Association of 'Candidatus Liberibacter solanacearum' with a vegetative disorder of celery in Spain and development of a real-time PCR method for its detection.

A new symptomatology was observed in celery (Apium graveolens) in Villena, Spain in 2008. Symptomatology included an abnormal amount of shoots per plant and curled stems. These vegetative disorders were associated with 'Candidatus Liberibacter solanacearum' and not with phytoplasmas. Samples from plant sap were immobilized on membranes based on the spot procedure and tested using a newly developed real-time polymerase chain reaction assay to detect 'Ca. L. solanacearum'. Then, a test kit was developed and validated by intralaboratory assays with an accuracy of 100%. Bacterial-like cells with typical morphology of 'Ca. Liberibacter' were observed using electron microscopy in celery plant tissues. A fifth haplotype of 'Ca. L. solanacearum', named E, was identified in celery and in carrot after analyzing partial sequences of 16S and 50S ribosomal RNA genes. From our results, celery (family Apiaceae) can be listed as a new natural host of this emerging bacterium.

On the morphology of the digestive system of two Monomorium ant species.

The digestive system of adults and mature larvae of two ant species of Monomorium Mayr (Hymoneptera: Formicidae) were described with the aid of light and scanning electron microscopy, as there is a lack of studies in this area. These two ant species are recurrently found in urban habitats and are known as 'tramp species,' as they cause problems in households, businesses, and hospitals. The most interesting finds of the present study include the existence of spinules in the crop of adults, and the number of Malpighian tubules and rectal pads was constant among different castes, ages, and species.

Nitrogen-improved photosynthesis quantum yield is driven by increased thylakoid density, enhancing green light absorption.

A nitrogen supply is necessary for all plants. The multifaceted reasons why this nutrient stimulates plant dry weight accumulation are assessed herein. We compared tomato plants grown in full sunlight and in low light environments under four N doses and evaluated plant growth, photosynthetic and calorimetric parameters, leaf anatomy, chloroplast transmission electron microscopy (TEM) and a high resolution profile of optical leaf properties. Increases in N supplies allow tomato plants to grow faster in low light environments (91.5% shading), displaying a robust light harvesting machinery and, consequently, improved light harvesting efficiency. Ultrastructurally, high N doses were associated to a high number of grana per chloroplast and greater thylakoid stacking, as well as high electrodensity by TEM. Robust photosynthetic machinery improves green light absorption, but not blue or red. In addition, low construction and dark respiration costs were related to improved total dry weight accumulation in shade conditions. By applying multivariate analyses, we conclude that improved green light absorbance, improved quantum yield and greater palisade parenchyma cell area are the primary components that drive increased plant growth under natural light-limited photosynthesis.

Carfentrazone-ethyl and glyphosate drift inhibits uredinial formation of Austropuccinia psidii on Eucalyptus grandis leaves.

BACKGROUND: The response to infection of Austropuccinia psidii in resistant (CLR-383) and susceptible (CLR-384) Eucalyptus grandis clones, exposed to herbicide drift of carfentrazone-ethyl, glyphosate and a mixture of these two herbicides, was evaluated at microscopic and physiological levels. RESULTS: Plants of the two clones showed symptoms of phytotoxicity caused by herbicide drift. However, net CO2 assimilation rate, height and shoot dry matter were lower in CLR-384 than in CLR-383. At the ultrastructure level, the leaves of both clones exposed to the herbicides showed thylakoid disorganization and accumulation of starch grains in the chloroplasts. Only plants of CLR-384 were infected by A. psidii, but when exposed to herbicide drift, rust severity was lower than in control plants. Six days after inoculation (dai), plants of this clone exposed to the herbicides had smaller uredinia than control plants. At 12 dai, non-herbicide treated plants showed normal uredinia, containing abundant urediniospores. By contrast, plants exposed to the herbicides were less colonized by the fungus, and the uredinia were smaller with reduced production of urediniospores, which were sometimes not even detected. CONCLUSION: Glyphosate and carfentrazone-ethyl herbicide drift reduce infection and uredinial formation of A. psidii and to some extent induce basal resistance in a susceptible clone of E. grandis. © 2018 Society of Chemical Industry.

Loss of type-IV glandular trichomes is a heterochronic trait in tomato and can be reverted by promoting juvenility.

Glandular trichomes are structures with widespread distribution and deep ecological significance. In the Solanum genus, type-IV glandular trichomes provide resistance to insect pests. The occurrence of these structures is, however, poorly described and controversial in cultivated tomato (Solanum lycopersicum). Optical and scanning electron microscopy were used to screen a series of well-known commercial tomato cultivars, revealing the presence of type-IV trichomes on embryonic (cotyledons) and juvenile leaves. A tomato line overexpressing the microRNA miR156, known to promote heterochronic development, and mutants affecting KNOX and CLAVATA3 genes possessed type-IV trichomes in adult leaves. A re-analysis of the Woolly (Wo) mutant, previously described as enhancing glandular trichome density, showed that this effect only occurs at the juvenile phase of vegetative development. Our results suggest the existence of at least two levels of regulation of multicellular trichome formation in tomato: one enhancing different types of trichomes, such as that controlled by the WOOLLY gene, and another dependent on developmental stage, which is fundamental for type-IV trichome formation. Their combined manipulation could represent an avenue for biotechnological engineering of trichome development in plants.

Morpho-histological, histochemical, and molecular evidences related to cellular reprogramming during somatic embryogenesis of the model grass Brachypodium distachyon.

