A freeze-fracture study of the tegumental membrane of Schistosoma mansoni (Platyhelminthes:Trematoda).

Two fracture faces in each half of the freeze-fractured tegumental membrane of adult Schistosoma mansoni indicate the presence of two trilaminate membranes. This result is compatible with the heptalaminate appearance of the tegumental membrane in ultrathin sections. Intramembranous particles are located mainly in the outermost leaflet of the outer membrane and in the cytoplasmic leaflet of the inner membrane. The tegumental membrane of the cercaria (infective larva) has a single fracture plane, which conforms with its trilaminate appearance in sections. Intramembranous particles are extremely numerous and are almost all located in the cytoplasmic leaflet.     

Cyton II: A subtegumental cell type in the cercaria of Schistosoma mansoni

In Schistosoma mansoni cercaria, an aggregate of subtegumental cells is found in a small, dorsoanterior area of the body (middivision). These cells are nestled between two laterally positioned flame cells and the muscle that delimits the anterior end of the body, and the anterior end of the central ganglion. This highly amorphous cell type, designated as cyton II, has a heterochromatic nucleus and a cytoplasm that is elaborated into coarse, tortuous processes. Its cytoplasm contains ribosomes, mitochondria, sparse amounts of endoplasmic reticulum, and two types of circular-to-oval concentric membranous bodies. One type has an electron-dense core and measures 200–250 nm on the short axis, and the other is completely membranous and measures 100–125 nm on the short axis. The cell body of cyton II communicates with the tegument that covers a small, dorsoposterior area of the anterior organ (oral sucker); however, we could not confirm a tegumental connection with the body division. When cercariae transform into schistosomules, the concentric membranous bodies of cyton II migrate into the anterior organ's tegument via cytoplasmic processes of the cell. The major function of previously described cells that have similar membranous bodies is to supply additional membranes to the outer tegument during development into an adult worm. A multilaminated outer membrane is an adaptation to the survival of the schistosomule and adult worm in the bloodstream of the vertebrate host (Hockley amd McLaren [′73]). The presence of membranous bodies from cyton II in the tegument does not confirm that this cell type participates in the formation of multilaminated membranes. Its precise function remains to be determined. © 1995 Wiley-Liss, Inc.     

Electron microscopy of the tumulus and origin of associated structures within the tegument of Eubothrium salvelini Schrank, 1790 (Cestoidea: Pseudophyllidea)

Tedesco J. L. and Coggins J. R. 1979. Electron microscopy of the tumulus and origin of associated structures within the tegument of Eubothrium salvelini Schrank, 1790 (Cestoidea: Pseudophyllidea). International Journal for Parasitology10: 275–280. Validity of the tumulus, a new organ recently described on the tegument of Eubothrium salvelini, is confirmed. Spherical, dense inclusions approximately 0.23–0.29 μm were associated with the tumulus and with subtegumental cell bodies. Origin of these inclusions within subtegumental cell bodies and their transport via ducts to the tumulus is described. Inclusions are synthesized within granular endoplasmic reticulum and packaged by Golgi apparatus prior to transport. Inclusions were observed only in association with the tumulus within the tegumental distal cytoplasm.     

Characterisation of a family of Schistosoma japonicum proteins related to dynein light chains.

Dynein light chains (DLC) are components of dynein, an enzyme complex involved in various aspects of microtubule-based motility. We report here the molecular cloning and sequencing of cDNAs encoding a family of DLC-like polypeptides (SjcDLC1-5) from the human bloodfluke Schistosoma japonicum with open reading frames of 87-104 amino acids and deduced molecular masses ranging from 10.5 to 12.3 kDa. Two-dimensional Western blot analysis confirmed the presence of several S. japonicum DLC isoforms with differing pI values and molecular sizes. We also describe the molecular characterisation, genomic organisation and expression of clone SjcDLC1, and the immunological characterisation and localisation of its encoded protein. Northern blot analysis of adult worm RNA indicated SjcDLC1 is encoded by a single message of approximately 650 bp and Southern analysis suggested one SjcDLC1 gene exists in the S. japonicum genome. Immunolocalisation studies demonstrated that the SjcDLC1 protein is present in the tegument of the adult and cercarial stages of S. japonicum. SjcDLC1 and the other SjcDLC may function in the transport of specialised organelles, comprising membranous and discoid bodies, through the tegument to the schistosome-unique heptalaminate tegumental membrane at the external surface of the adult worm. As a consequence, they may provide novel targets for anti-schistosome vaccine and/or drug development.     

