Differential adherence of M line Biomphalaria glabrata hemocytes to Schistosoma mansoni and Echinostoma paraensei larvae, and experimental manipulation of hemocyte binding

The ability of M line strain Biomphalaria glabrata hemocytes to adhere to mother sporocysts (MS) of PR1 Schistosoma mansoni or to MS or daughter rediae (DR) of Echinostoma paraensei was studied using an in vitro hemocyte adherence assay. Hemocytes were significantly more likely to bind to S. mansoni MS than to E. paraensei MS or DR. Hemocyte adherence to E. paraensei MS or DR was significantly increased if glutaraldehyde-fixed larvae were used as targets. Also, E. paraensei MS pretreated with the lectin concanavalin A (Con A) were more likely to be bound by hemocytes than MS pretreated with Con A in the presence of the competing sugar, alpha-methyl mannoside. Pretreatment of hemocytes with Con A increased their ability to bind E. paraensei sporocysts, but the effect was small compared to that achieved by pretreatment of MS with Con A. The lectin probably did not function as a bridging molecule between hemocytes and MS but, rather, altered the MS surface in a way that facilitated adherence. Similarly, adherence to E. paraensei MS was significantly increased if the MS were pretreated with cell-free M line plasma prior to use in adherence assays. Our results indicate that the two parasites provoke fundamentally different responses from M line hemocytes in vitro and that the living tegument can be modified by host humoral factors and by lectins such that hemocyte binding is significantly increased.     

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 avidin-biotin-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.     

A comparative study of mechanisms underlying digenean-snail specificity: in vitro interactions between hemocytes and digenean larvae

A panel of 4 digenetic trematode species (Echinostoma paraensei, E. trivolvis, Schistosoma mansoni, and Schistosomatium douthitti) and 5 snail species (Biomphalaria glabrata, Helisoma trivolvis, Lymnaea stagnalis, Stagnicola elodes, and Helix aspersa) was examined to determine if known patterns of host specificity could be explained by the tendency of digenean larvae to be bound by snail hemocytes, or by the ability of larvae to influence the spreading behavior of hemocytes. In short-term (1 hr) in vitro adherence assays, there was no overall pattern to suggest that sporocysts were more likely to be bound by hemocytes from incompatible than compatible snails. Compared with the other parasites, sporocysts of E. paraensei were less likely to be bound by hemocytes from any of the snail species tested. All rediae examined, including those of another species Echinoparyphium sp., were also remarkably refractory to binding by hemocytes from any of the snails. Of all the larvae examined, only sporocysts and young daughter rediae of E. paraensei caused hemocytes to round up in their presence. This was true for hemocytes from the compatible species B. glabrata and the incompatible lymnaeid species S. elodes and L. stagnalis. The patterns of host specificity shown by this particular panel of parasites and snails were not predicted by either the extent of hemocyte adherence to digenean larvae or by the ability of larvae to affect hemocyte spreading behavior. The results of this study suggest that a role for hemocytes, although likely, may require different assays, possibly of a more prolonged nature, for its detection. Also, different parasite species (notably E. paraensei) and intramolluscan stages have distinctive interactions with host hemocytes, suggesting that the determinants of specificity vary with the host-parasite combination, and with the parasite life cycle stage.     

Expression profiling and binding properties of fibrinogen-related proteins (FREPs), plasma proteins from the schistosome snail host Biomphalaria glabrata

A growing body of evidence suggests an important role for fibrinogen-like proteins in innate immunity in both vertebrates and invertebrates. It has been shown that fibrinogen-related proteins (FREPs), plasma proteins present in the freshwater snail Biomphalaria glabrata, the intermediate host for the human blood fluke Schistosoma mansoni, are diverse and involved in snail innate defense responses. To gain further insight into the functions of FREPs, recombinant FREP proteins (rFREPs) were produced in Escherichia coli and antibodies (Abs) were raised against the corresponding rFREPs. We first show that most FREP proteins exist in their native conformation in snail hemolymph as multimeric proteins. Western blot analyses reveal that expression of multiple FREPs including FREP4 in plasma from M line and BS-90 snails, which are susceptible and resistant to S. mansoni infection, respectively, is up-regulated significantly after infection with the trematode Echinostoma paraensei. Moreover, our assays demonstrate that FREPs are able to bind E. paraensei sporocysts and their secretory/excretory products (SEPs), and a variety of microbes (Gram-positive and Gram-negative bacteria and yeast). Furthermore, this binding capability shows evidence of specificity with respect to pathogen type; for example, 65-75-kDa FREPs (mainly FREP4) bind to E. paraensei sporocysts and their SEPs whereas 95-kDa and 125-kDa FREPs bind the microbes assayed. Our results suggest that FREPs can recognize a wide range of pathogens, from prokaryotes to eukaryotes, and different categories of FREPs seem to exhibit functional specialization with respect to the pathogen encountered.     

