Two New Proschizorhynchus Species from the Coast of Massachusetts, USA (Turbellaria, Kalyptorhynchia)1

Proschizorhynchus nahantensis sp. n. and Proschizorhynchus papillatus sp. n. are described and their taxonomy discussed. P. nahantensis is characterized by its penis papilla which has a proximal spherical and distal tubular portion. A simple stylet is present. The female system is distinguished by the external opening of the vagina and the prominent bursal mouthpiece. P. papillatus is distinguished by the two muscle layers of the copulatory organ, the large straight stylet in the penis papilla, a single gonopore and uterus. Notes on the ecology of the species are included. Views are given on the taxonomy of the genus Proschizorhynchus Meixner, 1928.     

Redescription of Syndesmis echinorum Fran?ois, 1886 (Turbellaria: Neorhabdocoela: Umagillidae), with comments on distinctions between Syndesmis and Syndisyrinx

Syndesmis echinorum is redescribed from specimens taken from the intestine of the sea urchin Paracentrotus lividus at the type locality, Banyuls-sur-Mer, France. This umagillid is shown to have a seminal bursa-bursal valve complex comparable to that of Syndisyrinx franciscanus. Thus, it is suggested that the genera Syndesmis and Syndisyrinx be distinguished on the basis of the form and proportions of the ejaculatory duct, male antrum, and penis stylet, rather than on the basis of the presence or absence of a seminal bursa and bursal valve. The sea urchin Sphaerechinus granularis at Banyuls harbors a worm that is, on the average, slightly more robust than S. echinorum from P. lividus; its morphology, however, agrees with that of S. echinorum. In addition to S. echinorum, P. lividus harbors at least 1 undescribed umagillid. Although 2 species found in Echinus esculentus have been referred to S. echinorum by previous workers, neither is conspecific with it.     

豚鼠卵巢囊淋巴孔的发现及卵巢囊超微结构研究

In order to explore its structure and speciality of species, the ovary bursa of guinea pig was studied by using dissecting microscopy and scanning electron microscopy. In this paper, the lymphatic stomata on both internal and external layer of the ovary bursa was frist reported in guinea pig. The results suggested that the stomata in bursa not only provided a pathway to connect the bursal cavity with the lymphatic vessels of bursa and the peritoneal cavity, but also may be involved in the reproduction and local immunity of the ovary. The stomata may play an important role in physiological function of the ovary. At the same time, the structural differences were identified in ovary bursa between guinea pig and golden hamster.     

A molecular analysis of the phylogenetic position of the suborder Cavernicola within the Tricladida (Platyhelminthes), with the description of a new species of stygobiont flatworm from Benin

This paper presents a new species of cavernicolan planarian from the African continent, which enabled us to determine the position of the suborder Cavernicola within the order Tricladida on a molecular basis. A recent paper suggested a sister‐group relationship between Cavernicola and marine triclads. However, our work is the first molecular study to demonstrate unequivocally that the Cavernicola does not group with either the freshwater planarians or the land planarians, i.e. is not closely related to the suborder Continenticola, and corroborates the suggested sister‐group relationship with the marine triclads or Maricola. Two possible scenarios are proposed for the evolution of the Cavernicola from marine ancestors. The new species is assigned to the genus Novomitchellia and is characterized by the absence of eyes. Testicular follicles are numerous and only dorsal, extending from behind the ovaries to the posterior end of the body. The vasa deferentia separately penetrate the penis bulb and the seminal vesicle. The penis papilla is very short and blunt, and the penis bulb musculature is weak. A long and narrow copulatory bursa is situated behind the gonopore. The common oviduct is orientated perpendicularly to the short bursal canal.     

Effect of environmental antigens on the Ig diversification and the selection of productive V-J joints in the bursa

In chickens, a single set of unique functional segments of both Ig H and L chain genes is rearranged during early embryogenesis to generate a pool of B cell progenitors that will be diversified in the bursa by gene conversion, forming the preimmune repertoire. After hatching, bursal cells are exposed to environmental Ags in the bursal lumen. We prepared B cells from each single bursal follicle and used PCR-directed Ig L chain gene analysis to study the differentiation of B cells and the effect of antigenic stimulation from the bursal lumen on the neonatal chicken B cell repertoire formation. Selective amplification of B cell clones with a productive V-J joint was observed during the late embryonic stage, possibly by the interaction with ligands expressed on the bursal stroma and further accelerated in the neonatal chicken. Administration of the artificial Ags into the bursal lumen before the isolation of bursa by bursal duct ligation in the embryo caused a significant increase in lymphocytes with a productive V-J joint in the neonatal chicken bursa compared with the isolated bursa. Intra- and interclonal diversity of a complementarity-determining region measured by an evolutionary distance increased during bursal development. Clonal diversification did not require stimulation by artificial Ags from the bursal lumen. Thus, the preimmune repertoire in the bursa is generated by gene conversion during Ag-independent B cell proliferation, and antigenic stimulation from the bursal epithelium to bursal B cells plays roles in the selection of clones with a productive V-J joint.     