Ultrastructural localization of Sm15 and Sm25, two major tegumental adult worm antigens of Schistosoma mansoni.

Sm15 and Sm25 are two of the principal tegumental antigens recognized by antibodies from mice protectively vaccinated with adult worm tegumental membranes and may therefore be potential vaccine candidate antigens. Using antibodies affinity purified from anti-tegumental membrane anti-sera, and antibodies raised against the recombinant antigens, Sm15 and Sm25 were shown to be located specifically in the tegument of adult worms being distributed throughout the syncitium but not associated with the outer membrane.     

Ultrastructural aspects of early immune damage to Taenia crassiceps metacestodes

Siebert, A. E. Jr., Good A. H. and Simmons J. E. 1978. Ultrastructural aspects of early immune damage to Taenia crassiceps metacestodes. International Journal for Parasitology8: 45–53. Changes in the ultrastructure of the tegument and subtegumental cells of intraperitoneally implanted Taenia crassiceps metacestodes were studied by transmission electron microscopy over a 4-week period. Death of metacestodes without involvement of host inflammatory cells is indicated initially by vacuolization of the larval tegument followed by loss of the tegument and subsequent death of the larvae. Changes in the tegument involve loss of the glycocalyx, reduction in the numbers of mitochondria and microtriches present, and loss of secretory capacity. Subtegumental cells show an accumulation of secretory vacuoles and marked disruption of nuclear morphology. Tegument damage is attributed to a complement-mediated lysis of the outer tegument membrane and death of the larvae probably results from loss of tegument function.     

Hymenolepis diminuta: ultrastructural observations on complement-mediated tegumental lysis and destrobilation of 4-day-old worms in vitro

Changes in the ultrastructure of the tegument and subtegumental cells of 4-day-old Hymenolepis diminuta were studied in vitro in 50% fresh normal rat serum over a 5-h period and compared with heat-inactivated serum and saline controls. First, membrane-bound vesicles accumulate above the microthrix-border. After 30–40 min large vacuoles, which may contain membranous elements, appear in the tegument at a time when the surface of the young strobila is virtually denuded of the microthrix-border. With prolonged incubations there are subtegumental secretory inclusions with dark, enveloping cytoplasm in the tegument and finally the apical plasma membrane, together with the majority of the matrix, is lost. The disrupted portion of the worm is abruptly demarcated from the comparatively intact scolex/anterior neck region by a constriction. Even after 5 h incubation there is no evidence of loss of tegumental matrix components from regions anterior to the constriction but the neck region shows a significant denudation of the microthrix layer and the tegument contains numerous inclusions. The scolex tegument only showed little evidence of loss of membrane from the surface. Possible mechanisms for the avoidance of complement-mediated lysis in the anterior region are discussed.     

Schistosoma mansoni Annexin 2: Molecular characterization and immunolocalization

We here describe the cloning and characterization of the Schistosoma mansoni Annexin 2, previously identified in the tegument by proteomic studies, and as an up-regulated gene in schistosomulum stage by microarray data. In silico analysis predicts a conserved core containing four repeat domains of Annexin (ANX) and a variable N-terminal region similar to that described for mammalian isoforms. Real-time RT-PCR and Western blot analysis determined that S. mansoni Annexin 2 is significantly up-regulated in the transition from free-living cercaria to schistosomulum and adult worm parasitic stages. Immunolocalization experiments and tegument membrane preparations confirmed Annexin 2 as a protein mainly localized in the tegument of schistosomula and adult worms. Furthermore, it binds to the tegument surface membranes in a calcium-dependent manner. These results suggest that S. mansoni Annexin 2 is closely associated to the tegument arrangement, being a potential target for immune intervention.     

东方杯叶吸虫体被的超微结构

扫描电镜显示东方杯叶吸虫体被有许多族生棘,具纤毛乳突和无纤毛窝状乳突,附着器皮层特化为微毛。透射电镜观察表明,皮层是由合胞体、基膜和肌肉层组成,合胞体通过细胞通道与皮层细胞相连。文中详细描述了这些结构,揭示无纤毛窝状乳突为腺乳突,皮层细胞间存在线粒体细胞和附着器皮层形成微毛,探讨了这些结构的生理功能。体外培养成虫皮层结构的某些不正常变化,如皮层细胞的空虚松散,可作为评价吸虫体外培养的一种指标。     

Wound healing in the trematode Schistosoma

The ability of adult Schistosoma mansoni to effect wound healing over an exposed surface has been demonstrated. In transected worm segments a new external plasma membrane formed over the exposed tegumental cytoplasm. An elevated leading edge of tegument developed around the margin of the wound; the surface of this region was highly convoluted and there was a proliferation of membranous bodies within its cytoplasm. Inward migration of the leading edge over the exposed internal tissues took place. The resulting new tegument lacked spines and sensory endings. There was no regeneration of basal lamina or tegumentary cytons. In vitro maintenance of worm segments for 3 weeks did not give rise to any major ultrastructural changes in the tissues away from the wound.     

