Daughter Sporocysts Research Articles

Infection patterns in the first intermediate host of Dicrocoelium dendriticum

In order to establish the infection patterns in the first intermediate host of Dicrocoelium dendriticum, investigations were carried out on sheep pastures situated in the eastern part of Brandenburg, Germany, during the grazing periods in 1986, 1987, 1989 and 1990. It was found that the majority of specimens of the first intermediate host, Helicella obvia, become infected in the autumn of their second year of life, when their shell diameter is of medium size. The percentage of snails containing daughter sporocysts was highest in spring. Slimeball output could be provoked only in May and June.

Schistosoma mansoni: host cell adhesion to the different stages of the parasite, in vivo.

The peritoneal cavity of laboratory mice was used to study the phenomenon of host cell adhesion to different evolutive stages of the Schistosoma mansoni (cercaria, adult worm, developing and mature eggs, miracidium, young and mature daughter sporocysts). Material recovered from the peritoneal cavity 30 and 180 min after the inoculation of each evolutive form was examined with the help of a stereomicroscope. The free swimming larvae (cercaria and miracidium), and the evolutive forms producing such larvae (mature egg and mature daughter sporocyst) elicited the host cell adhesion phenomenon. In all forms but cercariae the adherent cells remained as so till 180 minutes after inoculation.

Isolation of mother and daughter sporocysts of Trichobilharzia ocellata from Lymnaea stagnalis.

Isolation of mother and daughter sporocysts of Trichobilharzia ocellata from Lymnaea stagnalis.

Lecithochirium furcolabiatum (Jones, 1933), Dawes 1947: the miracidium and mother sporocyst

Experimental infections of the marine topshell Gibbula umbilicalis with Lecithochirium furcolabiatum (Digenea: Hemiuroidea) have allowed the development of a model system which will enable further studies of the molluscan host response. The long-lived intertidal prosobranch host is easily maintained in the laboratory, and experimental infection rates of 98% were consistently achieved. The miracidium and mother sporocyst have been studied at both light and ultrastructural levels, providing the first account of the morphology of these stages in Hemiuridae. The ingested egg hatches within the host intestine, treatment with L-cysteine and alkaline pH stimulating miracidial emergence in vitro. The general body surface of the miracidium is devoid of spines or cilia, the latter being restricted to four plates near the anterior extremity. The miracidium swims actively prior to penetration of the gut wall, the sporocyst being released from the miracidial epidermal coat within the haemocoel. Within 5 weeks of infection, the filamentous mother sporocyst contains 1 to 3 oval germ balls, daughter sporocysts being recorded free within the digestive gland haemocoel 7 weeks later. Twenty three weeks after ingestion of eggs, the daughter sporocyst extends into the host gill filaments, containing cystophorous cercariae ready for emergence.

Interactions between the snail Lymnaea truncatula and the plagiorchiid trematode Haplometra cylindracea

Ultrastructural observations of Haplometra cylindracea sporocysts revealed that the so-called “paletot”, a peculiar structure covoring the daughter sporocysts of various Plagiorchiid trematodes, derives from modified snail host hemocytes. This study emphasizes an unusual interference of the parasite with the snail's internal defense system; free hemocytes, whether attracted or trapped, are modified by their contact with the parasite, which seems to use them as a covering and as a source of nutrients.

Schistosoma mansoni: Influence of infection on levels of translatable mRNA and on polypeptide synthesis in the ovotestis and albumen gland of Biomphalaria glabrata

