M. Grilli Caiola

Ultrastructure of chromoplasts and other plastids in Crocus sativus L. (Iridaceae)

Saffron (Crocus sativus L. Iridaceae) chromoplasts and other plastids were studied by electron microscope to determine their structure, origin and pigment localization. Plastids from pistils of floral buds and flowers at anthesis, dried and decoloured stigmas, and green and senescent leaves were examined. Results indicated that mature saffron chromoplasts occur in the red parts of stigmas and have a reticulo-tubular structure. They contain a reticulum of tubules and plastoglobules. Tubules formed dilated vesicles mainly while plastoglobules appeared numerous and scattered on the whole chromoplast. Chromoplasts appeared in red stigma of very young floral buds. They originated from amyloplasts, the only plastids occurring in the colourless basal portion of style, as well as in the parenchyma of ovary and corm. Transition forms of plastid as amylo-chromoplast, occur in the yellow parts of stigma and style. Senescent leaves did not show plastids with structure similar to the chromoplast of red stigma. Red pigmented and scented stigmas might cooperate in saffron reproduction by attracting pollinator.

Localisation of a carbohydrate epitope recognised by human IgE in pollen of Cupressaceae

The objectives of the present study were: (1) to localise, at the subcellular level, the allergens in pollen of Cupressaceae species, using a monoclonal antibody (mAb 5E6) that is specific for carbohydrate epitopes of allergenic components of Cupressus arizonica pollen extract; (2) to determine whether the glycidic epitope recognised by mAb 5E6 was present in pollen of allergenic species taxonomically unrelated to Cupressaceae; and (3) to determine whether human IgE purified from monosensitive patients recognises the same epitope as mAb 5E6 in Cupressaceae pollen. Immunogold labelling of mAb 5E6 showed a high density of gold particles on the orbicules, supporting the hypothesis that they are important vectors of allergens. A high density was also found on the exine and in the cytoplasm, with the latter finding confirming that fragments of pollen ruptured under humid conditions can represent a vector. The glycidic epitope recognised by mAb 5E6 was detected in all of the species taxonomically unrelated to Cupressaceae, although with varying density. Human IgE recognised the same epitope as mAb 5E6. These findings are consistent with observations of diffuse allergenic cross-reactivity among various allergens. The in situ localisation of a common epitope recognised by both a monoclonal antibody and human IgE could be of importance in immunotherapy.

Embryo origin and development in Crocus sativus L. (Iridaceae)

Abstract Fruit and seed set were obtained in the triploid Crocus sativus (saffron) either by hand crossing with pollen of the diploid C. cartwrightianus or by free pollination. The morphology of the capsules and seeds was similar in both Crocus species. The embryo is comparatively small, whereas the endosperm is formed of numerous layers of cells with hemicellulose-thickened walls and a cytoplasm containing few lipid and protein reserves. Seed germination gave rise to a white prophyll from which a green leaf emerged. At the base of this leaf, a small corm without tunics developed, which, after leaf desiccation, became dormant. These corms sown separately in pots were again capable of growth and emitted a single long green leaf. Vegetative development of new corms was similar in the diploid and hybrid saffron plants. In sites of past saffron cultivation the beds were found to contain degenerated corms and corms at different development stages. The absence of fruit and seed set in emasculated and self- and out-pollinated plants suggests that saffron does not form apomictic embryos.

Intracellular localization of calcium, phosphorus and nitrogen in common bean seeds (Phaseolus vulgaris L. cv. Borlotto) by SEM, ESI and EELS techniques

ABSTRACT Scanning, transmission electron microscopy, electron spectroscopic imaging (ESI) and electron energy loss spectroscopy (EELS) were used to localize calcium, phosphorus and nitrogen in seeds of Phaseolus vulgaris. Prismatic, straight and kinked twin calcium oxalate crystals were detected in the crystalligen layer of seed teguments. Cytoplasmic calcium and phosphorus-rich particles, probably of phytin, were detected in cotyledon parenchyma cells by ESI and EELS. Moreover, higher levels of calcium were mapped on osmiophilic droplets close to cell walls. Crystals, cytoplasmic phytin particles, osmiophilic droplets and calcium in cell walls disappeared after treatment with critic acid. By chemical assays, 1.1 g/100 g of phytic acid was detected in control seeds, and an 81% decline was observed in citric acid-treated samples. These data suggest that crystals and phytin complexes could play a role in determining the low calcium availability in common beans. Also, nitrogen found in protein bodies of cotyledons could represent an important site of protein reserves.

Pollen-pistil interactions in Hermodactylus tuberosus Mill. (Iridaceae)

ABSTRACT Hermodactylus tuberosus Mill. (Iridaceae) is a species with a very short style and a tricarpellar unilocular ovary. For ornamental purposes it is usually reproduced by means of tubers, although both vegetative and seed reproduction occur in nature. However, its capsules do not contain many seeds in comparison to the high number of ovules inside the ovary. To understand this conflicting process a study on H. tuberosus pollen-pistil interaction was carried out. Pollen viability was evaluated by different techniques. Germination rate was tested in vitro and in vivo after stigma, intrastylar and intraovarian pollinations. A high percentage of the pollen grains were found to be viable resulting in a high percentage of in vitro germination and a low rate of germination on the stigma. Indeed, many pollen tubes developed after intrastylar and intraovarian pollinations. Several anomalies characterized pollen germination and pollen tube growth in vitro and in vivo. Observations from field trials indicated ...