Echinostoma paraensei and Schistosoma mansoni: adherence of unaltered or modified latex beads to hemocytes of the host snail Biomphalaria glabrata.

Hemocytes derived from a strain (13-16-R1) of Biomphalaria glabrata resistant to Schistosoma mansoni were significantly more likely to bind untreated latex beads than hemocytes from the schistosome-susceptible M line strain. Beads preincubated in 13-16-R1 plasma were more readily bound by both 13-16-R1 and M line hemocytes than beads preincubated in M line plasma. Beads preincubated in plasma derived from snails of either strain infected with the trematode Echinostoma paraensei were more readily bound by hemocytes than beads preincubated in plasma from control snails of the corresponding strain. Plasma from snails exposed to S. mansoni did not have a similar effect. Throughout these experiments, beads receiving a particular treatment were consistently bound at higher rates by 13-16-R1 than M line hemocytes. SDS-PAGE of plasma components eluted from beads revealed differences between treatments, particularly in diffuse bands falling into two groups, of 75-130 and 150-220 kDa. The results indicate that both hemocytes and plasma components from the two host strains differ and identify plasma molecules deserving of additional study as possible modulators of hemocyte effector functions. Also, S. mansoni and E. paraensei provoked different responses in the same host snail.     

Echinostoma paraensei: hemocytes of Biomphalaria glabrata as targets of echinostome mediated interference with host snail resistance to Schistosoma mansoni

Earlier in vivo work by Lie et al. (1977) indicated that the innate resistance of the 10R2 strain of Biomphalaria glabrata to PR1 Schistosoma mansoni could be interfered with if the snails were infected previously with another trematode, Echinostoma paraensei. We have studied this interference phenomenon using in vitro methods in an attempt to understand its mechanistic basis. Hemolymph, derived from 10R2 snails infected with E. paraensei for 14-28 days, killed 25% of S. mansoni sporocysts in vitro, significantly less (P less than 0.001) than the 90% killing rate observed with hemolymph from uninfected, control 10R2 snails. Hemolymph from the infected 10R2 snails and from schistosome susceptible M line snails did not differ significantly (P greater than 0.1) in their relative inability to kill S. mansoni sporocysts in vitro. The defect in sporocyst killing exhibited by echinostome infected 10R2 snails was traced to the cellular, rather than the humoral, component of the hemolymph. Preparations containing uninfected 10R2 snail hemolymph and echinostome daughter rediae exhibited significantly less (P less than 0.001) killing of S. mansoni sporocysts than did controls containing only 10R2 hemolymph and S. mansoni sporocysts. Our results suggest that echinostome larvae release factors that interfere with the ability of B. glabrata hemocytes to kill S. mansoni sporocysts.     

Differential lectin labelling of circulating hemocytes from Biomphalaria glabrata and Biomphalaria tenagophila resistant or susceptible to Schistosoma mansoni infection

Lectins/carbohydrate binding can be involved in the Schistosoma mansoni recognition and activation of the Biomphalaria hemocytes. Therefore, expression of lectin ligands on Biomphalaria hemocytes would be associated with snail resistance against S. mansoni infection. To test this hypothesis, circulating hemocytes were isolated from B. glabrata BH (snail strain highy susceptible to S. mansoni), B. tenagophila Cabo Frio (moderate susceptibility), and B. tenagophila Taim (completely resistant strains), labelled with FITC conjugated lectins (ConA, PNA, SBA, and WGA) and analyzed under fluorescence microscopy. The results demonstrated that although lectin-labelled hemocytes were detected in hemolymph of all snail species tested, circulating hemocytes from both strains of B. tenagophila showed a larger number of lectin-labelled cells than B. glabrata. Moreover, most of circulating hemocytes of B. tenagophila were intensively labelled by lectins PNA-FITC and WGA-FITC, while in B. glabrata small hemocytes were labeled mainly by ConA. Upon S. mansoni infection, lectin-labelled hemocytes almost disappeared from the hemolymph of Taim and accumulated in B. glabrata BH. The role of lectins/carbohydrate binding in resistance of B. tengophila infection to S. mansoni is still not fully understood, but the data suggest that there may be a correlation to its presence with susceptibility or resistance to the parasite.     

