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PubMed1985

Enzyme histochemical comparison of biomphalaria glabrata amebocytes with human granuloma macrophages.

In fresh water snails, amebocytes are the principal cells that react to parasitic infection. Ultrastructurally, amebocytes resemble mammalian macrophages. To clarify the relationship between amebocytes and macrophages, we compared the histochemical staining for seven enzymes in Biomphalaria glabrata snail amebocytes, both in the amebocyte-producing organ (APO) and in the encapsulation reaction formed around parasite sporocysts with the staining in macrophages from the lymph nodes of patients with sarcoid or tuberculosis. Snails were infected with Echinostoma paraensei and Schistosoma mansoni miracidia. APOs and ventricular tissue with encapsulated parasites were fixed and embedded in glycol methacrylate monomer. Hardened blocks were sectioned at 2 micron and stained for alkaline phosphatase, acid phosphatase, alpha-naphthyl acetate esterase (ANAE), ATPase, peroxidase, 5'nucleotidase, and chloroacetate esterase. The amebocyte-producing organ contained cells that were positive for acid phosphatase, ANAE, and ATPase. Amebocytes in the capsules formed around echinostome sporocysts showed stronger staining for the same three enzymes. Capsules did not form around schistosome sporocysts, but the connective tissue around them contained numerous amebocytes that were also positive for these three enzymes. The amebocyte enzyme histochemistry resembled that in human granuloma macrophages, but differed from that in neutrophils. The increased expression of enzymes in amebocytes involved in the encapsulation reaction as compared to those in the APO was reminiscent of the alterations observed when human monocytes convert to tissue macrophages. These studies support the hypothesis that the amebocyte is an "invertebrate macrophage."

Acid Phosphatase↗
PubMed1987

Structure and function of the spermatheca in a snail host of schistosomiasis, Biomphalaria glabrata.

The histochemistry and ultrastructure (SEM and TEM) of the spermatheca of Biomphalaria glabrata was investigated to elucidate the function of this organ and to compare its structure and function to similar organs found in other species. The spermatheca has a debris-filled lumen surrounded by a thin wall of tissue. The cells adjacent to the lumen are of three columnar epithelial cell types. Two cell types have abundant microvilli and mammalian cell-like organelle distribution and morphology. The above cell types differ in the electron density of their cytoplasms, nuclear morphologies, and organelle content. The third cell type differs from the other two in its cytoplasmic makeup. However, the most distinctive difference is the presence of large numbers of cilia at the apical surface with no evidence of microvilli. These columnar cells rest on a basal lamina adjacent to a two to three cell thick muscle layer. The entire organ is surrounded by an adventitia of unusual morphology. Histochemical investigation demonstrated that DNAase, RNAase, and protease are present in the lumen, alkaline phosphatase is associated primarily with the microvilli, small amounts of acid phosphatase are concentrated in the midcell area of the columnar epithelium, and ATPase activity is localized in the muscle cells and just below the absorptive surface of the microvillous cells. The luminal contents and adventitial areas are Sudan Black B positive, all areas of the lumen and organ wall are PAS positive, the cell nuclei and amorphous masses in the lumen showed Feulgen staining, and large vesicles in the columnar cells were Oil Red O positive. Apparently, the spermatheca of B. glabrata is both a digestive and absorptive structure. Although this organ shares functional similarities with those found in opisthobranchs and terrestrial pulmonates, the epithelia of the spermatheca differ dramatically in these groups.

Animals↗
PubMed2006

Baylis-Hillman adducts with molluscicidal activity against Biomphalaria glabrata.

The molluscicidal activities of ten Baylis-Hillman adducts against Biomphalaria glabrata (Say) snails, the intermediate host of schistosomiasis, have been determined. Nine of these compounds showed significant molluscicidal activity against B. glabrata, falling below the threshold of 100 microg ml(-1) set for potential molluscicidal activity by the World Health Organisation. Among these compounds, 3-hydroxy-2-methylene-3-(4-nitrophenyl)propanenitrile had the highest activity, with LC(50) = 6.64 microg ml(-1).

Animals↗
PubMed1996

New possible molluscicides from Calendula micrantha officinalis and Ammi majus. II. Molluscicidal, physiological, and egg-laying effects against Biomphalaria alexandrina and Bulinus truncatus.

In the present study, the effects of CuSo4 and crude extracts of the different parts of Calendula micrantha officinalis and Ammi majus, i. e., leaves, stems, roots, and flowers, on adult Biomphalaria alexandrina and Bulinus truncatus were investigated. Generally, leaves and flowers of both plants exhibited marked potency in killing the snail vectors of schistosomiasis. The recorded LC50 and LC90 values showed that C. officinalis was more toxic to both snails than A. majus, and B. truncatus are more sensitive to the extracts of both plants than B. alexandrina. Snails that are produced from snails previously exposed to low doses were more sensitive to the tested extracts, which may give primary indication of no possibility of inherited resistance. Moreover, prolonged exposure to the sublethal concentrations of A. majus have a definite lethal effect on the egg laying and longevity of both snails. Also, treatment with sublethal doses of both plants clearly inhibited the transaminase activity (ALAT, ASAT), diminished the total protein content, and increased markedly total lipid contents in the hemolymph of both snails.

Alanine Transaminase↗
PubMed1994

Schistosoma mansoni: modulation of hemocyte surface polypeptides detected in individual snails, Biomphalaria glabrata, following larval exposure.

