, in the shoot and from two distinguishable meristems in the root? (4) If the PTM forms only the cortex, what kind of meristem forms the vascular tissue? Methods: Light microscopy was used to examine stem and root anatomy in 16 species from 10 monocotyledonous families. Results: It was observed that radially aligned cortical cells extend outwards from endodermal initial cells in the cortex of the roots and the stems in all the species. The radial gradation in size observed indicates that the cortical cells are derivatives of a meristematic endodermis. In addition, perfect continuity was observed between the endodermis of the root and that of the stem. Meristematic activity in the pericycle gives rise to cauline vascular bundles composed of metaxylem and metaphloem. Conclusion: No evidence was obtained for the existence in monocotyledons of a PTM. Monocotyledons appear to resemble other vascular plants in this respect.

The Tomato (Solanum Lycopersicum cv. Micro-Tom) Natural Genetic Variation Rg1 and the DELLA Mutant Procera Control the Competence Necessary to Form Adventitious Roots and Shoots

Despite the wide use of plant regeneration for biotechnological purposes, the signals that allow cells to become competent to assume different fates remain largely unknown. Here, it is demonstrated that the Regeneration1 (Rg1) allele, a natural genetic variation from the tomato wild relative Solanum peruvianum, increases the capacity to form both roots and shoots in vitro; and that the gibberellin constitutive mutant procera (pro) presented the opposite phenotype, reducing organogenesis on either root-inducing medium (RIM) or shoot-inducing medium (SIM). Mutants showing alterations in the formation of specific organs in vitro were the auxin low-sensitivity diageotropica (dgt), the lateral suppresser (ls), and the KNOX-overexpressing Mouse ears (Me). dgt failed to form roots on RIM, Me increased shoot formation on SIM, and the high capacity for in vitro shoot formation of ls contrasted with its recalcitrance to form axillary meristems. Interestingly, Rg1 rescued the in vitro organ formation capacity in proRg1 and dgtRg1 double mutants and the ex vitro low lateral shoot formation in pro and ls. Such epistatic interactions were also confirmed in gene expression and histological analyses conducted in the single and double mutants. Although Me phenocopied the high shoot formation of Rg1 on SIM, it failed to increase rooting on RIM and to rescue the non-branching phenotype of ls. Taken together, these results suggest REGENERATION1 and the DELLA mutant PROCERA as controlling a common competence to assume distinct cell fates, rather than the specific induction of adventitious roots or shoots, which is controlled by DIAGEOTROPICA and MOUSE EARS, respectively.