Lectin and human blood group determinants of Schistosoma mansoni: alteration following in vitro transformation of miracidium to mother sporocyst.

A mixed agglutination assay method was employed to detect the presence of surface determinants for various lectins and human blood group antibodies on Schistosoma mansoni miracidia and cultured mother sporocysts. Miracidia were found to possess surface receptors for the lectins Con A (concanavalin A), anti-Heel (eel serum agglutinin), and anti-ADb (Dolichos seed extract), as well as human anti-A antibodies. Following in vitro transformation of the miracidium to mother sporocyst, anti-Heel and human anti-A receptors were no longer detectable on the sporocyst surface, while determinants for Con A and anti-ADb remained essentially unaltered. It is concluded that transition of the miracidium to the sporocyst results in the alteration of surface molecular structures on schistosome larve. Furthermore, since determinants for Con A, anti-Heel, anti ADb, and human anti-A have been found associated with macromolecules in the hemolymph of the snail Biomphalaria glabrata (Stnislawski et al., 1976), there is now evidence that miracidia and mother sporocysts of S. mansoni and their snail host share molecules with common lectin and human blood group determinants.     

Calcium dynamics of hemocytes of the gastropod Biomphalaria glabrata: effects of digenetic trematodes and selected bioactive compounds

Abstract. Two fluorescent calcium indicators, Calcium Green AM (CG) and Fura Red AM (FR), were used in conjunction with confocal microscopy to monitor hemocyte calcium dynamics following exposure to digenetic trematode larvae or relevant bioactive compounds. Changes in intracellular calcium levels, as measured by fluctuations in the CG/FR ratio, were correlated with hemocyte morphological changes. Hemocytes exposed to culture medium remained spread and had few calcium transients. However, following exposure to sporocysts, sporocyst secretory-excretory products, or small rediae of Echinostoma paraensei in culture medium, significantly more hemocytes both rounded up and exhibited calcium transients, though some hemocytes showed one response or the other but not both. Hemocytes did not respond significantly to large rediae, to sporocysts of another digenean ( Schistosoma mansoni ), or to bacterial lipopolysaccharides. Exposure to either zymosan particles or mannose BSA provoked responses similar to those seen with sporocysts of E. paraensei Caffeine caused rounding but no calcium transients, and phorbol myristate acetate provoked calcium transients but no rounding. The results show that sporocysts and small rediae of E. paraensei have pronounced effects on hemocyte rounding and calcium dynamics, and that these two events can occur independently of one another. This suggests that parasites may influence hemocytes in at least two separate ways.     

Inhibition of sea urchin fertilization by plant lectins

Effects of plant lectins on sea urchin (Lytechinus variegatus) fertilization and a partial characterization of lectin-binding involved in the process were evaluated. IC50 doses for inhibition of fertilization varied from 4.1 to 135.5 microg/ml when the lectins were pre-incubated with sperms and from 0.7 to 33.4 microg/ml when pre-incubated with eggs. Such effects were reversed when the lectins were heat inactivated. FITC-labeled lectins bound egg surfaces while their denatured forms did not. Glucose/mannose specific lectins bound weaker to eggs when pre-incubated with the glycoprotein bovine lactotransferrin. None of the glycoproteins assayed diminished FITC patterns of the Gal/GalNAc binding lectins. Pre-incubation of Glucose/mannose binding lectins with eggs did not alter binding of Gal/GalNAc lectins. Lectins with distinct competencies for binding monosaccharide and glycoconjugates were able to inhibit sea urchin fertilization.     

Changes in the cell surface coat during the development ofXenopus laevis embryos,detected by lectins

