Schistosoma mansoni TGF-beta receptor II: role in host ligand-induced regulation of a schistosome target gene

Members of transforming growth factor-beta (TGF-beta) superfamily play pivotal roles in development in multicellular organisms. We report the functional characterization of the Schistosoma mansoni type II receptor (SmTbetaRII). Mining of the S. mansoni expressed sequence tag (EST) database identified an EST clone that shows homology to the kinase domain of type II receptors from different species. The amplified EST sequence was used as a probe to isolate a cDNA clone spanning the entire coding region of a type II serine/threonine kinase receptor. The interaction of SmTbetaRII with SmTbetaRI was elucidated and shown to be dependent on TGF-beta ligand binding. Furthermore, in the presence of human TGF-beta1, SmTbetaRII was able to activate SmTbetaRI, which in turn activated SmSmad2 and promoted its interaction with SmSmad4, proving the transfer of the signal from the receptor complex to the Smad proteins. Gynaecophoral canal protein (GCP), whose expression in male worms is limited to the gynaecophoric canal, was identified as a potential TGF-beta target gene in schistosomes. Knocking down the expression of SmTbetaRII using short interfering RNA molecules (siRNA) resulted in a concomitant reduction in the expression of GCP. These data provide evidence for the direct involvement of SmTbetaRII in mediating TGF-beta-induced activation of the TGF-beta target gene, SmGCP, within schistosome parasites. The results also provide additional evidence for a role for the TGF-beta signaling pathway in male-induced female reproductive development.     

Schistosoma mansoni: effect of a protein-free host diet on tegument.

The effect of a protein deficiency in the host's diet on the tegument of Schistosoma mansoni is described. Both the infected and the uninfected hamsters, fed on the diet, were stunted in growth; but the effect of the diet was more pronounced on the infected hamsters. The parasites recovered from both the liver and the mesenteric veins of animals fed on the diet from the time of infection were also retarded in growth. The tegument of both groups of parasites were reduced in height as compared with the tegument of control worms. The worms recovered from the liver of the hamsters were less adversely affected than the worms recovered from the mesenteric veins, in the sense that the tegument did not show any sign of degeneration, as was found in the latter group of parasites. In the worms from the mesenteric veins, the external plasma membrane was approximately half the thickness of the external plasma membrane of control worms. The invaginations of the external plasma membrane of experimental worms penetrated deeply into the tegument and in most instances almost reached the basal plasma membrane. Prolonged feeding of the hosts on the experimental diet resulted in the disintegration of the tegument in localised areas of the body. There was no adverse effect on adult worms of an established infection after the hosts were transferred to the protein-free diet for up to 3 wk. The ability of the tegument to regenerate after transferring the hosts from the experimental diet to normal laboratory diet (control diet) was demonstrated.     

Effects of intermediate host genetic background on parasite transmission dynamics: a case study using Schistosoma mansoni

For parasites that require multiple hosts to complete their development, genetic interplay with one host may impact parasite transmission and establishment in subsequent hosts. In this study, we used microsatellite loci to address whether the genetic background of snail intermediate hosts influences life-history traits and transmission patterns of dioecious trematode parasites in their definitive hosts. We performed experimental Schistosoma mansoni infections utilizing two allopatric populations of Biomphalaria glabrata snails and assessed intensities and sex ratios of adult parasites in mouse definitive hosts. Our results suggest that the genetic background of hosts at one point in a parasite’s life cycle can influence the intensities and sex ratios of worms in subsequent hosts.     

Possible mechanisms of the decoy effect in Schistosoma mansoni transmission

Combes C. and Moné H. 1987. Possible mechanisms of the decoy effect in Schistosoma mansoni transmission. International Journal for Parasitology17: 971–975. Three main possible mechanisms of the decoy effect have been showed in Schistosoma mansoni transmission: (1) miracidia penetrated into the non-target molluscs and then degenerated; (2) miracidia were exhausted by trying to penetrate the nontarget molluscs, excitation of miracidia was more specific than the penetration behaviour; (3) the defense mechanisms of the target mollusc Biomphalaria glabrata were stimulated by the miracidia that have been in contact with non target molluscs. The authors concluded in classifying the molluscs in three types of ranges: interference range, penetration range, and developmental range.     