Larval trematode infection causes a disruption of normal reproductive activity in the molluscan intermediate host. Because relatively little is known about the dynamics of this host-parasite interaction, the effect of Schistosoma mansoni infection on translatable mRNA pools and on polypeptide synthesis was examined in the ovotestis (OT) and albumen gland (AG) of Biomphalaria glabrata. Total RNA was isolated from OTs and AGs from uninfected control snails and snails at 14, 21, and 28 days postinfection (pi) with 20 S. mansoni miracidia and subjected to a rabbit reticulocyte in vitro translation system. Quantitative densitometry of autofluorograms of one-dimensional SDS-PAGE slab gels revealed reductions in quantities of total proteins synthesized in vitro from RNA isolated from infected OTs at 21 and 28 days pi, but not at Day 14 pi. Similar reductions were seen in 10 individual polypeptides selected for a more detailed analysis. In contrast to the OT, Day 14 pi-infected AGs exhibited an initial increase in total protein synthesized in the in vitro translation system utilized, followed by significant reductions at 21 and 28 days pi. Selective modulation of labeled polypeptides was evident in 11 polypeptides chosen for a more detailed analysis. This general pattern of parasite inhibitory effects was also seen in parallel pulse-chase studies using [ 35S]methionine metabolic labeling of in vitro-cultured OTs and AGs. In these experiments, significant reductions in the amounts of labeled polypeptides found in culture supernatants at 14, 21, and 28 days pi were evident. Total polypeptide synthesis also was reduced in solubilized AGs from infected snails at 21 and 28 days pi. Results indicate that larval trematode infection induced a generalized disruption of polypeptide metabolism in OTs and AGs of B. glabrata. Such inhibition may occur at both the transcriptional and the translational levels and is initially manifested early in infection, during the time that daughter sporocysts begin to migrate and colonize the digestive gland and OT.

Effects of Trichobilharzia ocellata on hemocytes of Lymnaea stagnalis

We analyzed the effects of infection with Trichobilharzia ocellata on hemocytes of its snail host, Lymnaea stagnalis, and correlated them with successive stages of parasite development. Circulating hemocytes were studied at 0, 2, 4, 6, and 8 weeks post exposure (p.e.) with respect to cell number, distribution of subpopulations (as characterized by morphology, determinants recognized by either of two lectins and a monoclonal antibody) and to proliferative, phagocytic and endogenous peroxidase activity. Infection results in a net elevated level of activity of circulating hemocytes at 2 weeks p.e., when mother sporocysts are present in the head-foot-mantle region, as well as at 4 weeks p.e., when daughter sporocysts are migrating to and growing in the digestive gland region. A lower level of activity was observed at 6 weeks p.e., when cercariae are differentiating within daughter sporocysts. A net activation was again found at 8 weeks p.e., when cercariae are escaping. So, infection with T. ocellata results in a net general activation of the internal defense system of L. stagnalis, during several stages of development of the parasite.

Trichobilharzia ocellata: influence of infection on the fecundity of its intermediate snail host Lymnaea stagnalis and cercarial induction of the release of schistosomin, a snail neuropeptide antagonizing female gonadotropic hormones

Subadult and adult specimens of the pond snail Lymnaea stagnalis were infected with the schistosome Trichobilharzia ocellata. Egg production and growth of the snails were monitored over an 8-week period post-infection (p.i.). Snail haemolymph was collected and analysed for the presence of schistosomin, a neuropeptide which antagonizes the action of the snails' female gonadotropic hormones. Snails infected as subadults showed an increase in fecundity during the first 4 weeks p.i. compared with non-infected controls. The possibility is discussed that this increase is caused by an accelerated maturation of the female sex organs due to elevated levels of Dorsal Body Hormone, a female gonadotropic hormone. No difference in fecundity was found between snails infected as adults and control snails during the first 4 weeks p.i. Snails infected as subadults and as adults showed a decrease in fecundity from week 5 p.i. and onwards. This decrease coincided with the appearance of schistosomin in the haemolymph of the snails and with that of differentiating cercariae in the daughter sporocysts. Cercariae are probably involved in the induction of schistosomin release from the snails' CNS into the haemolymph. Snails infected as subadults or as adults grew at approximately the same rate as uninfected snails.