Summary The composition of the surface coat in embryonic cells ofXenopus laevis was examined by agglutination and fluorescent staining with lectins.Cells of early and mid gastrula stages were agglutinated by lectins specific for D-mannose, D-galactose, L-fucose, N-acetyl-D-glucosamine and N-acetyl-D-galactosamine. No differences in agglutinability among ectoderm, mesoderm and endoderm cells were observed with lectins specific for D-mannose, D-galactose and N-acetyl-D-galactosamine, though agglutination of gastrula cells with fluorescent lectins revealed considerable differences in the intensity of lectin binding among cells within an aggregate. These differences in amount of lectin bound were not related to cell size or morphology. Patches of fluorescent material formed on the cells, suggesting that lectin receptors are mobile in the plane of the plasma membrane.In the early cleavage stages intensive lectin binding occurs only at the boundary between preexisting and nascent plasma membranes. The external surface of the embryo has few lectin receptors up to the late gastrula stage. The unpigmented nascent plasma membranes, when exposed to fluorescent lectins, do not assume any fluorescence distinguishable from the background autofluorescence of yolk, in stages up to the mid-blastula. From this stage onwards lectin binding was observed on the membranes of the reverse side of surface layer cells and on the membranes of deep layer cells. During gastrulation there is an accumulation of lectin-binding material on surfaces involved in intercellular contacts.The significance of lectin binding material for morphogenesis is discussed.     

Mechanisms underlying digenean-snail specificity: role of miracidial attachment and host plasma factors

Digenetic trematodes usually show a high degree of specificity for their molluscan intermediate hosts. A panel of 4 digenean species (Echinostoma paraensei, E. trivolvis, Schistosoma mansoni, and Schistosomatium douthitti) and 5 snail species (Biomphalaria glabrata, Helisoma trivolvis, Lymnaea stagnalis, Stagnicola elodes, and Helix aspersa representing 3 gastropod families) was used to assess the relative contributions of miracidial behavior, host plasma osmolality, and host plasma factors in dictating specificity. Additional experiments were undertaken with a fifth digenean, Echinoparyphium sp. Expected patterns of compatibility were first confirmed; each parasite species produced patent infections in its known snail host, but not in the other snail species. One exception was S. douthitti, which unexpectedly did not infect L. stagnalis. As judged by direct observation and by noting their disappearance after exposure to snails, miracidia were generally less likely to attach to or penetrate incompatible than compatible hosts. However, over half of the miracidia of each parasite species attached to or attempted penetration of both compatible and incompatible hosts, suggesting that under the experimental conditions used, miracidial host location and attachment behaviors were not of overriding importance in dictating observed patterns of specificity. For each digenean species, the percentage of larvae that became immobile, rounded, showed tegumental damage, or died over a 6-hr interval in plasma of the various snails was assessed. In no case was plasma from a compatible host harmful to sporocysts or rediae. In contrast, in 8 of 16 (50%) incompatible combinations, snail plasma had a significant negative effect on sporocyst condition. In 4 of 12 (33%) incompatible combinations, plasma had a significant negative effect on rediae. In 9 of 10 combinations tested, lymnaeid plasma was toxic for the parasites of planorbid snails and in 2 of 4 combinations, planorbid plasma was toxic for the parasites of lymnaeid snails. Toxicity was not attributable to differences in plasma osmolality between snail species. The ability of plasma from incompatible snails to affect viability of both sporocysts and rediae was surprisingly strong, suggesting that humoral factors play a greater role in dictating patterns of digenean-snail specificity than previously appreciated.     

Carbohydrate localization on Gyrodactylus salaris and G. derjavini and corresponding carbohydrate binding capacity of their hosts Salmo salar and S. trutta

The congeners Gyrodactylus salaris and G. derjavini are specific ectoparasites of Atlantic salmon Salmo salar and brown trout S. trutta, respectively. To elucidate the involvement of lectin-carbohydrate interactions in this host specificity, carbohydrates on the tegument of the two species and the corresponding lectin activity of their hosts have been studied. Carbohydrate composition on the tegument differed significantly between the two gyrodactylids. Three of four commercially available peroxidase-labelled lectins with primary affinity towards D-mannoside, D-GalNAc and L-fucose bound more strongly to G. derjavini than to G. salaris. Lectins with an affinity towards D-mannoside and D-GalNAc bound significantly stronger to the cephalic lobes on G. derjavini compared to the tegument and sheaths of the hamuli. One brown trout strain and three different salmon strains were tested for lectin activity in skin and plasma. Two Baltic salmon strains and one strain from the Atlantic region were included. Brown trout differed significantly from the salmon strains when skin samples were tested for D-GalNAc activity. Lectins binding to other carbohydrates showed trends for similar host differences. The implications of carbohydrate-lectin interactions for host specificity in gyrodactylids are discussed.     

Germinal elements and their development in Echinostoma caproni and Echinostoma paraensei (Trematoda) miracidia.