Schistosoma mansoni: ultrastructural observations on the small intestine of murine host

Between 6 and 29 weeks, the small intestines of mice infected with Schistosoma mansoni were studied with the electron microscope. Granulomas were confined to the serosal-muscularis regions of the small intestine. Early granulomas were characterized by having several cell types with the most conspicuous type being the eosinophil. Older granulomas were more fibrotic. These were compared with hepatic granulomas of comparable age. In the vicinity of active eggs either in granulomas or in the lamina propria, mitochrondria from epithelial cells displayed intracristal granules. Intraperitoneal injections of soluble egg antigen isolated from viable S. mansoni eggs produced identical mitochondrial abberations. Cytochrome c-cytochrome oxidase activity was visualized in the mitochondria by the diaminobenzidine method.     

Inter-specific parasite competition: mixed infections of Schistosoma mansoni and S. rodhaini in the definitive host

Competition between parasite species has been predicted to be an important force shaping parasite and host ecology and evolution, although empirical data are often lacking. Using the Mus musculus-Schistosoma mansoni and Schistosoma rodhaini host-parasite systems we characterized mate choice and inter-specific competition between these two schistosome species. Simultaneous infections revealed species-specific mate preferences for both species as well as suggesting mating competition, with male S. rodhaini appearing dominant over male S. mansoni. S. rodhaini homologous pairs were also shown to have increased reproduction per paired female in the presence of a competitor in simultaneous infections. Overall total reproductive success was, however, similar between the two species under conditions of direct competition due to the greater initial infectivity of S. mansoni in comparison to S. rodhaini. Inter-specific competition was also implicated as increased parasite virulence to the host. The potential effects of such interactions on parasite and host ecology and evolution in nature are discussed.     

Schistosoma mansoni: degradation of host extracellular matrix by eggs and miracidia

A radioactively labeled in vitro model of the extracellular matrix of the mammalian intestinal wall and of snail tissue was used to determine whether proteolytic enzymes released by eggs and miracidia of Schistosoma mansoni could degrade connective tissue macromolecules in the type of interactive framework found in vivo. Eggs were collected and miracidia hatched in the presence of antibiotics to eliminate bacterial contamination. Uninfected livers were used as controls to ensure that the tissue dissociation and egg collection procedures did not produce proteolytic activity. One thousand live eggs incubated with the extracellular matrix for 72 hr at 37 C degraded 31% of the glycoprotein in the matrix; there was no degradation of elastin or collagen. Medium conditioned by incubation with eggs degraded 60% as much of the matrix as the live eggs themselves. The proteolytic activity of the egg-conditioned medium was greater in the presence of dithiothreitol. Miracidia incubated with the extracellular matrix in tissue culture medium at 27 or 37 C rapidly transformed to living sporocysts. This transformation was accompanied by a release of proteolytic activity, resulting in the degradation of 49 to 58% of the glycoprotein in the extracellular matrix by 1000 miracidia. Again, no elastin or collagen was degraded. The time course of degradation by miracidia was rapid over 24 hr and thus similar to that previously reported for cercariae. Degradation by eggs occurred more slowly over 72 hr. These data confirm that both eggs and miracidia secrete proteinases which are capable of degrading at least the glycoprotein components of extracellular matrix to facilitate their migration through intestinal wall or penetration of snail tissue.     

Schistosoma mansoni: functional proteasomes are required for development in the vertebrate host

Proteasomes are multi-subunit proteases involved in several mechanisms and thought to contribute to the regulation of cellular homeostasis. Here, we report for the first time biochemical evidence for the existence of a ubiquitin-proteasome proteolytic pathway in this parasite. Proteasomes from both cercariae and adult worms exhibited a high preference for hydrolysis of the substrate Suc-LLVY-AMC, although in the cercariae extract the rate of hydrolysis was 50% lower when compared to adult worms extracts. The same difference in proteasome activities was observed when endogenous proteins were broken down in the presence of ATP and ubiquitin. Additionally, accumulation of high molecular weight conjugates was observed when cercariae were pre-incubated with proteasome inhibitors. Finally, we present evidence that during experimental schistosomiasis, proteasome inhibitors were able to reduce the number of lung stage schistosomula, reduce the worm burden and consequently decrease the egg output in infected mice.     

