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The Alismataceae family, widely distributed across tropical temperate swamps and wetlands, includes 15 genera post-merger with Limnocharitaceae. In Argentina, six genera are represented across three clades. Embryological characters, notably the male gametophyte and anther, are crucial in taxonomy due to their stability against environmental changes. This study aims to analyze the ultrastructure of the tapetum and pollen grain development in three economically and ecologically important species representing each clade: Sagittaria montevidensis (Clade A), Hydrocleys nymphoides (Clade B), and Alisma plantago-aquatica (Clade C). Anthers at different developmental stages were processed according to classic techniques for their observation with bright-field and transmission electron microscopy. The three studied species within the Alismataceae family exhibit similar reproductive characteristics. Seven stages of pollen grain development were identified. The microsporogenesis is successive with a regular meiosis. The ultrastructure of the tapetal cells shows similarities to other species with plasmodial tapetum. During the microspore tetrad stage, there is tapetal hyperactivity and an increase in secretion processes. In the free microspore stage, the tapetal cells lose their walls and increase the amount of rough endoplasmic reticulum forming a network of cisternae that extend into evaginations. Later cells completely invade the anther locule and fuse to form a tapetal plasmodium. No peritapetal membrane with orbicules was observed. Pollen is released at the tricellular stage. The pollen grain wall presents an ectexine with a basal layer, columellae, and tectum with supratectal spines while an endexine is not observed in any of the three species. This research enhances the understanding of tapetal cell interactions with developing pollen grains and contributes to the knowledge of the ultrastructure of plasmodial tapetum. Moreover, these findings highlight evolutionary reproductive patterns in Alismataceae, suggesting the plasmodial tapetum as a synapomorphy for the order., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Floral nectaries are essential for plant reproduction but little is known about the relationship between these secretory structures and pollination system in cacti. To test phenotypic patterns in nectaries associated with pollination syndromes and/or with its pollinators, we selected from evolutionarily related genera Cleistocactus, Denmoza, and Echinopsis, a set of species with bird-pollinated flowers and floral traits that may fit with ornithophily or with sphingophily, and other set of sphingophilous species with moths as effective pollinator. Observations were made under light microscope and scanning and transmission electron microscopes. Nectaries are located at the base of the filaments welded to the tube, forming a chamber. The nectary consists of the epidermis with distinctive features in each genus, a secretory parenchyma which may be vascularized and a non-secretory vascularized parenchyma. Anatomical variants observed in nectaries of different species are not consistent with the floral pollination syndromes neither with groups of pollinators. The basic structure of the nectar chamber is relatively conserved, a fact that may be explained by phylogenetic conservatism among the genera investigated. Our results revalue the role of anatomical traits for the systematics of Cactaceae.

Studies focusing on terminal drought combined with heat impacts on plants of agronomic value remain scarce, and even less under field conditions. The objective of this study was to investigate leaf structural and ultrastructural changes induced by heat stress (HS) and drought stress (DS) during seed filling and their relationship with physiological variables and yield determination. Two soybean cultivars were grown in field conditions. During seed filling four treatments were applied, including a control (without manipulation, at ambient temperature and field capacity), HS (episodes exceeding 32°C for 6 h d-1) during 21-d, DS (20% of field capacity soil water content) during 35-d, and HS×DS. Drought principally reduced leaf area, whereas heat decreased leaf thickness, possible as acclimation strategies, but also irreversible reducing CO2 assimilation sites. Both stresses damaged the outer and inner membranes of chloroplasts, causing swollen chloroplasts and accumulation of plastoglobules, loss of chlorophyll content, and negatively affecting chlorophyll fluorescence. Thus, the performance and integrity of the photosynthetic machinery were reduced. Through a morpho-functional perspective and a holistic multiscale approach, our results provide evidence of photosynthesis impairment and yield drops under stressful conditions which were associated with structural and ultrastructural (particularly at the level of chloroplasts) modifications of leaves.

The aim of this paper is to investigate the ultrastructural events that occur during pollen grains development, with emphasis in pollen grain wall and tapetum ontogeny in Ziziphus jujuba, Z. mucronata, Paliurus spina-christi (Paliureae) and Gouania ulmifolia (Gouanieae). Anthers at different developmental stages were processed according to classic techniques for transmission electron microscopy. Differences in the number of endothecium layers and in the number of tapetal cell nuclei were found. Tapetal cells present an anastomosing tubular network and large vesicles with fibrillar content in the cytoplasm. Pollen grain development and ontogeny of pollen grain wall are similar in the four species. The number of endothecium layers, the number of nuclei of the tapetal cells and tapetal cells ultrastructure of the four species support the phylogenetic relationships previously published for the Rhamnaceae family. Tapetal vesicles with fibrillar or polysaccharide content seem to be an exclusive characteristic of the tribes Paliureae and Gouanieae. Some ultrastructural characters of the pollen grain wall development are common to other species of Rhamnaceae, such as the primexine matrix present at the microspore mother cell stage, the aperture entirely built up during the tetrad stage, the thick and fibrillar intine, and the granular infractectum.