Among the large cells located in the posterior of Echinostoma caproni and E. paraensei miracidia are secretory cells, germinal cells (GC), and undifferentiated cells. Secretory cells do not give rise to progeny, whereas GC do. Undifferentiated cells develop into GC that can also divide to produce embryos. Cleavage of GC of E. caproni occurs only after the parasite has entered the snail host and develops into a sporocyst. With E. paraensei, GC are larger than noted for E. caproni, and in 3 of 23 miracidia examined, germinal cell cleavage had occurred in the miracidium such that an embryo containing 20-25 blastomeres was present. Observations on the germinal elements of miracidia help to explain previous results showing that (1) E. paraensei sporocysts release mother rediae a few days earlier than do sporocysts of E. caproni, and that (2) a single mother redia is produced ahead of all others by sporocysts of E. paraensei, but not by sporocysts of E. caproni. The present study adds support to the concept that E. caproni and E. paraensei are distinct species.     

Helminth communities of Nectomys squamipes naturally infected by the exotic trematode Schistosoma mansoni in southeastern Brazil

The water rat Nectomys squamipes is endemic in Brazil and found naturally infected with Schistosoma mansoni. Helminth communities, their prevalences, intensity of infection and abundance in N. squamipes in an endemic area of schistosomiasis in the state of Rio de Janeiro, Brazil were studied. Four species of nematodes (Physaloptera bispiculata, Syphacia venteli, Hassalstrongylus epsilon and Litomosoides chagasfilhoi) were recovered in 85.3%, two trematodes (Schistosoma mansoni and Echinostoma paraensei) in 38.8% and one cestode species (Raillietina sp.) in 1.7% of rats examined. Rats were infected with up to five helminth species each, and these were highly aggregated in distribution. For H. epsilon and S. venteli, intensities and abundances were higher in adult male and subadult female hosts, respectively. Hassaltrongylus epsilon, P. bispiculata, S. venteli and S. mansoni were classified as dominant species, L. chagasfilhoi and E. paraensei as co-dominant and Raillietina sp. as subordinated. No significant correlation was found in the intensity of infecton between each pair of helminth species. Schistosoma mansoni was not related to any other helminth species according to their infection rates, althougth S. mansoni was well established in the natural helminth comunity of the water rat.     

Schistosoma mansoni modulation of phagocytosis in Biomphalaria glabrata

Both short-term (3 hr) exposure of Biomphalaria glabrata snails (M-line and 13-16-R1) to Schistosoma mansoni (PR1) miracidia and in vitro incubation of parasite sporocysts with host hemolymph components altered host phagocytic ability. Hemocytes obtained from susceptible (M-line) snails that had been exposed to parasite miracidia for 3 hr showed reduced levels of phagocytosis of yeast cells in vitro compared to hemocytes from unexposed individuals. Incubation of whole hemolymph with sporocysts in vitro also reduced yeast phagocytosis in this susceptible strain. In contrast, resistant (13-16-R1) hemocytes showed increased levels of yeast phagocytosis after in vitro incubation with the parasite, and the opsonic properties of 13-16-R1 plasma were greater after exposure of snails to miracidia. These strain-specific effects of S. mansoni on host hemocyte phagocytosis and plasma opsonization were seen only when both plasma and hemocytes were present at the time of exposure to the parasite.     

Stage-specific Differences in Lectin Binding to the Surface of Anguina tritici and Meloidogyne incognita

The occurrence and distribution of several lectin binding sites on the outer surfaces of eggs, preparasitic second-stage juveniles (J2), parasitic second-stage juveniles (PJ2), females, and males of two tylenchid nematodes, Anguina tritici and Meloidogyne incognita race 3, were compared. In both species, a greater variety of lectins bound to the eggs than to other life stages; lectin binding to eggs was also more intense than it was to other life stages. Species-specific differences also occurred. More lectins bound to the amphids or amphidial secretions of M. incognita J2 than to the amphids or amphidial secretions of A. tritici J2. Lectins also bound to the amphids or amphidial secretions of adult male and female A. tritici, but binding to the cuticle occurred only at the head and tail and was not consistent in all specimens. Canavalia ensiformis and Ulex europaeus lectins bound specifically to the outer cuticle of M. incognita. Several other lectins bound nonspecifically. Oxidation of the cuticle with periodate under mild conditions, as well as pretreatment of the nematodes with lipase, markedly increased the binding of lectins to the cuticle of A. tritici J2 but not, in most cases, to M. incognita J2 or eggs of either species.     