Trichobilharzia ocellata: Quantification of effects on haemocytes of the pond snail Lymnaea stagnalis by morphometric means

To identify functionally different subpopulations, we quantified by morphometric means the spreading activity of circulating haemocytes of the pond snail Lymnaea stagnalis, recognized by monoclonal antibodies (5 surface and 5 cytoplasmic). The influence of snail age and of the different intramolluscan stages of the compatible avian schistosome Trichobilharzia ocellata on this activity were studied. The antibody-recognized cells could be separated into two groups, differing in their spreading activities. The probes detecting cytoplasmic markers recognized the majority of cells (78–95%). These were active (well spreading), differentiated cells. The surface probes recognized a smaller part (12–38%) of the total haemocyte population. These harmocytes were less active (less spreading), and were predominantly immature cells. The relative sizes of the antibody-detected subpopulations were not affected by snail age or infection with T. ocellata. The maximal size (planimetric area) attained after attaching to glass of all cells increased between day 0 (juvenile snails) and about 6 weeks post (sham) exposure and then decreased. Infection had little effect on the spreading activity of the more differentiated cells. The less differentiated cells showed a larger spreading activity when the parasite was present as mother sporocyst and also when the digestive gland area became colonized by growing daughter sporocysts.

Influence of larval schistosomes on polysaccharide synthesis in albumin glands of Biomphalaria glabrata

An in vitro bioassay was used to examine [14C]glucose incorporation into polysaccharides in albumen glands (AGs) of susceptible M-line Biomphalaria glabrata infected with the NMRI strain of Schistosoma mansoni. Polysaccharide and galactogen synthesis were unaffected by larval trematode infection in AGs of snails at days 14, 21, and 28 post-infection (p.i.) when compared to uninfected controls. Further experiments were conducted to determine if daughter sporocysts, hypothesized to be primary mediators of parasitic castration in this system, were able to exert direct effects on synthetic activity of uninfected AGs via haemolymph-borne molecules or in vitro culture-generated larval excretory-secretory (ES) products. When AGs were incubated in the presence of infected snail haemolymph, significant differences in quantities of polysaccharides and galactogen were detected only in test organs incubated in day 28 p.i. haemolymph. Daughter sporocyst ES products generated during the first 48 h of culture caused a significant reduction in polysaccharide and galactogen synthesis in test organs. When ES products from days 3 to 6 of in vitro culture were tested similarly, no significant differences in either polysaccharide or galactogen synthesis were observed between control and test organs. These data demonstrate that daughter sporocysts are able to modulate a specific aspect of the reproductive activity of the snail host through haemolymph-borne molecules of host or parasite origin, or directly through in vitro culture-generated ES products.

Praziquantel treatment of Biomphalaria glabrata infected with Schistosoma mansoni– influence on snail fecundity

Praziquantel, administered over a 72 h period in the food of mature Biomphalaria glabrata harbouring 7-week-old Schistosoma mansoni infections, dramatically reduced the numbers of cercariae shed. Doses of 20-30 micrograms/g body weight (including shell weight) reduced shedding by 85-95% over 5 weeks before recovery was evident. Suppression of cercarial shedding in these infections was accompanied by the temporary recovery of the snail reproductive regression due to S. mansoni infection. Snail fecundity was subsequently re-suppressed 2 weeks prior to recovery of the parasite as evidenced by a resumption of cercarial shedding. Praziquantel destroyed mature and developing cercariae within the daughter sporocysts but had no apparent effect on daughter sporocysts; this may account for the eventual resumption of cercarial production. Reproductive failure of the snail is apparently related to the latter stages of cercarial development specifically. Reproductive recovery did not occur when snails were infected as juveniles or when mature-infected snails harboured 12.5 week i.e. older infections: drug treatment temporarily inhibited cercarial production but no snails produced eggs.

Lectin binding to cells of Schistosoma mansoni sporocysts and surrounding Biomphalaria glabrata tissue

The binding of different lectins to the surface of mother and daughter sporocysts of Schistosoma mansoni (Trematoda) and to cells of its intermediate host Biomphalaria glabrata (Gastropoda) was investigated. The test system consisted of several biotin-labeled lectins, an avidinbiotin-peroxidase complex and 3,3′-diaminobenzidine. The fixatives used were Formalin, Bouin's and Zenker's solutions; unfixed material was also studied. Most lectins reacted equally with host tissue and parasite tissue. However, receptors for Ulex europaeus I (most probably fucose) were only demonstrated on daughter sporocysts. Thus, a method was found to specifically mark Schistosoma mansoni daughter sporocysts in the digestive gland tissue of its intermediate host. Mother sporocysts and surrounding host tissue differed in their distribution of galactosyl groups. Both lack fucose and N-acetyl-galactosamine. The differences in lectin binding of galactosyl determinants were also observed during the in vitro development of mother to daughter sporocysts.

