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.

The in vitro transformation of the miracidium to the mother sporocyst of Schistosoma margrebowiei was initiated by placing the miracidium in mammalian physiological saline. The transformation occurs in stages: the cilia cease beating; the ciliated plates become detached from the intercellular ridges and underlying muscle layers; the intercellular ridges spread over the body surface eventually forming a new tegument; the sporocyst changes from an ovoid to a tubular shape in about 48 h at room temperature. The surfaces of the miracidium, sporocyst and cercaria of S. margrebowiei display stage-specific carbohydrates on their surfaces as indicated by lectin staining. Ricin120 stains the cilia alone of the miracidium whereas peanut agglutinin stains the larval surface except for the cilia. The intercellular ridges of the miracidium stain with concanavalin A and wheat germ agglutinin, and these lectins stain the entire surface of the mature mother sporocyst. The cercaria is the only larval stage which stains positively with asparagus pea lectin. Bulinus nasutus is incompatible with Schistosoma margrebowiei; the haemolymph of this snail contains an agglutinin which agglutinates a wide variety of mammalian erythrocytes including those of human ABO blood groups. The haemagglutinin titre of B. nasutus plasma is reduced after incubation with miracidia of S. margrebowiei indicating that the agglutinin is absorbed onto the surface of this larval stage but not that of the mother sporocyst or cercaria. The possible roles of agglutinins in host–parasite interactions together with the significance of the differences in the surface carbohydrates of the larval stages are discussed.

In the present study ultrastructural immunocytochemical techniques have been used to localize the cells producing two soluble egg antigens present in the hatching fluid of Schistosoma mansoni, using two recently described monoclonal antibodies (MAbs). In the miracidium of S. mansoni, the one apical and the two lateral unicellular glands showed immunoreactivity for the egg antigens, using a post-embedding immunogold technique. Immunoreactivity was associated with the contents of vesicles for both the MAbs, although the intravesicular distribution was not identical. This difference in ultrastructural staining pattern provides additional evidence that the two MAbs are binding with different epitopes of the same antigen or with different antigens altogether. The finding of free unbound label around the embedded miracidium is consistent with secretion having been released from the glands.

The routes of penetration and the strategies of invasion of Schistosoma japonicum miracidia in the snail vector Oncomelania hupensis were observed in a histological study. In all species of the genus Schistosoma, it is usually assumed that the miracidia achieve penetration through the tegument. Our results showed that at least 57% of S. japonicum miracidia penetrated the snail by natural openings (branchial cavity, mouth and rectum). Throughout the invasion phase, the larvae were observed in all the tissues and organs with the exception of the genital gland. The spatial distribution of parasites in the snail revealed that the migration towards the visceral organs such as the kidney, heart and sinuses (which are the most usual microhabitats of the mother sporocysts of S. japonicum) appeared to take place via the circulatory system. Using natural openings as routes for penetration probably provides a selective advantage in a host–parasite system in which the target mollusc is amphibious: we presume that the miracidia inside these natural openings are protected against desiccation when the snail leaves the water, and that they can subsequently invade the tissues.

The localization and distribution of SALMFamide immunoreactivity (IR), SI(GFNSALMFamide), in the nervous system of both the adult and larval stages of the trematode Schistosoma mansoni has been determined by an indirect immunofluorescent technique in conjunction with confocal scanning laser microscopy (CSLM). Immunostaining was widespread in the nervous system of adult male and female S. mansoni. In the central nervous system (CNS), IR was evident in nerve cells and fibres in the anterior ganglia, cerebral commissure and dorsal and ventral nerve cords. In the peripheral nervous system (PNS), IR was apparent in nerve plexuses associated with the subtegmental musculature, oral and ventral suckers, the lining of the gynaecophoric canal, and in fine nerve fibres innervating the dorsal tubercles of the male worm. In the reproductive system of male and female worms, Sl-IR was only observed around the ootype/Mehlis' gland complex in the female. Immunostaining was also evident in the nervous system of both miracidium and cercarial larval stages. A post-embedding, IgG-conjugated colloidal gold immunostaining technique was employed to examine the subcellular distribution of SALMFamide-IR in the CNS of S. mansoni. Gold labelling of peptide was localized over dense-cored vesicles within nerve cell bodies and fibres constituting the neuropile of the anterior ganglia, cerebral commissure and nerve cords of the CNS. Antigen pre-absorption studies indicated that the results obtained do suggest S1-like immunostaining and not cross-reactivity with other peptides, in particular FMRFamide.

The miracidium of Sanguinicola inermis contains a stylet and rodlet complex located within the single apical gland, which lies between a pair of lateral glands. The miracidial stylet consists of an arrangement of hollow tubes, composed of microtubules, connected at their base to large secretory granules and opening anteriorly through the apical papilla. A single rodlet lies adjacent to the stylet base and at least 8 rodlets are arranged at the apical tip of the stylet. The surface plasma membrane of the apical papilla is drawn into knob-like projections and 2 ciliated sensory endings are associated with the papilla. The ultrastructure of the apical gland, especially the stylet and rodlet complex, is related to the requirement for sanguinicolid miracidia to penetrate the double barrier of fish and snail hosts.