Schistosoma mansoni sporocysts in culture: host plasma hemoglobin contributes to in vitro oxidative stress

The initiation and promotion of sporocyst propagation and subsequent production of cercariae by intramolluscan larval stages of digenic trematodes are thought to depend on mollusc-derived factors. The ability to investigate this using in vitro cultures of Schistosoma mansoni sporocysts has been impeded by the fact that plasma from the host, Biomphalaria glabrata, becomes toxic to the parasite in long-term cultures. The present study identifies hemoglobin as the plasma component responsible for this toxicity. The addition of the enzyme catalase to sporocyst cultures neutralized the toxic effects of both purified hemoglobin and whole plasma, suggesting that the generation of H2O2 as a consequence of hemoglobin oxidation is the mechanism of plasma toxicity. Furthermore, cultures incubated in unconditioned schistosome medium with plasma plus catalase yielded significantly higher numbers of daughter sporocysts than cultures with media or plasma alone, but not higher than cultures with catalase alone. These latter results suggest that the oxidative environment and the antioxidant capacity of the media are critical factors for in vitro propagation of S. mansoni sporocysts.     

Humoral response of the snail Biomphalaria glabrata to trematode infection: observations on a circulating hemagglutinin

The hemolymph of invertebrates often contains molecules that agglutinate vertebrate erythrocytes and that may function as humoral mediators of "non-self" recognition. The objectives of this study were to 1) determine if exposure of M line or 10-R2 strain Biomphalaria glabrata snails to infection with the trematodes Echinostoma paraensei and Schistosoma mansoni could increase agglutinating activity in snail hemolymph, and 2) identify particular hemolymph molecules with such activity. In some host-parasite combinations, such as juvenile M line snails and E. paraensei, infection provoked significant elevations in titer from as early as 2 days postinfection (dpi) through 15 dpi. In other combinations, as with 10-R2 snails and E. paraensei or S. mansoni, host responses were comparatively modest, yet still measurable. In general, E. paraensei and S. mansoni elicited different responses from the same host strain, and M line and 10-R2 snails responded differently to the same parasite. Further study of the response of juvenile M line snails to E. paraensei indicated that hemolymph agglutinating activity could be inhibited by several monosaccharides (including L-fucose) and by EDTA and EGTA. An affinity column containing L-fucose agarose beads was used to purify molecules with agglutinating activity from the hemolymph of such snails. The fraction eluted from the column by 0.2 M L-fucose was shown by SDS-PAGE to contain a broad band of 80-120 kD and, less consistently, a 200 kD band. Following extensive dialysis to remove L-fucose, this fraction had agglutinating activity. As a previous study has shown that the hemolymph of E. paraensei-infected snails contains significantly increased quantities of 80-120 kD polypeptides, it is concluded that polypeptides in this size range are responsible, at least in part, for the increased hemolymph agglutination activity in such snails.     

Excretory-secretory products of Echinostoma paraensei sporocysts mediate interference with Biomphalaria glabrata hemocyte functions.

Miracidia of Echinostoma paraensei were cultured in medium containing 14C-labeled amino acids, allowed to transform into sporocysts, and their excretory/secretory products (E-S) were collected and characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis and autoradiography. Effects of E-S on hemocytes of Biomphalaria glabrata were also assessed. E-S collected during day 1 of culture (E-S1) contained several polypeptides, none of which were labeled, suggesting that E-S1 are largely preformed. E-S1 significantly depressed the ability of hemocytes to phagocytose sheep red blood cells (SRBC), but otherwise had little effect on hemocyte structure or behavior. E-S released by sporocysts in day-2 cultures (E-S2) and in older cultures generally were similar and also contained several polypeptides, many of which were labeled, indicating active synthesis of E-S in vitro. E-S2 strongly inhibited hemocyte uptake of SRBC. Also, hemocytes pretreated with E-S2 assumed a spherical shape and failed to spread normally. E-S obtained through 10 days of culture mediated this effect. Active components of E-S2 were greater than 100 kDa in their native configuration, were heat- and trypsin-labile, and were bound by anti-E-S antibodies. Both greater than 200- and 80-kDa bands were prominent in anti-E-S immunoprecipitates. Hemocytes derived from snails of the 13-16-R1 strain of B. glabrata (a strain resistant to infection with Schistosoma mansoni), when pretreated with E-S2, bound to sporocysts of S. mansoni but lost their ability to damage such sporocysts. E-S2 interfered with hemocyte functions in ways inferred from earlier classic in vivo studies of trematode-snail interactions.