To study the ontogeny of the extrafloral nectaries present in the inflorescences of Vigna luteola (Jacq.) Benth (Leguminosae, Phaseolinae), the location, morphology, anatomy of the earliest stages, histology of the definitive structures and ultrastructure of the secretory stage were analyzed. The extrafloral nectaries at different developmental stages were examined with light microscopy and scanning electron microscopy. The secretory stage was also examined with transmission electron microscopy. The racemose inflorescence of V. luteola has six nodes. At each node, a short globose secondary axis bears two flowers and one to three extrafloral nectaries. Each extrafloral nectary originates from the abscission of a flower bud and is formed by two differentiated zones: a ring of epidermal cells surrounding a group of longitudinally enlarged papillose central cells, both with underlying secretory parenchyma. The primary secretory tissue consists of the central cells, while the ring contributes to secretion to a lesser degree. Secretion is granulocrine, by means of exocytotic vesicles and plasmalemma invaginations. Four developmental stages succeed; the third one being the secretory. The extrafloral nectaries activity period starts when the flowers of the same secondary axis open and ceases before fruit development.

Although the presence of scent was described for several species of Rhamnaceae, localization, morphology and structure of osmophores were unknown. We studied different species of the tribes Rhamneae (Rhamnoids clade), Pomaderreae, Colletieae, Paliureae (Ziziphoids clade) and the species Alphitonia excelsa (unknown tribe, Ziziphoids clade). We expect to have a better comprehension of these structures and provide information on which morphological and anatomical characters may support the phylogeny of the family. We localized the osmophores in the margins and top of the sepals using neutral red. Histochemical tests were made on transverse hand-cut sections of fresh sepals. Observations were made with stereoscopic and bright field microscopes, scanning and transmission electron microscopes. Papillae were observed in the zones with positive reaction to reagents. Different kinds of hairs are present in the sepal epidermis besides papillae. Epidermal cells present a striate cuticle with canals and cavities. Druses are abundant in most species. The ultrastructure of epidermal and subepidermal cells shows high metabolic activity: there are vesicles, mitochondria, endoplasmic reticulum, dictyosomes, plastids with lipids and starch. The vascularization is well developed and reaches the top of the sepal where the principal area of volatile components production is localized. The location and abundance of papillae are the most important traits that allow us recognize and characterize the osmophores in Rhamnaceae. There are no clear anatomical and morphological features exclusive of one clade or tribe. Therefore, in contrast to other sporophytic structures of this family, osmophores do not seem to have any systematic value.

The ultrastructure of the style and pollen tube pathway before, during and after anthesis were studied in 13 species belonging to the tribes Pomaderreae, Paliureae, Colletieae and Gouanieae (Ziziphoid clade) and Rhamneae (Rhamnoid clade) using light microscopy and transmission electron microscopy. The aim of this study is to provide new morphological characters useful for phylogenetic analysis at suprageneric level in Rhamnaceae. The patterns of pollen tube growth and the ultrastructural changes undergone by cells of the style were also described. Species of Rhamneae (Scutia buxifolia and Condalia buxifolia) have a solid style, with the transmitting tissue forming three independent strands (S. buxifolia) or a central, single horseshoe-shaped strand as seen in transversal section (C. buxifolia) which could derive from the fusion of formerly independent strands. In contrast, Pomaderreae, Gouanieae and Paliureae showed semi-solid styles, while in Colletieae, as previously reported, the style is hollow with two or three stylar canals. The style anatomy and the ultrastructure of the pollen tube pathway show that there is a tendency towards a solid style with a single strand of transmitting tissue within the family. The three-canalled hollow style could be the plesiomorphic state of the character "type of style" in the family, the semi-solid style the synapomorphic state and the solid style with three strands of transmitting tissue the apomorphic state, with the solid style with a single strand of transmitting tissue as the most derived state. Therefore, Colletieae would be the most basal tribe of the Ziziphoid clade.

The style morphology and anatomy vary among different species. Three basic types are: open, closed, and semi-closed. Cells involved in the pollen tube pathway in the different types of styles present abundant endoplasmic reticulum, dictyosomes, mitochondria, and ribosomes. These secretory characteristics are related to the secretion where pollen tube grows. This secretion can be represented by the substances either in the canal or in the intercellular matrix or in the cell wall. Most studies suggest that pollen tubes only grow through the secretion of the canal in open styles. However, some species present pollen tubes that penetrate the epithelial cells of the canal, or grow through the middle lamella between these cells and subepithelial cells. In species with a closed style, a pathway is provided by the presence of an extracellular matrix, or by the thickened cell walls of the stylar transmitting tissue. There are reports in some species where pollen tubes can also penetrate the transmitting tissue cells and continue their growth through the cell lumen. In this review, we define subtypes of styles according to the path of the pollen tube. Style types were mapped on an angiosperm phylogenetic tree following the maximum parsimony principle. In line with this, it could be hypothesized that: the open style appeared in the early divergent angiosperms; the closed type of style originated in Asparagales, Poales, and Eudicots; and the semi-closed style appeared in Rosids, Ericales, and Gentianales. The open style seems to have been lost in core Eudicots, with reversions in some Rosids and Asterids.