Schistosoma mansoni: relationship between cercarial production levels and snail host susceptibility

Two populations of Biomphalaria glabrata snails differing slightly in their susceptibility to Schistosoma mansoni infection showed dramatic differences in cercarial output per snail. Exposed to five or more miracidia, snails from a group with a 90-100% susceptibility rate (Group A) produced nearly twice the number of cercariae as those from a group with a 70-80% susceptibility rate (Group B). Exposure of individual snails to known numbers of miracidia resulted in higher numbers of primary (mother) sporocysts in Group A snails than in Group B snails. However, monomiracidial exposure of snails from both groups resulted in equivalent numbers of cercariae produced per positive snail, indicating that, once established, all primary sporocysts possess a similar reproductive potential. Morphometric analysis of serially sectioned 9-day-old primary sporocysts supported this conclusion; the size of the primary sporocysts and the size and numbers of secondary (daughter) sporocysts within each primary sporocyst were comparable in snails from both groups. The data indicate cercarial production in this system is regulated prior to, and/or during, early development of the primary sporocyst.     

Schistosoma mansoni: immediate hypersensitivity to adult antigens in the rat

Antigen fractions from adult S. mansoni, obtained from infected mice, were isolated by a variety of methods. A readily soluble fraction was obtained in good yield by freezing and thawing the schistosomes, while the less soluble residue was fractionated by the use of a number of the methods currently used for the extraction of tissue and cell surface antigens. The dialyzed, centrifuged products were characterized by acrylamide gel disc electrophoresis methods, agar gel precipitin reactions with antisera from rabbits immunized with whole schistosome homogenate, and by Prausnitz-Kustner (P-K) assay with sera from schistosome infected rats. The pattern of P-K reactivity suggested that there were a number of different antigen specificities involved in the reaginic antibody response to schistosome infection in rats. With repeated infection and increased duration of infection, more different antigens seemed to be involved in the reagin response. The schistosome antigen fraction obtained by freezing and thawing was especially reactive with both early infection rat sera and sera from multiply infected rats. Both the soluble fraction isolated by freezing and thawing and residue solubilized materials were found to be able to induce the formation of reagin antibodies on immunization with alum and B. pertussis vaccine.     

Schistosoma mansoni: the egg, biosynthesis of the shell and interaction with the host

The schistosome eggshell is a hardened and tanned structure made from cross-linked proteins. It is synthesized within the female worm from many different kinds of proteins and glycoproteins. Once the egg is released in the circulation, the outer surface of the eggshell is exposed and hence a direct site of interaction between the parasite and the host. The major eggshell protein is p14, but about one third of the eggshell is made from common cellular proteins, some of which are known to be immunogenic. This has many consequences for parasite-host interactions. However, so far, the eggshell has gained little attention from researchers. We will discuss the structure of the eggshell and its role in granuloma formation, host factor binding and egg excretion.     

Schistosoma mansoni: sex-specific modulation of parasite growth by host immune signals

Development of female schistosomes from infectious cercariae to mature egg-producing adults requires both male schistosomes and an intact adaptive immune system. By examining single sex infections in immunodeficient mice, we provide evidence that female schistosome development is not directly influenced by the adaptive immune system, whereas male development is. Our data are consistent with a sequential model of schistosome development, where the adaptive immune system signals development of mature males, which subsequently stimulate development of mature females. The male schistosome therefore appears to play a central role both in transducing signals from the adaptive immune system and in facilitating female development.     

Schistosoma mansoni: cryopreservation of schistosomules.