Functional association between female sperm storage organs and male sperm removal organs in calopterygid damselflies

Female damselflies in the family Calopterygidae have two sperm storage organs: a spherical bursa copulatrix and a tubular spermatheca. Male flies have a peculiar aedeagus with a recurved head with which to remove bursal sperm, and lateral spiny processes to remove spermathecal sperm. The lateral processes differ among species and populations in terms of their width relative to the spermathecal duct: the narrower processes are physically able to access spermathecal sperm, while the wider ones are not. In the present study, sperm storage patterns and aedeagal structures were compared between two calopterygid species with different spermathecal structures –Calopteryx cornelia and Mnais pruinosa– with respect to not only sperm quantity (number) but also sperm quality (viability), by using a recently developed method based on live/dead dual fluorescence. Calopteryx cornelia is a typical spermathecal sperm remover. In this species, viability was similar between bursal and spermathecal sperm. In contrast, in M. pruinosa, the spermatheca was much smaller than the bursa and often contained no sperm. Even when the spermatheca of this species did contain sperm, a high percentage of it was dead. Although the spermatheca of M. pruinosa has such atrophic tendencies, males have nevertheless developed long and spiny lateral processes similar to those of C. cornelia, suggesting the processes have functions other than spermathecal sperm removal. They possibly function as stoppers or guides for manipulating the aedeagal head to remove the sperm mass from the bursa.     

A novel method to bursectomize avian embryos and obtain quail----chick bursal chimeras. II. Immune response of bursectomized chicks and chimeras and post-natal rejection of the grafted quail bursas

Two methods to bursectomize chick embryos before hemopoietic cell seeding of the bursa of Fabricius were compared in this work: section of the tail region at E3 including the presumptive bursal territory, and selective removal of the bursa at E5. Hatching ability is better with the former method, but survival rate and effectiveness of bursectomy are favored with the second, novel technique. Moreover, selective removal of the bursa at E5 can be followed by in situ engraftment of a quail bursa and construction of quail-chick bursal chimeras. The immune response of bursaless birds and bursal chimeras has been studied. Total absence of the bursa does not prevent a few B cells from differentiating and nonspecific Ig (IgM and/or IgG) from being secreted. As reported previously, bursaless birds, however, are unable to mount an immune response by producing specific antibodies. This immune function is restored by the graft of a quail bursa. The microenvironment of the bursa, although heterospecific, allows the expansion of the B cell population and generates the repertoire of the B cell antigen receptors. This process takes place during late embryonic and early postnatal life because the grafted quail bursal stroma is subjected to immune rejection from 2 to 3 wk after birth in all chimeras, which are, however, perfectly immunocompetent.     

豚鼠卵巢囊淋巴孔的发现及卵巢囊超微结构研究

本实验通过扫描电镜等方法研究豚鼠卵巢囊及卵巢囊淋巴孔的结构,并探讨了卵巢囊及其淋巴孔的种属差异.实验结果首次报道豚鼠卵巢囊内、外层上皮均存在淋巴孔;豚鼠卵巢囊结构在输卵管走行、囊闭合程度及囊表面超微结构等方面与金仓鼠存在差异.结果提示豚鼠卵巢囊内、外层淋巴孔是沟通卵巢囊腔、卵巢囊淋巴系及腹膜腔的形态基础,可能是三者间物质转运的重要途径.卵巢囊淋巴孔可能影响物种的繁殖并参与囊腔内局部免疫反应.卵巢囊结构和发育程度与对应的输卵管伞端等结构及其他生殖特点相适应,研究结果丰富了生殖形态学和比较解剖学资料.     