MANGANESE DETECTED BY ELECTRON SPECTROSCOPY IMAGING AND ELECTRON ENERGY LOSS SPECTROSCOPY IN MITOCHONDRIA OF NORMAL RAT BRAIN CELLS

ABSTRACT Electron spectroscopy imaging (ESI) and electron energy loss spectroscopy (EELS) provide an opportunity to investigate the presence of different trace elements directly in the structure of fixed tissues. We evaluated the usefulness of such techniques in detecting the sub-cellular localization and relative content of manganese in the mitochondria of cells in the brain of normal rats. ESI provided a granular appearance to manganese ions aggregated at any site summed through the thickness of ultra-thin sections. EELS confirmed the presence of manganese in the organelles shown by ESI. To demonstrate the specificity of signals for manganese, a comparative study was also performed. Electron-spectroscopy analysis of manganese, iron and fluorine in the same mitochondria demonstrated the presence of strong and specific signals for manganese, but the absence of detectable iron, or of fluorine, which was expected to be normally absent. However, iron and manganese were both detected in myelin sheaths by ESI and the spectral analysis afforded by EELS revealed distinct peaks for manganese and iron in the same samples. The manganese concentration in mitochondria as measured by the relative optical density on photomicrographs was highly variable probably due to variability in organelle characteristics and subtle differences in section thickness. These results show that ESI and EELS are sensitive and valuable techniques to detect bound manganese within sub-cellular structures.

Pollen structure and germination of Crocus thomasii Ten. (Iridaceae)

Abstract Mature pollen of C. thomasii Ten. (Iridaceae) has been studied from a morphological and physiological point of view and compared to that of C. sativus L., C. thomasii pollen is roundish, with a 83 μm diameter and a 14% of anomalous grains. Percentage of the positive alcoholdehydrogenase (ADH +) pollen grains is about 90% whereas the in vitro germinated rate reaches 55%. Exine is homogeneously thickened (2.5 μm) with randomly distributed thinner zones and 5 μm thick intine. Cytoplasm of vegetative and generative cells is very rich of small and large smooth vesicles and vesicles coated by ribosomes. The generative cell shows a thin ondulated pectocellulosic wall. Its nucleus is intensely fluorescent after treatment with the 4,6-diamidino-2-phenylindole (DAPI) fluorochrome. Although many ultrastructural aspects of the C. thomasii pollen are common in C. sativus. L., C. thomasii pollen is smaller, more regularly structured and germinates in vivo and in vitro in higher percentage than that of C. sativus.

Structure and physiology of Cycad coralloid roots

Abstract The gonidial zone organization of coralloid roots of Cycas revoluta is reported together with the taxonomy of cyanobiont, the physiology of the association and the significance of nitrogen fixation. Recent data on gonidial zone composition obtained by microelectrodes and immunogold labeling are referred in order to determine the mechanisms of defence of nitrogenase against oxygen and ammonium inhibiting effects.

Calcium(II) and the reproductive biology of Crocus biflorus Miller subsp. biflorus (Iridaceae)

ABSTRACT Calcium is known to be involved in pollen germination and pollen tube growth in vitro, and in incompatibility processes such as the rejection of incompatible pollen. Previous studies on calcium distribution in Crocus cartwrightianus Herb, and C. sativus L. pistils have indicated an increase in calcium concentration in the ovary of cross-fertile species after fertilisation. In order to compare the distribution of calcium in other Crocus species, a study was undertaken on pistils of C. biflorus Miller subsp. biflorus by means of calcium(II)-selective microelectrodes and chlorotetracycline. Moreover, the effect of calcium on pollen germination and pollen tube growth in vitro was studied. Results show that C. biflorus subsp. biflorus is a self- and cross-compatible plant; a peak in calcium content in the ovary at day 7 after self- and cross- pollinations was revealed, indicating an involvement of calcium in fertilisation. Furthermore, calcium in the germination media was found to influence germinatio...

Azolla: An efficient N2-fixing association with three components

Abstract Bacterial population is found together with the cyanobacterium Anabaena azollae in the leaf cavities and the sporocarps of Azolla. Different Arthrobacter species have been invariably isolated from the leaf cavities and sporocarps of several fern species. About the role of the bacteria in the association, it is probable that the bacteria affect the carbon and nitrogen metabolism of the cavity by competing with the Anabaena for the nutrients and in the meantime it is likely that their high respiratory activity favours the establishment of a microaerobic environment and consequently the nitrogenase activity. Furthermore Arthrobacter may contribute to the production of the mucilage present in the algal packets by excreting extracellular polysaccharides, which components can be involved in lectin binding with Azolla and Anabaena. Finally the IAA formation by Arthrobacter may influence the hormonal balance during the establishment of the association.