The wild grass species Brachypodium distachyon (L.) has been proposed as a new model for temperate grasses. Among the biotechnological tools already developed for the species, an efficient induction protocol of somatic embryogenesis (SE) using immature zygotic embryos has provided the basis for genetic transformation studies. However, a systematic work to better understanding the basic cellular and molecular mechanisms that underlie the SE process of this grass species is still missing. Here, we present new insights at the morpho-histological, histochemical, and molecular aspects of B. distachyon SE pathway. Somatic embryos arose from embryogenic callus formed by cells derived from the protodermal-dividing cells of the scutellum. These protodermal cells showed typical meristematic features and high protein accumulation which were interpreted as the first observable steps towards the acquisition of a competent state. Starch content decreased along embryogenic callus differentiation supporting the idea that carbohydrate reserves are essential to morphogenetic processes. Interestingly, starch accumulation was also observed at late stages of SE process. Searches in databanks revealed three sequences available annotated as BdSERK, being two copies corresponding to SERK1 and one showing greater identity to SERK2. In silico analysis confirmed the presence of characteristic domains in a B. distachyon Somatic Embryogenesis Receptor Kinase genes candidates (BdSERKs), which suggests SERK functions are conserved in B. distachyon. In situ hybridization demonstrated the presence of transcripts of BdSERK1 in all development since globular until scutellar stages. The results reported in this study convey important information about the morphogenetic events in the embryogenic pathway which has been lacking in B. distachyon. This study also demonstrates that B. distachyon provides a useful model system for investigating the genetic regulation of SE in grass species.

Cellular and molecular changes associated with competence acquisition during passion fruit somatic embryogenesis: ultrastructural characterization and analysis of SERK gene expression.

The integration of cellular and molecular data is essential for understanding the mechanisms involved in the acquisition of competence by plant somatic cells and the cytological changes that underlie this process. In the present study, we investigated the dynamics and fate of Passiflora edulis Sims cotyledon explants that were committed to somatic embryogenesis by characterizing the associated ultrastructural events and analysing the expression of a putative P. edulis ortholog of the Somatic Embryogenesis Receptor-like Kinase (SERK) gene. Embryogenic calli were obtained from zygotic embryo explants cultured on Murashige and Skoog medium supplemented with 2,4-dichlorophenoxyacetic acid and 6-benzyladenine. Callus formation was initiated by the division of cells derived from the protodermal and subprotodermal cells on the abaxial side of the cotyledons. The isodiametric protodermal cells of the cotyledon explants adopted a columnar shape and became meristematic at the onset of PeSERK expression, which was not initially detected in explant cells. Therefore, we propose that these changes represent the first observable steps towards the acquisition of a competent state within this regeneration system. PeSERK expression was limited to the early stages of somatic embryogenesis; the expression of this gene was confined to proembryogenic zones and was absent in the embryos after the globular stage. Our data also demonstrated that the dynamics of the mobilization of reserve compounds correlated with the differentiation of the embryogenic callus.

Diaporthe endophytica and D. terebinthifolii from medicinal plants for biological control of Phyllosticta citricarpa.

The citrus industry is severely affected by citrus black spot (CBS), a disease caused by the pathogen Phyllosticta citricarpa. This disease causes loss of production, decrease in the market price of the fruit, and reduction in its export to the European Union. Currently, CBS disease is being treated in orchards with various pesticides and fungicides every year. One alternative to CBS disease control without harming the environment is the use of microorganisms for biological control. Diaporthe endophytica and D. terebinthifolii, isolated from the medicinal plants Maytenus ilicifolia and Schinus terebinthifolius have an inhibitory effect against P. citricarpa in vitro and in detached fruits. Moreover, D. endophytica and D. terebinthifolii were transformed by Agrobacterium tumefaciens for in vivo studies. The transformants retained the ability to control of phytopathogenic fungus P. citricarpa after transformation process. Furthermore, D. endophytica and D. terebinthifolii were able to infect and colonize citrus plants, which is confirmed by reisolation of transformants from inoculated and uninoculated leaves. Light microscopic analysis showed fungus mycelium colonizing intercellular region and oil glands of citrus, suggesting that these two new species are capable of colonizing citrus plants, in addition to controlling the pathogen P. citricarpa.

Somatic embryogenesis of a wild passion fruit species Passiflora cincinnata Masters: histocytological and histochemical evidences.

The characterization of cellular changes that occur during somatic embryogenesis is essential for understanding the factors involved in the transition of somatic cells into embryogenically competent cells and determination of cells and/or tissues involved. The present study describes the anatomical and ultrastructural events that lead to the formation of somatic embryos in the model system of the wild passion fruit (Passiflora cincinnata). Mature zygotic embryos were inoculated in Murashige and Skoog induction media supplemented with 2,4-dichlorophenoxyacetic acid and 6-benzyladenine. Zygotic embryo explants at different development stages were collected and processed by conventional methods for studies using light, scanning, and transmission electron microscopy (TEM). Histochemical tests were used to examine the mobilization of reserves. The differentiation of the somatic embryos began in the abaxial side of the cotyledon region. Protuberances were formed from the meristematic proliferation of the epidermal and mesophyll cells. These cells had large nuclei, dense cytoplasm with a predominance of mitochondria, and a few reserve compounds. The protuberances extended throughout the abaxial surface of the cotyledons. The ongoing differentiation of peripheral cells of these structures led to the formation of proembryogenic zones, which, in turn, dedifferentiated into somatic embryos of multicellular origin. In the initial stages of embryogenesis, the epidermal and mesophyll cells showed starch grains and less lipids and protein reserves than the starting explant. These results provide detailed information on anatomical and ultrastructural changes involved in the acquisition of embryogenic competence and embryo differentiation that has been lacking so far in Passiflora.