The localization and distribution of the serotoninergic and peptidergic elements of the nervous system of larval Schistosoma mansoni have been investigated using an indirect immunofluorescence technique in conjunction with confocal scanning laser microscopy (CSLM). A range of antisera was used, raised to the biogenic amine, 5-hydroxytryptamine (5-HT, or serotonin), the vertebrate peptides pancreatic polypeptide (PP), peptide YY (PYY) and neuropeptide Y (NPY) and to the native invertebrate peptide, FMRFamide; all these antisera were shown previously to be immunopositive in the adult worm. No immunoreactivity to 5-HT was detected in any of the larval stages, but both miracidia and cercariae were consistently immunoreactive to all 4 peptides. The peptidergic nervous system of the miracidium is relatively simple, taking the form of a central neural mass with associated paired anterior and posterior nerve tracts. The cercarial peptidergic nervous system comprises a central commissure joining paired anterior ganglia, from which emanate paired dorsal and ventral nerve tracts, which terminate at the body/tail junction. The excretory bladder region of the tail is also immunoreactive for the 4 peptides, and a fine pair of nerve tracts extends the length of the tail shaft. Immunoreactive nerve cell bodies are also evident in the midbody region of the intrasprocystic cercariae, these same structures being immunoreactive for the neuronal marker, neurone-specific enolase (NSE). The organization of the larval peptidergic nervous system is compared to that of the cholinergic nervous system and contrasted with the peptidergic system in the adult worm. The absence of immunoreactivity to 5-HT is discussed in relation to the proposed development of the aminergic nervous system upon establishment in the definitive host.

Schistosoma mansoni eggs, miracidia and primary sporocysts were labelled with phalloidin-rhodamine to visualize filamentous actin structures. Analysis of these forms by confocal fluorescence microscopy revealed the presence of previously well-defined circular and longitudinal muscle layers. Besides these muscular layers that sustain and provide motility to these parasite forms, we found in these 3 consecutive developmental stages of the parasite previously unidentified actin-rich tubular structures. In the 3 forms, 4 actin-rich tubules could be observed by optical sectioning underneath the well-developed muscle layers. The tubules appear in pairs, transversal to the length of the parasite, and located towards the extremities. By using an anti-flame cell specific antibody we confirmed that the tubules co-localize with flame cells and also determined that the tubule core is filled with microtubules. The additional presence of myosin in these tubules strongly suggests that they are contractile structures.

It is generally accepted that free-living stages of parasitic helminths are dependent on aerobic degradation of endogenous energy sources for their energy generation. This concept, however, is not the result of extensive experimental evidence, but originated mainly intuitively as oxygen is widely available in their habitat and these stages generally have a small size. Schistosoma mansoni, the sole parasitic helminth whose energy metabolism has been studied throughout its life-cycle indeed has aerobically functioning free-living stages. However, large differences exist in energy metabolism between adult stages of distinct parasitic helminths, and caution should be taken in predicting that all free-living stages of all parasitic helminths have the same, aerobic energy metabolism. Hence, this report studied the energy metabolism of Fasciola hepatica miracidia and demonstrated that F. hepatica miracidia are also dependent on aerobic degradation of their endogenous glycogen stores by glycolysis and on Krebs cycle activity for energy generation. However, in contrast to S. mansoni, F. hepatica miracidia cannot function anaerobically, as inhibition of the respiratory chain blocked motility and carbohydrate degradation, and finally resulted in death of the miracidia. Therefore, this report demonstrated that differences exist between miracidia of distinct species, in pre-adaptation of their energy metabolism to the occasional hypoxic conditions within their next host.

The avian schistosomatid Trichobilharzia ocellata plays an important role as causative agent of cercarial dermatitis of humans in Europe. In order to improve the taxonomic knowledge on this parasite, studies of miracidial chemo-orientation as well as experimental infections of different snail species were conducted using strains of T. ocellata and T. franki. Both schistosomes exhibited a high intermediate host specificity. The miracidia clearly preferred the SCW (snail-conditioned water) of the respective natural intermediate host to SCW of other sympatric snail species. T. ocellata proved to be capable of infecting Lymnaea stagnalis and Stagnicola palustris, but could not develop in Radix ovata or R. auricularia. T. franki established an infection in specimens of R. auricularia and R. ovata, but not in L. stagnalis or S. palustris. The results imply that the intermediate host spectrum of T. ocellata is limited to L. stagnalis and S. palustris. Findings of T. ocellata (or Cercaria ocellata) that originated from snails of the genus Radix are likely to have actually belonged to species such as T. franki or T. regenti. The assumption that T. szidati is synonymous to T. ocellata is also discussed.