Bursa,bursa canal,and female antrum of Dugesia tigrina (Plathelminthes,Tricladida)

Summary The bursa of Dugesia tigrina is located between pharynx and penis and is suspended in a network of muscle fibers. Two cell types are present in the bursa: small outer cells which likely represent replacement cells and tall inner cells. During copulation seminal material, consisting of secretion products, fibrillar and tubular material, and small clusters of spermatozoa, is injected through the bursa canal into the bursa. During the 48 h post-copulatory period the seminal material is absorbed by the inner cells. Phagocytosis of the seminal material is facilitated by broad apical cytoplasm and modified distal cell membranes. Phagocytosed spermatozoa are enclosed in vaculoes and morphologic breakdown occurs as early as 4 h after copulation. Fibrillar and tubular material is phagocytosed directly into the apical, organelle-free cytoplasm without vacuole formation.The bursa canal is ciliated and the distal cell ends are studded with ultrarhabdites. These are also present in the epithelial cells of the female antrum. Antrum epithelial cells furthermore display apical organelle-free cell areas. Numerous glands penetrate through the antrum wall.     

Comparative ultrastructure of the terminal portions of the male copulatory apparatus in Planorbidae (Gastropoda: Pulmonata)

Terminal portions of the male copulatory apparatus of Planorbis planorbis, Segmentina oelandica, and Anisus vortex were studied using whole-mount preparations, serial semi-thin sections, and transmission electron microscopy. In the latter species, stylet formation was investigated at several stages of postembryonic development. Organization of the penial distal portion in the species studied varies greatly. In P. planorbis, the distal end of the penis lacks developed papillae and is armed with a stylet built up of the covering epithelial cells of the penis proper. In A. vortex, the stylet is formed by the secretory activity of the middle cells of the distal portion of the penis. To the time of maturation, the cells encompassing the stylet are broken down exposing its solid chitinous structure and characteristic shape. In S. oelandica, the distal end of the penis bears the long probably flexible papilla with the characteristics of an internal ‘skeleton,’ organized as a line of connective tissue cells and a system of hydrocoelic cavities.     

Ultrastructural investigations on the differentiation of genital hard structures in free-living platyhelminths and their phylogenetic significance

Ultrastructure and differentiation of penis stylets and stylet needles have been investigated in representatives of various groups of free-living platyhelminths, viz. the Acoela, Macrostomida, Typhloplanoida, Kalyptorhynchia, and Dalyellioida. In all these groups, the differentiation of such hard parts occurs intracellularly but in different ways in the different groups. The ultrastructure of the bursal mouth piece in an acoel platyhelminth is not comparable to the hard structures in male copulatory organs. The presence of penial copulatory organs having intracellular hard structures appears to be an autapomorphy of the Euplatyhelminthes. Several characters in the ultrastructure and development of these structures can be used as autapomorphies for various platyhelminth groups.     

Ultrastruktur und Differenzierung des Penisstiletts von Petaliella spiracauda (Plathelminthes, Rhabdocoela)

Petaliellu spiracauda Ehlers, 1974, has a complex funncl-shaped penis stylet, differentiating intracellularly in a synchronous mode. First, a stylet-forming cell is built which in shape is adapted to the future stylet; this cell is lined by interstitial cells and secretory cells proximally and enveloped by cover cells at the outside. The differentiation of the hard structurc starts at the same time in all parts. Electron dcnsc material is added to the inner surface of thc cell membrane of the styletforming cell, in which the number and size of layers differ in places. During the phase of formation, small tubules are arranged parallel on the inner surface of the hard structure. In the stylet base, however, many separate hard elements, joined by electron dense material, are formed at the membrane of the stylet-forming cell, as well as freely in the cytoplasm. All parts of the stylet are completed at the same time due to a varying growth in wall thickness. The base of the hard structure is embedded in a finely fibrous intracellular matrix.     

Studies of testosterone-induced involution of the bursa of Fabricius

The present investigation deals with the effect of testosterone on each of the tissue components of the bursa of Fabricius: the endodermal epithelium, the mesenchyme, and the hemopoietic stem cells. Tissue combination experiments between testosterone-treated endoderm and normal mesenchyme and vice versa have shown that the androgen damages irreversibly the bursal epithelium. The latter is not seeded by hemopoietic stem cells and cannot undergo follicle formation when treated with high doses of testosterone. This occurs even if it is associated with a nontreated bursal mesenchyme. On the contrary, associations of testosterone-treated mesenchyme with normal endoderm result in normal bursa histogenesis. By using an original test of viability for lymphoid cells based on the application of the quail-chick marker system, we demonstrate that disappearance of hemopoietic cells in the endoderm results from their expulsion from the bursa and not from their death in situ. The conspicuous effect of testosterone on the bursa of Fabricius can be related to the levels of androgen receptors found in the organ. Typical cytosol androgen receptors are demonstrated in both bursal endoderm and mesoderm, although the amount in the former is higher. The concentration of binding sites in the bursa is >10 times higher than that in other organs such as lung and small intestine whose development is not affected by testosterone, contrasting with glucocorticosteroid receptor (measured by labeling with dexamethasone) found in the same concentration in all tissues.     