Histofluorescent and ultrastructural demonstration of biogenic amines in daughter sporocyst ofDiplostomum pseudospathaceumNiewiadomska, 1984 (Digenea)

The aminergic neurons in D. pseudospathaceum daughter sporocysts were studied by fluorescence histochemistry and electron microscopy. Green fluorescence showing the presence of catecholamines was present in the central and peripheral nervous systems. Ultrastructurally different types of vesicles were identified on the basis of their size and morphology. Their possible function was discussed.

Ultrastructural Evidence for the Destruction of Schistosoma mansoni Sporocysts Associated with Elevated Lysosomal Enzyme Levels in Biomphalaria glabrata

The activity levels of serum acid phosphtase, aminopeptidase, and lysozyme in a Brazilian strain of Biomphalaria glabrata were ascertained at 1, 2, and 3 hr after mechanical wounding or injection with albumin on the 30th day postexposure to a compatible strain of Schistosoma mansoni miracidia and found to be elevated. Parallel transmission electron microscope studies on daughter sporocysts and developing cercariae at these time intervals revealed progressive disintegration of the parasites that was associated with increased numbers of host granulocytes abutting the sporocyst surfaces. Furthermore, host granulocytes were observed to have passed through eroded sporocyst walls and attacked developing cercarial embryos. It is proposed that the elevated levels of lysosomal hydrolases released from activated host granulocytes as a result of challenge altered the parasite's surfaces so that these were recognized as nonself. Consequently, additional host granulocytic response, which included additional release of lysosomal enzymes into serum as well as phagocytosis of remnants of both sporocysts and developing cercariae, was elicited.

Ultrastructural studies on the daughter sporocysts of Meiogymnophallus minutus (Digenea: Gymnophallidae)

The daughter sporocyst body wall is a syncytial microvillous tegument resting on a basement lamina, followed by circular and longitudinal muscle fibres and five kinds of subtegumentary cells, namely flame, myoblast, germinal, secretory and digestive cells. The last two have cytoplasmic bridges connecting them to the tegument and their principal function is probably nutrition. Secretory cells probably synthesize hydrolytic enzymes which pass via the tegument into the host's haemolymph for exogenous digestion of nutrients. Simple nutrients are probably absorbed over the entire surface of the microvilli whereas undigested or partly digested nutrients are absorbed by micropinocytosis at their base. Some nutrients may be digested in the tegument but most pass into the digestive cells which undergo a cycle including synthetic, absorptive, digestive, storage, autophagic and resting phases. Nutrients are supplied to developing embryos by extensions of digestive cells which line the brood chamber. Residual bodies are shed into the host's haemolymph, or into the brood chamber. In ageing sporocysts, microvilli and later the tegument completely degenerate and subtegumental cells undergo extensive autolysis, finally, the body wall ruptures, releasing the remaining fully formed cercariae.

Schistosoma rodhaini: dynamics and cercarial production for mono- and pluri-miracidial infections of Biomphalaria glabrata.

No significant difference in cercarial production was noted when Biomphalaria glabrata from Brazil were exposed to one or several miracidia of Schistosoma rodhaini from Burundi, even though the dynamics seemed different. The minor differences can be explained by the intramolluscal larval development of this parasite characterized by the production of a high number of daughter sporocysts over a long period.

