Ultrastructure of the female gonad of two temnocephalids (Platyhelminthes,Rhabdocoela)

The female gonad of Temnocephala dendyi and T. minor consists of a single germarium and two rows of vitellaria. It is enveloped by an extracellular lamina and accessory cells. Accessory cells are only peripherally located in the germarium while their cytoplasmic projections also fill the spaces between vitellocytes in the vitellarium. The main feature of oocyte maturation is the appearance of chromatoid bodies and the development of rough endoplasmic reticulum (R.E.R.) and Golgi complexes which appear to be correlated with the production of double-structured egg granules. The egg granules, which are localized in the cortical cytoplasm of mature oocytes, contain glycoproteins, are devoid of polyphenols and are similar in structure and composition to the cortical granules observed in some Digenea and Monogenea. Vitellocytes are typical secretory cells with well-developed R.E.R. and Golgi complexes which are involved in the production of shell globules and yolk. The multigranular pattern and the polyphenolic composition of the shell globules of the temnocephalids investigated are similar to those observed in other rhabdocoels, and in some Prolecithophora and Neodermata. This feature may represent a synapomorphy shared by these taxa. This revised version was published online in July 2006 with corrections to the Cover Date.     

Ultrastructural and cytochemical studies of the female gonad of Prorhynchus sp. (Platyhelminthes,Lecithoepitheliata)

The female gonad of Prorhynchus is heterocellular (neoophoran organization) and consists of an unpaired, elongate germovitellarium enveloped by a finely granular extracellular lamina. It is composed of a posterior germinative area where early oocytes are randomly associated with differentiating vitellocytes and a growth area with follicular organization. In each follicle a single oocyte is surrounded by a layer of vitellocytes. By electron microscopy, the oocytes showed features typical of non-vitellogenic germ cells; they had chromatoid bodies, annulate lamellae, lipid droplets and R.E.R. and Golgi complexes producing small granules with a multilamellar pattern. Vitellocytes showed features typical of secretory cells with the R.E.R. and Golgi complex developed to a great extent and involved in the production of type A and type B globules, respectively. We speculate that type A globules are shell-globules and type B globules are yolk. The structure, composition and role of vitellocyte globules of Prorhynchus are compared with those of homologous inclusions from other Platyhelminthes.Abbreviations A type A globule - B type B globule - ECL extracellular lamina - GC Golgi complex - L lipid - RER rough endoplasmic reticulum - O oocyte - V vitellocyte     

Oogenesis in Actinoposthia beklemischevi (Platyhelminthes, Acoela): an ultrastructural and cytochemical study

The oogenesis of the acoel Actinoposthia beklemischevi can be divided into a previtellogenic and a vitellogenic stage. Maturing oocytes are surrounded by accessory cells (a.c.) that produce electrondense granules, the content of which is released into the space between the oocyte and a.c. and gives rise to a thin primary egg envelope. The a.c. may also contribute to yolk synthesis by transferring low molecular weight precursors to the oocyte. Two types of inclusion are produced in maturing oocytes. Type I inclusions are small, roundish granules produced by the Golgi complex. They have a proteinaceous non-polyphenolic content which is discharged in the intercellular space and produce a thicker secondary egg envelope. Type I inclusions represent eggshell-forming granules (EFGs). Type II inclusions are variably sized globules progressively changing their shape from round to crescent. They appear to be produced by the ER, contain glycoproteins and remain scattered throughout the cytoplasm in large oocytes. Type II inclusions represent yolk. The main features of oogenesis in Actinoposthia are: (a) EFGs have a non-polyphenolic composition; (b) the egg envelope has a double origin and is not sclerotinized; (c) yolk production appears to be autosynthetic. The present ultrastructural findings are compared with those from other Acoelomorpha and Turbellaria.     

Ultrastructural and cytochemical aspects of the female gonad of Geoplana burmeisteri (Platyhelminthes, Tricladida, Terricola)