Sclerotised spines in the female bursa associated with male’s spermatophore production in cantharidin-producing false blister beetles

Cantharidin is a defence chemical synthesised in only two beetle families Meloidae and Oedemeridae. In Meloidae, cantharidin is used as a defence chemical in eggs. However, in Oedemeridae the function of cantharidin remains unclear. Based on morphological comparison of female internal reproductive organs in 39 species of Oedemeridae, we found that some species have sclerotised spines in the bursa copulatrix (bursal spines), while others have no such spines. Molecular phylogenetic trees inferred from mitochondrial 16S and nuclear 28S rRNA gene sequences suggested multiple evolutionary origins of bursal spines from an ancestor without spines. In the species which lacked spines, males transferred small amounts of ejaculates to females; however, in species with spines, males transferred large spermatophores. Deposited spermatophores gradually disappeared in the bursa, probably owing to absorption. To compare the amounts of cantharidin in eggs laid by species with and without bursal spines, we constructed a new bioassay system using the small beetle Mecynotarsus tenuipes from the family Anthicidae. M. tenuipes individuals were attracted to droplets of cantharidin/acetone solution, and the level of attraction increased with cantharidin concentration. This bioassay demonstrated that the eggs of Nacerdes caudata and N. katoi, both of which species have conspicuous bursal spines, contain more cantharidin than the eggs of N. waterhousei, which lacks spines. In the former species, males transfer large spermatophores to the female, and spermatophores are eventually broken down and digested within the female’s spiny bursa. Thus, females with bursal spines may be able to provide more cantharidin to their eggs.     

Postnatal development of the ovarian bursa of the golden hamster (Mesocricetus auratus): its complete closure and morphogenesis of lymphatic stomata

The golden hamster ovarian bursa was studied by light and electron microscopy to clarify the process of its complete closure and the development of lymphatics that leads to morphogenesis of stomata. The results were as follows. 1) The bursa completely closed at 9 days of age primarily due to development of the mesotubarium superius. 2) With the closure, the ovary and bursa became closely apposed, and most of the original bursal cavity disappeared. 3) Between 9 and 12 days of age U-shaped folds of the bursal mesothelium began to invade the connective tissue of the bursa. 4) Widening of the internal angle of the U-shaped folds contributed to reappearance of the bursal cavity, and thus separation of the bursa from the ovary. It also contributed to future geometrical proximity of lymphatics to the cavity of the bursa. 5) The separation of the bursa from the ovary began as early as 12 days of age in the cephalic half of the bursa. It occurred remarkably late in the caudal half. Juxtaposition of the window portion of the bursa to the ovary remained in some adult animals. 6) Development of lymphatics in the cephalic half of the bursa was divided into two stages, before and after days 21-24 of life. In the first stage, lymphatics grew in the submesothelial connective tissue, and the framework of lymphatics was formed. In the second stage, lymphatics extended small branches to form the submesothelial plexus or lymphatic lacuna. 7) Intercellular junctions between contiguous lymphatic endothelial cells were mostly tight and desmosomelike. Open junctions were, if they occurred at all, rare. (8) A smooth-surfaced area lined with the lymphatic endothelium was found in the bursa on day 27 of life, before the initiation of ovulation. Valvelike stomal orifices were absent before the initiation of ovulation and extremely rare even after the first ovulation. They were commonly present in the bursae after the fourth ovulation, however. The process of complete closure of the ovarian bursa is very complex and may be related to the later development of the bursal mesothelium and lymphatics. Some liplike stomal orifices are of purely developmental origin. However, all valvelike stomal orifices are assumed to be formed as a result of damage to the bursal mesothelium, as well as to the submesothelial connective tissue and lymphatics, by repetition of ovulation. It is possible that liplike stomal orifices may be formed in the process of repairing the damage.     