Ultrastructure and cytochemistry of the tegument of Orthocoelium scoliocoelium and Paramphistomum cervi (Trematoda: Digenea)

The tegument of Orthocoelium scoliocoelium and Paramphistomum cervi was examined using histochemical techniques and electron microscopy. On the basis of the distribution of acid and alkaline phosphatase (E.C. 3.1.3.2, E.C. 3.1.3.1), non-specific esterase (E.C. 3.1.1.1), cholinesterase (E.C. 3.1.1.7) and succinate dehydrogenase (E.C. 1.3.99.1) at light microscope level two distinct regions were recognized, an outer and an inner zone. Electron microscopy revealed that the tegument comprises an outer surface syncytium underlain by a thick subsyncytial zone and musculature. Deeper still occur the nucleated "tegumental cells". The latter are in cytoplasmic continuity with the surface syncytium via vacuolated cytoplasmic trabeculae which traverse the muscle layers and the subsyncytial zone. Three types of tegumental cells each lacking mitochondria were observed. The T1 cells synthesize discoid and electron dense T1 bodies while T2 cells produce oval and electron lucent T2 bodies. The third type of tegumental cells apparently produce no secretory bodies and may represent an embryonic cell type. The surface syncytium contains T1 and T2 secretory bodies and is bounded apically by a plasma membrane invested externally by a fuzzy and filamentous glycocalyx. The surface syncytium lacks mitochondria and is traversed by infoldings of the basal plasma membrane. Beneath the surface syncytium the subsyncytial zone is largely comprised of fibrous interstitial material. This zone, which is particularly thick in the amphistomes, is traversed by trabeculae and extensions of underlying parenchymal cells which usually contain mitochondria and lysosomes. The subsyncytial zone overlies numerous circular and longitudinal muscle fibres. The absence of mitochondria and enzymes associated with active transport suggests that the amphistome tegument may be mainly specialized for protection of the worm against mechanical and chemical conditions prevailing in the rumen. Active uptake of nutrients is probably not a primary function.     

The in vitro culture and tegumental dynamics of the plerocercoid of Ligula intestinalis (Cestoda: Pseudophyllidea)

Hoole D. and Arme C. 1985. The in vitro culture and tegumental dynamics of the plerocercoid of Ligula intestinalis (Cestoda: Pseudophyllidea). International Journal for Parasitology14: 609–615. An in vitro system for maintaining Ligula has been developed and used in an autoradiographic study on the tegumental dynamics. Parasite tissue was cultured in teleost Ringer, Eagle's medium, M199 medium and modified Leibovitz' medium. Small membrane-bound bodies within the microthrix border increased in number in all media, and abnormal mitochondria were found in the distal tegument in teleost Ringer, M199 and Eagle's. Autophagy occurred after 24 h culture in teleost Ringer. Ultrastructural and physiological (¹⁴C-D-glucose uptake) evidence suggests that parasite tissue maintained in modified Leibovitz' medium remains viable for at least 24 h. The turnover rate of the surface components of L. intestinalis in this medium is approx. 12–24 h.     

Fasciola gigantica: Immunolocalization of 28.5 kDa antigen in the tegument of metacercaria and juvenile fluke

Specific monoclonal antibody (MoAb) to 28.5 kDa tegumental antigen (TA) was used to localize this antigen in the tissues of metacercariae, newly excysted juvenile (NEJ), 1, 3, 5, and 7-week-old juveniles of Fasciola gigantica by using indirect immunofluorescence, immunoperoxidase and immunogold techniques. Both indirect immunofluorescence and immunoperoxidase detections showed that this antigen was concentrated in the tegument particularly in its outer rim, tegumental cells and their processes as well as epithelial linings of the oral sucker. Unlike adult F. gigantica, it was not detected in spermatogenic cells in the testes, cells of Mehlis’gland, oocytes within the ovary, and ovum within the egg of parasites. At the ultrastructural level, the immunogold labeling showed deposit of gold particles specifically in G₂ tegumental granules and on the surface membrane. Thus, this 28.5 kDa antigen is expressed in the tegument and associated structures of juvenile parasites, and it could be a major component of the G₂ granules which are shown to fuse with the surface membrane and contribute material to replace the casted-off membrane. This process is the replenishment and turnover of the surface membrane to prevent the attachment of the host immune effector cells.     