The ultrastructure of the female gonad of the land planarian Geoplana burmeisteri was investigated by means of electron microscopy and cytochemical techniques. It consists of two small germaria located ventral to the intestine and of two irregular, lateral rows of vitelline follicles, both enveloped by a tunica composed of an extracellular lamina and an inner sheath of accessory cells. Accessory cell projections completely surround developing oocytes and vitellocytes. The main feature of oocyte maturation is the appearance of chromatoid bodies and the development of the rough endoplasmic reticulum (RER) and Golgi complexes. These organelles appear to be correlated with the production of egg inclusions of medium electron density, about 1.5-1.8 microm in diameter, which remain scattered in the ooplasm of mature oocytes. On the basis of cytochemical tests demonstrating their glycoprotein composition, these inclusions were interpreted as residual yolk globules. Vitellocytes are typical secretory cells with well-developed RER and Golgi complexes that are mainly involved in the production of yolk globules and eggshell globules, respectively. Eggshell globules appear to arise from repeated coalescence of small Golgi-derived vesicles and, at an intermediate stage of maturation, show a multigranular pattern. Later, after vesicle fusion, they reach a diameter of 1.3-1.6 microm when completely mature and show a meandering/concentric pattern, as is typical of the situation seen in most Proseriata and Tricladida. The content of yolk globules is completely digested by pronase, while the content of eggshell globules is unaffected. Mature vitellocytes contain, in addition, a large quantity of glycogen and lipid droplets as further reserve material. On the basis of the ultrastructural characteristics of the female gonad described above and in relation to the current literature, we conclude that G. burmeisteri appears to be more closely related to the freshwater triclads, in particular to members of the Dugesiidae, than to the marine triclads.     

Ultrastructure of the ciliary pits in the Geocentrophora group (Platyhelminthes,Lecithoepitheliata)

The ultrastructure of the paired lateral ciliary pits in several endemic species of Geocentrophora from Lake Baikal and in one cosmopolitan species, G. baltica, has been compared and the possible functional significance is discussed. The pit is composed of two distinctive parts; the bottom of the pit is an extensive sensitive area, filled with uni-and biciliary sensory receptors with reduced rootlets and numerous neurotubules. The walls of the pit are formed by several large dark cells, characterized by a dark cytoplasm with numerous mitochondria, a large nucleus, intracellular canaliculi, basal infoldings of the cell membrane, glycogen granules and a varying number of cilia. A protruding, densely ciliated ridge occurs along the anterior wall of the pit. The cilia have a strengthened rootlet system and seem to provide a strong water current into the pit. Dark cell processes penetrate the basement membrane of the pit and come into the vicinity of large cells with a cytoplasm similar to that of the dark cells of the pit. These large cells in their turn come close to the terminal parts of the protonephridial canals, containing a weir. Smaller protonephridial capillaries without a weir seem to open directly into the pit lumen. The morphological data obtained suggest that the ciliary pit in not only a sensory structure, but plays a part in osmoregulation and ion exchange as well.     

Fine-structural characteristics of female and male germ cells in Proseriata Otoplanidae (Platyhelminthes)

Fine-structural features of female germ cells differentiating within the germaria of Otoplanella baltica and Notocaryoplanella glandulosa are documented and compared with those of other free-living platyhelminths having ectolecithal eggs.In these species, encompassed in the taxon Proseriata Lithophora, insemination of the germocytes occurs within the germaria. A sperm cell, that has penetrated a germocyte differs in special features from mature male germ cells found in the testes, in parts of the male genital system, or even in other regions of the organism. The hypothesis that dense bodies correspond to acrosomal material is supported.     

Ultrastructure of the pharynx of Prorhynchus (Platyhelminthes, Lecithoepitheliata)

The pharynx variabilis of Prorhynchus is strongly muscular, with a small pharyngeal fold and a thin surrounding sheath. There is one row of inner longitudinal musclcs, up to six rows of inner circular muscles, many radial muscles, one row of outer circular and one row of outer longitudinal muscles, with no sphincter muscle groups. Three kinds of secretion, produced in a cluster of gland cell bodies posterior to the pharynx, enter the pharynx wall. They travel anteriorly in ducts and two kinds unite in a common duct just prior to discharging into the anterior region of the pharynx lumen. The perikarya of lumen epithelial cells lie within the pharynx musculature and, at the anterior and posterior margins of the pharynx, external to the pharynx. Bundles of ciliated receptors are numerous at the anterior and posterior constrictions. Similarities in the ultrastructure of flame bulbs of Rhabdocoela and Lecithoepitheliata suggest a relationship between these groups. However, the usefulness of pharynx ultrastructure for platyhelminth phylogeny cannot be assessed until complete ultrastructural studies of various groups of Rhabdocoela have been made.     