Rhopalobdella japonica n. gen., n. sp. (Hirudinea, Piscicolidae) from Dasyatis akajei (Chondrichthyes: Dasyatididae) in the northwestern Pacific

A new genus and species of piscicolid leech in the Platybdellinae inhabits the oral cavity of Dasyatis akajei in the northwestern Pacific Ocean near Tanabe, Japan. The genus Rhopalobdella n. gen. is characterized externally by very small oral and caudal suckers and a smooth body that is widest just posterior to the clitellum. Eyespots and ocelli are lacking. The coelom is spacious with large segmental connecting sinuses between dorsal and ventral sinuses. There are 5 pairs of testisacs, an unusually extensive epididymis, and a very large bursa. Conducting tissue is absent. There are 2 pairs of esophageal diverticula and very well developed nephridia. Rhopalobdella japonica n. gen. n. sp. is characterized by a urosome that tapers strongly to the caudal sucker and by a single gonopore; the common oviduct opens into the posterior portion of the bursa. The coelomic and excretory systems resemble Aestabdella, but in other respects the genera are quite different. This is the first marine leech reported from rays in the northwestern Pacific.     

NGF Retards apoptosis in chick embryo bursal cell in vitro

Abstract. Recent studies have demonstrated that the action of nerve growth factor (NGF) is not restricted to neuronal cells but also affects cells of the immune system. In a previous work on the effect of NGF on the chick embryo bursa of Fabricius both in vivo and in vitro, we observed that NGF prolongs bursal cell survival in vitro. In the present study we report that the increase of viable cells in NGF-treated cultures is not due to a proliferative effect of NGF on bursal cells but to a reduction of cell mortality. The morphological analysis revealed that bursal cells in cultures die by apoptosis, which was also shown by the typical pattern of DNA fragmentation, a hallmark of this cell death process. It is concluded that NGF, with an action similar to that described in sympathetic neurons and PC12, could retard bursal cell death by influencing apoptosis.     

Immune complexes of E. coli antigens and maternal IgG in the bursa of Fabricius

The bursa of Fabricius of the chicken is known to be both a primary lymphoid organ and a secondary lymphoid tissue. Bursal follicles are equipped with antigen-trapping follicle-associated epithelium. However, bioactive antigens such as protein and bacteria have not been detected in the bursal parenchyma. By immunoperoxidase staining with a polyspecific antibody (Ab) against Escherichia coli, we detected aggregated E. coli antigens in the medulla of bursal follicles after hatching. The distribution of aggregated E. coli antigens is restricted to the medulla of bursal follicles. The antigens are not found in the spleen or the parenchyma of the caecal tonsil. The bursa is thus a trapping site for E. coli antigens from the external environment. Furthermore, two-color immunostaining clarified that these antigens form immune complexes with maternal IgG (MIgG) and are retained by reticular cells. Additionally, immune complexes in the bursa were shown to induce the rapid development of serum IgM Ab for indigenous E. coli. Our results suggest that immune complexes of MIgG and environmental antigens in the medulla of bursal follicles exert positive effects on B-cell differentiation in the bursa in situ.     

Evidence for widespread sperm displacement ability among Zygoptera (Odonata) and the means for predicting its presence

Males of the coenagrionid damselflies Argia moesta, A. sedula and hchnura ramburii use similar penis morphology to remove and/or reposition sperm of previous males from the storage organs of females prior to inseminating them. Although the species vary in the degree to which sperm is removed from or packed into the spermatheca, in all three species, sperm is removed from the bursa copulatrix. Since sperm in the bursa probably has priority in fertilizing eggs in at least the first oviposition after mating, sperm precedence can be estimated as the percentage of sperm (by volume) in the bursa belonging to the last male to mate. Estimated sperm precedence for these species is approximately 71% for Argia sedula, 82% for I. ramburii and 93% for A. moesta. These results, combined with similar ones for other damselflies clearly indicate that the ability to displace sperm may be widespread among temperate-zone Zygoptera. Species with each of the four major variations in damselfly penis structure have now been shown to displace sperm using this morphology. The systematic distribution of these major variants suggests several origins of sperm displacement ability within the Zygoptera. Whether or not all damselflies are capable of sperm displacement depends on both the presence of micro-structures used in sperm removal or repositioning and on the presence of sperm of previous males in mating females. It is possible, therefore, to predict that sperm displacement occurs in a damselfly if (1) females mate more than once, (2) mating females store sperm in organs accessible to penis morphology, (3) the distal segment of the male penis has structures similar to those known to be involved in sperm removal or repositioning, and (4) oviposition occurs in tandem or with the male non-contact guarding his mate.