Conditions were established for recovery of active schistosomules of Schistosoma mansoni after cryopreservation and storage in liquid nitrogen (?196 C). Schistosomules prepared from cercariae by a shear pressure technique were subjected to a two-step cooling process consisting of a slow cooling rate to an intermediate temperature, followed by rapid quenching of the sample in liquid nitrogen. Overall averages of 39 and 44% of the schistosomules, with a maximum of 88%, were recovered retaining normal activity with cooling rates of 0.4 C/min to ?32 C or 0.8 C/min to ?35 C, respectively. Methanol at 17.5% in Earle's lactalbumin hydrolysate was the freezing medium. As compared with 24 hr storage in liquid nitrogen, no loss in schistosomule motility was observed after 1 month. Following cryopreservation, attenuated schistosomules derived from ⁶⁰Co-irradiated cercariae (50 kR) exhibited structure and activity equivalent to that of unattenuated schistosomules. Infectivity for mice of unattenuated schistosomules derived from ⁶⁰Co-irradiated cercariae (50 Krad) exhibited structure and activity of unfrozen schistosomules ranged from 0.4 to 15.2%.     

Intraspecific variations in the hemolymph of Biomphalaria glabrata, a snail host of Schistosoma mansoni.

An attempt was made to characterize the hemolymph of Biomphalaria glabrata with reference to "normal" intra-specific variation, i.e., both inter- and intra-strain differences. Total protein concentration, per cent hemoglobin, pH, and osmolarity were studied. Seven geographic strains of B, glabrata were examined. In addition, observations were made on the hemolymph of Biomphalaria straminea, several strains of Helisoma caribaeum, and on B. glabrata subjected to infection with Schistosoma mansoni or to periods of starvation. Intra-strain differences in total protein concentration and total hemoglobin concentration in B. glabrata appeared to be more closely related with snail size than with absolute age. Inter-strain variation in B. glabrata was also noted, but the differences were of the same magnitude as those from intra-strain samples. Significant differences in total protein concentration were observed, however, between the means of similar size B. glabrata, B. straminea and H. caribaeum. The osmolatity of the hemolymph from different size B. glabrata was similar as were the osmolalities of the hemolymph from similar size snails of different strains. However, all B. glabrata strains exhibited hemolymph osmolalities lower than observed in strains of H. caribaeum. Infection with S. mansoni reduced the protein concentration of B. glabrata hemolymph. Differences were noted as early as 1.5-24 hr post-infection, with significant alterations occurring at about 11 days post-infection. To a lesser extent, starvation also depleted the protein content of the hemolymph.     

Purification and characterization of 3-hydroxymethylglutaryl-coenzyme A reductase of Schistosoma mansoni: regulation of parasite enzyme activity differs from mammalian host

The enzyme 3-hydroxymethylglutaryl-CoA (HMG-CoA) reductase plays a critical role in regulating the production of cholesterol, dolichols, and ubiquinones in mammals. The inhibition of this enzyme in Schistosoma mansoni is accompanied by a cessation of egg production by the female parasite and a reduced ability of the parasite to properly glycoslyate their proteins. Furthermore, we recently demonstrated that mevinolin, if given continuously over a period of 10-14 days, is a potent antischistosomal drug. In this paper, we describe the properties of purified HMG-CoA reductase from S. mansoni. Using affinity chromatography, we were able to obtain a 417-fold purification of the enzyme which had Km values similar to the rat enzyme for HMG-CoA and NADPH. The Ki value for mevinolin, a potent and selective inhibitor of the rat reductase (Ki = 0.6 nM), was significantly higher (Ki = 46 nM) for the schistosome enzyme. SDS-PAGE and HPLC of the purified enzyme resulted in the appearance of a single protein, which had a molecular weight (66,000) in the range reported for the rat enzyme. Parasite reductase activity, unlike that of its host, did not display a circadian rhythm. Furthermore, agents which elevate (cholestyramine) or decrease (cholesterol) mammalian reductase activity had no effect on the parasite enzyme. Our results suggest that the mechanism which regulates production of the parasite's enzyme may differ from its mammalian host.