The evolution of protonephridia of the Platyhelminthes

Three types of flame bulbs are distinguished in the Platyhelminthes: type 1 has two cilia arising from a terminal cell and rootlets extending along the weir; type 2 has many cilia arising from a terminal cell and the proximal canal cell closely aligned with it; and type 3 has a non-terminal perikaryon forming many flame bulbs, each with many cilia and a single row of longitudinal ribs. Each type appears in various structural forms. Type 1 is found in the Catenulida; type 2 in the Macrostomida, Polycladida, Prolecithophora, Proseriata, Tricladida, Fecampiidae, and Neodermata; and type 3 in the Rhabdocoela and Lecithoepitheliata. The most likely evolutionary sequence is that type 3 is derived from type 2 and, perhaps, that type 2 is derived from type 1. Characters of the protonephridia show that the Rhabdocoela and the Neodermata form separate phylogenetic lineages; other similarities between these taxa are due to convergent evolution (or horizontal gene transfer?).     

Spermatozoa of tapeworms (Platyhelminthes,Eucestoda): advances in ultrastructural and phylogenetic studies

New data on spermiogenesis and the ultrastructure of spermatozoa of ‘true’ tapeworms (Eucestoda) are summarized. Since 2001, more than 50 species belonging to most orders of the Eucestoda have been studied or reinvestigated, particularly members of the Caryophyllidea, Spathebothriidea, Diphyllobothriidea, Bothriocephalidea, Trypanorhyncha, Tetraphyllidea, Proteocephalidea, and Cyclophyllidea. A new classification of spermatozoa of eucestodes into seven basic types is proposed and a key to their identification is given. For the first time, a phylogenetic tree inferred from spermatological characters is provided. New information obtained in the last decade has made it possible to fill numerous gaps in the character data matrix, enabling us to carry out a more reliable analysis of the evolution of ultrastructural characters of sperm and spermiogenesis in eucestodes. The tree is broadly congruent with those based on morphological and molecular data, indicating that convergent evolution of sperm characters in cestodes may not be as common as in other invertebrate taxa. The main gaps in the current knowledge of spermatological characters are mapped and topics for future research are outlined, with special emphasis on those characters that might provide additional information about the evolution of tapeworms and their spermatozoa. Future studies should be focused on representatives of those major groups (families and orders) in which molecular data indicate paraphyly or polyphyly (e.g. ‘Tetraphyllidea’ and Trypanorhyncha) and on those that have a key phylogenetic position among eucestodes (e.g. Diphyllidea, ‘Tetraphyllidea’, Lecanicephalidea, Nippotaeniidea).     

Characters of the female gonad and the phylogeny of Platyhelminthes

The female gonad of Platyhelminthes shows a wide variability in its anatomical and cellular organization and submicroscopic structure, and some characteristics may be useful for elucidating the questioned monophyly of some taxa and reconstructing the phylogenetic tree of the group. The morphological characters of the female gonad have been subdivided into three levels: macroscopic, microscopic and ultrastructural, and some examples with potential phylogenetic value are presented for each level. This revised version was published online in July 2006 with corrections to the Cover Date.     

Ultrastructure of the nerve cells and sensilla of Geocentrophora baltica (Platyhelminthes,Lecithoepitheliata) and the surface sensilla in the Geocentrophora group

Two types of nerve cell could be distinguished ultrastructurally in the central nervous system of Geocentrophora baltica (Prorhynchida, Lecithoepitheliata). Both show invaginations in the plasma membrane, but they differ in the character of the cytoplasm (light or densely stained) and the distribution of the neuronal vesicles (evenly or in groups). Different kinds of vesicles and neuronal release sites are observed. Special features of the synapses are pronounced local thickenings of the presynaptic membrane connected to paramembranous densities. In G. baltica and five endemic Geocentrophora spp. from Lake Baikal six types of surface sensillum were observed at the epidermal surface: 1. those with a long thin rootlet; 2. a short, balloon-shaped cilium with an aberrant axoneme and a reduced rootlet; 3. a rootlet branching into many striated bundles; 4. a thick rootlet; 5. a reduced rootlet and numerous neurotubules;and 6. collared sensilla each with one cilium in a deep pit surrounded by a collar of 11 to 12 microvilli. The variable number of microvilli in the collared sensillum is considered plesiomorphic relative to the stable number of eight microvilli known in sensilla of the Prolecithophora, Proseriata, and Rhabdocoela. The ultrastructure of the collar sensillum indicates that the Lecithoepitheliata is only distantly related to the Prolecithophora and higher turbellarians.     