Differentiation of the tegument and associated structures in Diphyllobothrium dendriticum Nitsch (1824) (Cestoda: Pseudophyllidea). An electron microscopical study

Grammeltvedt Anne-François 1973. Differentiation of the tegument and associated structures in Diphyllobothrium dendriticum Nitsch 1824 (Cestoda : Pseudophyllidea). An electron microscopical study. International Journal for Parasitology3: 321–327. The differentiation of the tegument and associated structures of the coracidium, procercoid, plerocercoid and adult is described. The embryophore is composed of four zones and is covered by a fibrous layer resembling a glycocalyx. The oncospheral plasma membrane is extensively folded. A typical cestode tegument, with a distal and perinuclear cytoplasm, is probably already existing in the coracidium. The formation of the microvilli starts after about three days in the copepod host. In young procercoids ribosomes and Golgi complexes were observed in the distal cytoplasm. These organelles disappear at later stages. The infective procercoid has a typical tegument. The microvilli are shaped like a thorn compressed from the sides. They have an electron dense tip and a less dense base in which microfilaments are seen. Bodies, called disc-shaped and lamellated bodies, are described. The microvilli of the plerocercoid are characterized by a great variation in shape. The villi are bounded by two unit membranes. The lamellated bodies are especially well developed. The adult microvilli are uniform in shape. The lamellated bodies are few in young adults and disappear in mature worms.     

The formation of the cyst wael of the metacercaria of Cloacitrema narrabeenensis (Bowell & Bearup, 1967) (digenea: Philophthalmidae)

Dixon K.E. and Colton M. 1978. The formation of the cyst wall of the metacercaria of Cloacitrema narrabeenensis (Howell &; Bearup, 1967) (Digenea: Philophthalmidae). International Journal for Parasitology8: 491–499. The cercaria of Cloacitrema narrabeenensis contains six different types of cystogenic cells which were distinguished on the basis of their position, ultrastructure and chemical composition. Four of the different types secrete carbohydrate-protein complexes and the other two protein granules. Early in development, the cercaria is bounded by a flattened, cellular envelope which contains a few mitochondria but lacks cytoplasmic ground substance and a nucleus. Later in development, this envelope is lost, shortly after a new cellular covering forms at the surface. The nuclei are later lost from this layer and it is suggested that they sink inwards, thus forming a cercarial tegument. The products of the cystogenic cells are gradually discharged through pseudopodial-like connections with the surface layer of the cercarial tegument to form the metacercarial cyst wall. The sequence of secretion is controlled so that the separate layers of the cyst wall are formed in their correct order.     

Induction cues for tegument formation during the transformation of Schistosoma mansoni cercariae

Adult schistosomes are parasitic blood flukes that have a continuous double lipid bilayered membrane surrounding the entire worm. This tegumental membrane is synthesised during invasion of the vertebrate host by free-swimming infectious forms called cercariae. As cercariae invade their final hosts they lose their tails and encounter a changing environment that includes altered temperature, sugar concentration and osmolarity. We have identified a glucose transporter protein designated SGTP4 that is found exclusively in the outer adult tegument and on membranous vesicles within the tegumental cytoplasm. By using immunofluorescence analysis to monitor the appearance and distribution of SGTP4 we can track the process of new tegumental membrane formation and examine the cues that trigger this developmental pathway. Cercariae in water do not transform their tegument while those incubated in rich medium do so rapidly. We have examined which of the many constituents of rich medium are responsible for triggering this transformation. Incubation in a solution of moderate osmolarity (120 mOsM PBS) is sufficient by itself to trigger tegument transformation, albeit at a slower rate relative to incubation in rich medium. Adding either glucose (to 100 mM) to the solution or increasing the temperature of incubation (from 22 degrees C to 37 degrees C) further increased the rate of tegument biogenesis. The introduction of glucose together with an increase in the incubation temperature further accelerated the process, suggesting that these factors act synergistically to promote transformation rates. The critical nature of osmolarity in inducing the process is highlighted by the fact that transformation proceeds as efficiently in 360 mOsM alone as it does in rich medium. While the fatty acids linolenic acid (cis-9, cis-12, cis-15-octadecatrienoic acid at 1 mM) and capric acid (Decanoic acid, at 0.1 mM) have both been proposed to stimulate tegumental transformation, we show that neither promotes the morphogenesis of a normal schistosomulum tegument. The schistosomicide praziquantel (to 1 mM) has no detectable effect on new tegument formation.     