The perianth organs of six species of Nymphaeaceae, representing Euryale, Nymphaea and Victoria, were studied on the basis of macroscopical, micromorphological, and anatomical characters. The aims were to determine whether perianth is differentiated among tepal whorls considering the presence of sepaloid and petaloid characters, and to evaluate the occurrence of both features in individual tepals. Selected perianth series were examined macroscopically, with light microscopy, and scanning electron microscopy. Osmophores were detected using neutral red and Sudan. In all tepals examined, stomata and hydropotes were present on the abaxial and adaxial surfaces. These are anomocytic or stephanocytic; hydropotes of irregular type are also present. The outer series of tepals display morpho-anatomical characters in most part related with photosynthetic and protective functions. Osmophore activity is very scarce and petaloid epidermal morphology is present only in N. lotus, thus allowing interpretation of this whorl as primarily sepaloid. The second series exhibits both petal-like and sepal-like characters; in N. amazonum and N. gardneriana sepaloid and petaloid group of cells are present on the abaxial surface of individual tepals. Therefore, this whorl is transitional between the outer and the innermost ones. Both the morpho-anatomy and presence of osmophore activity indicate that the innermost series is entirely petaloid. Inner tepals of E. ferox, N. alba, and V. cruziana share the presence of epidermal cells with predominantly smooth cuticle, whereas those of N. amazonum, N. gardneriana, and N. lotus share a cuticular ornamentation consisting of numerous papillae on each cell. Morphological characters of the perianth epidermis are in some respects congruent with the molecular phylogeny of Nymphaeaceae. Our results support the co-expression of sepaloidy and petaloidy within individual tepals and the mosaic model of perianth evolution proposed for the angiosperms.

Variations in pollen characters and tapetum behavior were recently acknowledged in the early-divergent family Nymphaeaceae and even within the genus Nymphaea, which probably is not monophyletic; some traits such as infratectum and tapetum type are also a matter of different interpretations. In this study, developmental characters of the pollen grains and tapetum in Nymphaea subgenus Hydrocallis are provided for the first time. Observations were made in N. amazonum, N. gardneriana, and N. prolifera using light, scanning, and transmission electron microscopy. Tapetum is of the secretory type and produces orbicules. At microspore and pollen grain stages, the distal and proximal walls differ considerably. This result supports the operculate condition of the aperture in Hydrocallis, and such aperture might be plesiomorphic for Nymphaeoideae. The infratectum is intermediate, composed of inter-columellae granular elements, robust columellae consisting of agglomerated granules, complete columellae, and fused columellae. Narrow microchannels are present and persist until the mature pollen grain stage. The membranous granular layer is often present in the pollen grains of Nymphaeaceae. In N. gardneriana, this layer is most probably a component of the intine because it is lost after acetolysis. Orbicules in the Nymphaeaceae are characterized as spherical or subspherical, with a smooth sporopolleninic wall that surrounds an electron-lucent core and with individual orbicules that usually merge to give irregular aggregations. The aperture, pollen wall ultrastructure, and the tapetum of the studied species are discussed in an evolutionary and systematic context, and these characters are also compared with those of other angiosperm lineages.

Despite that there is some literature on pollen morphology of Rhamnaceae, studies addressing general aspects of the microsporogenesis, microgametogenesis, and anther development are rare. The aim of this paper is to describe the ultrastructure of pollen grain ontogeny with special attention to tapetum cytology in Hovenia dulcis. Anthers at different stages of development were processed for transmission and scanning electron microscopy, bright-field microscopy, and fluorescence microscopy. Different histochemical reactions were carried out. The ultrastructural changes observed during the development of the tapetal cells and pollen grains are described. Large vesicles containing carbohydrates occur in the tapetal cell cytoplasm during the early stages of pollen development. Its origin and composition are described and discussed. This is the first report on the ontogeny and ultrastructure of the pollen grain and related sporophytic structures of H. dulcis.

Cabomba Aubl. is a genus that presents a range of features that have made it to be considered a potential genetic model for studies of early angiosperm evolution. Therefore, any study that expands our knowledge of this genus is potentially useful for the understanding of the evolution of early angiosperms. This paper reports the study of the anatomy and the ultrastructure of the stigma and the style of Cabomba caroliniana Gray during the 2 days of anthesis using bright-field microscope, fluorescence microscope and transmission electron microscope. The stigma is dry and has pluricellular papillae. The style is hollow with a central canal coated by an epithelium. The papillae have fewer organelles than those typical of glandular cells, and they are covered by a cuticle that is broken when pollen germinates. The ultrastructure of epithelial cells indicates that the cells lining the canal are secretory. The canal is filled with a fibrillar and granular substance. The pollen tubes grow inside the canal through this substance. The results are discussed in the context of what is known for other species of angiosperms.