The female gonad in two species of Microplana (Platyhelminthes,Tricladida, Rhynchodemidae): Ultrastructural and cytochemical investigations

The female gonad of the land planarians Microplana scharffi and Microplana terrestris consists of two small germaria located ventrally in the anterior third of the body and of two ventro‐lateral rows of oblong vitelline follicles distributed between the intestinal pouches. Both these structures are enveloped by a tunica composed of an outer extracellular lamina and an inner sheath of accessory cells. Oocyte maturation is characterized by the appearance of chromatoid bodies and the development of endoplasmic reticulum and Golgi complexes. These organelles appear to be correlated with the production of egg granules with a fenestrated/granular content of medium electron density, about 4–5 μm in diameter, which remain dispersed in the ooplasm of mature oocytes. On the basis of cytochemical tests showing their glycoprotein composition, and their localization in mature oocytes, these egg granules have been interpreted as yolk. In the vitelline follicles, vitellocytes show the typical features of secretory cells with well‐developed rough endoplasmic reticulum and Golgi complexes involved in the production of eggshell globules and yolk. The eggshell globules, which appear to arise from repeated coalescences of two types of Golgi‐derived vesicles, contain polyphenols and, when completely mature, they measure about 1–1,2 μm in diameter and show a meandering/concentric content pattern as is typical of the situation observed in most Proseriata and Tricladida. Mature vitellocytes also contain a large amount of glycogen and lipids as further reserve material. On the basis of the ultrastructural features of the female gonad and in relation to the current literature the two species of rhynchodemids investigated appear to be closely related to the freshwater planarians belonging to the family Dugesiidae. J. Morphol. 2009. © 2009 Wiley‐Liss, Inc.     

A phylogeny of the Platyhelminthes: towards a total-evidence solution

We advocate a total-evidence approach for the reconstruction of working phylogenies for the Turbellaria and the phylum Platyhelminthes. Few morphology-based character matrices are available in the systematic literature concerning flatworms, and molecular-based phylogenies are rapidly providing the only means by which we can estimate phylogenies cladistically. Character matrices based on gross morphology and ultrastructure are required and should be internally consistent, i.e. character coding should follow a set of a priori guidelines and character duplication and contradiction is avoided. In order to test our molecular phylogenies we need complementary data sets from morphology. To understand morphological homology we need phylogenetic evidence from independent (e.g. molecular) data. Fully complementary morphological and molecular data sets enable us to validate phylogenetic hypotheses and the combination of these sets in phylogenetic reconstruction utilises all statements of homology. Working phylogenies which include all phylogenetic information not only shed light on individual character evolution, but form a strong basis for comparative studies investigating the origin and evolutionary radiation of the taxonomic group under scrutiny. This revised version was published online in July 2006 with corrections to the Cover Date.     

Aspects of the phylogeny of Platyhelminthes based on 18S ribosomal DNA and protonephridial ultrastructure

DNA studies of 23 taxa (20 platyhelminths, 1 nemertean, Homo and Artemia) and electron-microscopic studies of the protonephridia of many platyhelminths (supported by some additional ultrastructural data) have led to the following conclusions: the Neodermata are monophyletic; Temnocephalida and Dalyelliida form one clade and are not the primitive sister group of the Neodermata; Gyrocotylidea, Amphilinidea and Eucestoda form one monophylum; Pterastericolidae and Umagillidae are dalyelliids and not the sister group of the Neodermata; and Proseriata are unlikely to be closely related with the Tricladida. A large taxon consisting of the Proseriata and some other turbellarians may represent the sister group of the Neodermata.     

Evolution of body-wall musculature in the Platyhelminthes (Acoelomorpha, Catenulida, Rhabditophora)

In an effort to understand the phylogeny of the Platyhelminthes, the patterns of body-wall musculature of flatworms were studied using fluorescence microscopy and Alexa-488-labeled phalloidin. Species of the Catenulida have a simple orthogonal gridwork of longitudinal and circular muscles. Members of the Rhabditophora have the same gridwork of musculature, but also have diagonal muscles over their entire body. Although a few species of Acoelomorpha possessed a simple orthogonal grid of musculature, most species typically have distinctly different patterns of dorsal and ventral body-wall musculature that include sets of longitudinal, circular, U-shaped, and several kinds of diagonal muscles. Several distinct patterns of musculature were identified, including 8 patterns in 11 families of acoels. These patterns have proven to be useful in clarifying the phylogeny of the Acoelomorpha, particularly with regard to the higher acoels. Patterns of musculature as well as other morphological characters are used here for revisions of acoel systematics, including the return of Eumecynostomum sanguineum (Mecynostomidae) to the genus Aphanostoma (Convolutidae), the revision of the family Childiidae, and the formation of a new family, Actinoposthiidae.     