Ultrastructural observations on the redial tegument of Paramphistomum epiclitum from the planorbid snail, Indoplanorbis exustus.

The morphology of the tegument in the redia of Paramphistomum epiclitum (Digenea: Paramphistomidae) resembles that shown by most larval and adult digeneans; an outer surface syncytium is in continuity with the cytoplasm of in-sunken, nucleated cytons. Although tegumental cytons usually contain a single nucleus, some display up to six nuclei. The tegumental syncytium lining the pharynx of P. epiclitum rediae lack underlying cytons. The apical membrane of the tegument is elaborated by folds and microvilli, which presumably facilitate uptake of nutrients and/or exchange of ions involved in osmoregulation. A single type of secretory body, resulting from the fusion of smaller vesicles produced at Golgi complexes in the cytons, occurs throughout the tegument. Uniciliate sensory receptors occur in the surface syncytium particularly around the oral opening.     

鱊头槽绦虫原尾蚴皮层的超微结构观察

利用透射电镜观察了头槽绦虫 (Bothriocephalusacheilognathi)原尾蚴皮层的超微结构。头槽绦虫原尾蚴的皮层为典型的合胞体结构。皮层表面有浓密的微毛与少量的突起。与成虫相比 ,原尾蚴的微毛长而且粗 ,呈现单态性 ,推测在进入终末宿主的过程中有脱落和再生现象。突起分布在头部与体侧 ,含有较少的电子致密颗粒。在原尾蚴皮层细胞质中观察到三种腺细胞的分泌过程 ,即顶分泌、外分泌和微分泌过程。原尾蚴身体后部观察到尾钩和穿刺腺管道的开口。在核周区和纤维层下可见发育较好的环肌与纵肌。本文还对原尾蚴皮层突起的功能进行了探讨。     

Schistosoma mekongi: the in vitro effect of praziquantel and artesunate on the adult fluke

The efficacy and tolerance of 80 microg/ml praziquantel (PZQ) and 40 microg/ml artesunate (ATS) against adult stage Schistosoma mekongi in vitro were investigated after 3, 6, 12, and 24h incubation by monitoring worm motility and compared tegumental changes using scanning electron microscopy (SEM). Thirty mice were infected with S. mekongi cercaria for 49 days. Adult worms were collected by perfusion method and prepared for in vitro study. Contraction and decreased motor activity were observed after as little as 3h incubation with PZQ and ATS. Some of the worms were immobile 12h after exposure, and died within 24h. The tegument of S. mekongi showed severe swelling, vacuolization and disruption, fusion of the tegumental ridges, collapse and peeling. After 12-24h incubation, PZQ induced similar but they less severe, tegumental changes to those observed after exposure to ATS. The direct observation of the fluke motility and SEM study suggest that ATS is more effective than PZQ in causing tegumental damage in adult S. mekongi, and provides a basis for subsequent clinical trials.     

Functions of the tegument of schistosomes: clues from the proteome and lipidome

The tegumental outer-surface of schistosomes is a unique double membrane structure that is of crucial importance for modulation of the host response and parasite survival. Although several tegumental proteins had been identified by classical biochemical approaches, knowledge on the entire molecular composition of the tegument was limited. The Schistosoma mansoni genome project, together with recently developed proteomic and lipidomic techniques, allowed studies on detailed characterisation of the proteins and lipids of the tegumental membranes. These studies identified tegumental proteins and lipids that confirm the function of the tegument in nutrient uptake and immune evasion. However, these studies also demonstrated that compared to the complete worm, the tegument is enriched in lipids that are absent in the host. The tegument is also enriched in proteins that share no sequence similarity to any sequence present in databases of species other than schistosomes. These results suggest that the unique tegumental structures comprise multiple unique components that are likely to fulfil yet unknown functions. The tegumental proteome and lipidome, therefore, imply that many unknown molecular mechanisms are employed by schistosomes to survive within their host.