Schistosoma mansoni: Effect of infection on reproduction and gonadal growth in Biomphalaria glabrata

Sexually mature Biomphalaria glabrata were exposed to 12 miracidia of Schistosoma mansoni, and egg production of snails was monitored over a period of 5 weeks. During the study period, exposed snails grew at approximately the same rate as unexposed controls. Castration, as measured by a reduction in the mean number of eggs laid per snail, occurred between 14 and 21 days postexposure (PE). The reduction in fecundity in infected snails coincided with the migration and establishment of daughter sporocysts in the digestive gland and gonad. Enumeration of individual oocytes in longitudinal sections of the ovotestis revealed that uninfected snails contained significantly more oocytes per section than infected snails at 27, 31, and 40 days PE. In addition, the mean area of gonadal sections of control snails increased over the 40-day experimental period, whereas there was no such increase in gonadal area of infected snails. These data suggest that there is an inhibition in gonadal growth in infected snails. When oocyte data were expressed in terms of mean gonadal area, the mean number of oocytes per mm 2 of gonad of uninfected and infected snails did not differ significantly over the study period, except at Day 14 PE, when infected snails contained a significantly greater number of oocytes per mm 2 of gonad than did uninfected controls. It is hypothesized that daughter sporocysts of S. mansoni are primarily responsible for the inhibition of host reproductive activity, and may be mediating their effects through mechanisms involved in the regulation of gonadal growth. However, it is not known whether such action is mediated directly on the gonads or indirectly through alterations in snail host metabolism.

Dynamics of the intramolluscan larval development of Schistosoma haematobium: replication of daughter sporocysts and cercarial production.

During the intramolluscan larval development of Schistosoma haematobium (Algerian strain) in Bulinus truncatus, two replication processes of daughter sporocysts occur. Replication by direct sporocystogenesis appears more important than sporocystogenesis post cercariogenesis. These mechanisms assure a periodic renewal of the sporocyst stock in the snail host and seem to be synchronized with the development of cercarial generations. The succession of several generations of cercariae is responsible for the alternation of high and low periods of productivity. The scheme proposed for the intramolluscan development of S. haemtobium is compared with those described for S. mansoni and S. bovis and interpreted in terms of demographic strategies adapted to a better exploitation of the snail host.

Schistosoma rodhaini: intramolluscan larval development, migration and replication processes of daughter sporocysts

The larval development of Schistosoma rodhaini in Biomphalaria glabrata presents two features of interest. The first concerns the migration of daughter sporocysts: only a fraction of the daughter sporocysts produced by the mother sporocyst migrate directly towards the digestive gland of the snail and become cercariogenous; the others remain in the anterior region of the snail where the majority transforms into small sporocystogenous sporocysts. The second point concerns the replication processes: only direct replication of daughter sporocysts was observed in sporocysts in a prehepatic localization. The demographic processes involved in the development of S. rodhaini are compared with other species of Schistosoma and interpreted as an adaptive character for better exploitation of the snail host.

Schistosoma mansoni: Origin and expression of a tegumental surface antigen on the miracidium and primary sporocyst

A monoclonal antibody, recognizing a carbohydrate epitope associated with several tegumental surface components on Schistosoma mansoni primary sporocysts, was used to follow tegumental formation during transformation of the miracidium to sporocyst and its subsequent development in vitro and in vivo. Indirect fluorescent antibody and direct immunogold labeling methods confirm a structural connection between the intercellular ridges and a submuscular, multinucleate syncytium in the miracidium. Immunoreactive vesicles within this latter system directly contribute to elaboration of the tegumental surface membrane, through the process of membrane fusion. Lateral expansion of intercellular ridges by vesicular fusion ultimately result in fully transformed sporocysts exhibiting vesicular membrane epitopes as prominent tegumental surface components. Light microscopical and ultrastructural observations, together with Western immunoblot analyses, suggest a gradual depletion of intracellular and surface immunoreactive material of vesicular origin in primary sporocysts grown in culture for up to 12 days. In contrast, similar immunoreactive vesicles appear to be continuously synthesized throughout in vivo primary sporocyst development. Monoclonal antibody reactive epitopes appear to be uniquely expressed in the miracidium/primary sporocyst since similar molecules are absent from daughter sporocysts, cercariae, adults, and snail tissues.