Unciliated body surface in three species of the Umagillidae (Dalyellioida,Platyhelminthes)

Anoplodiera voluta Westblad, Seritia elegans (Westblad) and Wahlia macrostylifera Westblad are species of the family Umagillidae living in the intestine of the holothurian Parastichopus tremulus. In all three species, part of the epidermis is unciliated, but unlike unciliated epidermis in the major parasitic flat-worm groups, it is cellular and has intraepithelial nuclei. The surface of the unciliated cells in A. voluta is convex with the cells separated by lateral gaps. The cells have two distinct regions: the basal, organelle-rich part and the apical part which contains few identifiable organelles except vesicles. In W. macrostylifera the unciliated cells have a flat surface and, between them, narrower gaps that in some cases widen to paracellular compartments below the cell surface. Apically the cells contain electron-dense vesicles, often in contact with the surface. S. elegans has unciliated cells separated by gaps. In all these species, apical vesicles indicate secretory activity. Comparison of the epidermis of these three species with the neodermis of the major parasitic flatworm groups within the Neodermata does not support a close relationship of the three to the Neodermata.     

Morphology and fine structure of the gravid uterus of three hymenolepidid tapeworm species (Platyhelminthes: Cestoda)

Summary

The ultrastructure of the uterus in gravid proglottids of Hymenolepis citelli, Vampirolepis nana and Vampirolepis microstoma was examined using transmission and scanning electron microscopy. The cellular and subcellular structures were similar in all three species, consisting basically of a syncytial layer attached to a basal extracellular matrix. All nuclei were juxtaluminal and each contained a single large centrally located nucleolus and prominent masses of central and peripheral heterochromatin. The cytoplasm showed evidence suggesting a high level of protein synthesis and secretion. It consisted primarily of granular endoplasmic reticulum with moderately dilated cisternae; most cisternae were completely filled with an electron-lucent flocculent material, but others contained electrondense granules. Free ribosomes and mitochondria were also present. The apical plasma membrane and a small amount of enclosed cytoplasm were folded into long microlamellae that extended into the uterine lumen. Larger epithelial folds and villi consisting of folded portions of the entire epithelium projected into the uterine lumen. The uterine epithelium along with its basul extracellular matrix and underlying cellular parenchyma were folded into the lumen at several points, forming epitheliomesenchymal folds and villi that closely apposed many of the eggs.     

Ultrastructure of spermiogenesis and spermatozoa in Prorhynchus sp. (Platyhelminthes,Lecithoepitheliata, Prorhynchidae)

Summary

Mature sperm of Prorhynchus sp. have an elongated nucleus, multiple mitochondria and dense bodies, and two free axonemes which are located in grooves of the main shaft for much of their length. The axonemes are subterminally inserted and have the typical 9+ ‘1’ arrangement unique to Platyhelminthes and synapomorphic for taxa of Trepaxonemata. The testis follicles examined had small numbers of developing spermatids and very few mature sperm were present. During spermiogenesis, spermatids remain joined in clusters by distinctive bridges. In each spermatid two centrioles (with an intercentriolar body between them) give rise to free axonemes which grow out in opposite directions from each other. Indistinct ciliary rootlets are present. The axonemes are carried distally from the main spermatid mass on an elongating process and turn back towards the main spermatid mass. Nucleus, mitochondria and dense bodies move into the shaft, and the spermatid elongates before detaching from others in the cluster. This is the first detailed study of sperm and spermiogenesis in Lecithoepitheliata. Mature sperm are distinctly different from those of prolecithophorans, to which they are reputedly related, the latter having aflagellate sperm without dense bodies.     

The freshwater planarian Dugesia sicula Lepori from Sardinia (Platyhelminthes,Tricladida)

Fissiparous strains of freshwater triclads of the Dugesia gonocephala group were collected from 12 localities in Sardinia all situated not more than 5 kilometers from the coast. Some environmental factors and the sexual status of the specimens were noted at the time of collection. During the laboratory rearing 30% of individuals of each strain became sexual (ex-fissiparous individuals). All the examined strains showed common karyological and morphological characteristics suggestive of the species Dugesia sicula Lepori. The chromosome complement, which was a constant 27+2–3 B chromosomes, was classified as aneutriploid, due to its clearly documented characteristics. The fissiparous populations of D. sicula appear to have a high degree of tolerance to variations in environmental factors, especially temperature. Within the D. gonocephala group this species has the broadest distribution in the Mediterranean region.