As an approach to better understand the molecular mechanisms underlying the differential cellular response to larval Schistosoma mansoni observed in Biomphalaria glabrata, we have analyzed hemocyte membrane-associated polypeptides from inbred susceptible and resistant strains of B. glabrata. A combination of techniques involving biotin-labeling of cell surface-exposed polypeptides and a highly sensitive chemiluminescent detection technique has allowed the identification of major polypeptides expressed on nonadherent and adherent hemocyte subpopulations from individual snails and an assessment of the variability in their expression in response to S. mansoni exposure and latex bead injection. Results showed that S. mansoni exposure and latex bead injection differentially modulate the expression of 4 of 10 major hemocyte surface polypeptides with estimated molecular weights of 155/130, 120/110, 85, and 66 kDa. Little correlation was observed between the expression of any given polypeptide and a specific response to S. mansoni exposure. Moreover, the complex pattern of hemocyte surface polypeptides observed in snails subjected to the two treatments suggests that multiple signals may be influencing either the expression of these four polypeptides at the hemocyte surface or the hemodynamic of specific peptide-bearing hemocyte subpopulations released into the circulation. The fact that a natural and an artificial insult in the two snail strains elicit a differential response in the occurrence of these four polypeptides suggests that they may be involved in defense-related activities.

Animals↗
PubMed1994

Schistosoma mansoni: relationship between low fecundity and reduced susceptibility to parasite infection in the snail Biomphalaria glabrata.

Biomphalaria glabrata snails which were not susceptible as juveniles to infection by Schistosoma mansoni were selectively bred (by self-fertilization) from the highly susceptible NMRI laboratory snail stock. The susceptibility rate among juvenile snails derived from interbreeding NMRI parents was initially 85-95%, but after several generations of selection, less than 5% of exposed snails became infected by the parasite. Selection for low susceptibility also resulted in a large proportion of snails that displayed low fecundity and produced abnormal egg masses. Individual adult snails which were isolated from an interbreeding population of nonselected NMRI snails usually produced well-developed egg masses each containing 15-30 embryos. However, when juvenile snails from this same population were reared in isolation and not allowed to cross-fertilize, many displayed a pattern of low fecundity and abnormal egg production similar to that observed in the selected low susceptible line. Furthermore, it was found that many of the isolated snails which exhibited low egg production were also not susceptible to parasitic infection.

Animals↗
PubMed1995

Schistosoma mansoni: interleukin-1 increases phagocytosis and superoxide production by hemocytes and decreases output of cercariae in schistosome-susceptible Biomphalaria glabrata.

Decreases in the number of Schistosoma mansoni cercariae released from susceptible M-line Biomphalaria glabrata were detected following injection with the recombinant human cytokine, interleukin-1. No differences in either the time post-exposure at which shedding began or the percentage of snails shedding cercariae were detected between interleukin-1 injected, heat-inactivated interleukin-1 injected, or sham injected controls. However, sham injected and heat-inactivated interleukin-1 injected snails maintained significantly higher (approximately three-fold) levels of cercarial production compared to interleukin-1 injected snails over 8 weeks of cercarial shedding. Injection of interleukin-1 into schistosome-susceptible (M-line) and resistant (13-16-R1) strains of B. glabrata increased hemocyte phagocytosis of target particles and phagocytosis stimulated O2- production in both snail strains at 24 hr postexposure to the parasite. Resistant 13-16-R1 snails maintained, on average, 2.4 times the number of O2- producing phagocytic cells than did M-line susceptible snails, indicating that the incomplete abrogation of cercarial shedding in M-line snails may be due to an inadequate number of activated circulating effector cells in these snails. These data strongly support the contention that the evolutionarily conserved cytokine, interleukin-1, or a molecule in snail plasma with interleukin-1-like immunospecificity, biological activity, and function plays a significant role in the maintenance of susceptibility or resistance to S. mansoni infection in B. glabrata. Finally, these data also supply evidence for the evolutionary conservation of the function and role of interleukin-1, O2-, and antioxidant defense mechanisms in this host-parasite relationship.

Animals↗
PubMed1995

Schistosoma mansoni: excretory-secretory polypeptides exhibit selective binding to plasma components of the snail Biomphalaria glabrata.

Previous studies have shown that Schistosoma mansoni excretory-secretory polypeptides (ESP) inhibit various internal defense functions of hemocytes from Biomphalaria glabrata and that plasma also may exert a modulatory effect on hemocyte activity. To better understand how plasma may influence hemocyte-schistosome interactions in inbred strains of snails, biotinylated ESP (b-ESP) were used as probes to identify ESP-reactive plasma components and partially characterize the nature of their binding interactions. In a plasma binding assay, b-ESP bound in a dose-dependent fashion to immobilized snail plasma, although no quantitative differences in ESP reactivity to plasma of schistosome-susceptible (M-line) and -resistant (13-16-R1) snail strains were detected. Moreover, co-incubation of b-ESP with homologous plasma or pretreatment of plasma with nonbiotinylated ESP in the plate assay significantly reduced b-ESP binding to immobilized plasma, indicating a specific interaction between ESP and plasma. Pretreatment of b-ESP with mannose, porcine stomach mucin, fetuin, and asialofetuin resulted in a significant inhibition of b-ESP binding to plasma, whereas pretreatment of immobilized plasma with a cocktail of monosaccharides (including mannose), porcine stomach mucin, and fetuin had no inhibitory effect. These data suggest the presence of a carbohydrate-binding protein(s) in sporocyst ESP that is targeted to plasma glycoconjugates. Based on the carbohydrate content of the major inhibitory glycoproteins, galactosyl and n-acetyl-galactosaminyl sugars may represent putative determinants for the lectin-like ESP molecule(s). However, ESP binding to plasma from M-line and 13-16-R1 B. glabrata strains exhibited similar sugar and glycoprotein inhibition patterns. SDS-PAGE and electroblot analyses further demonstrated that ESP bound to a subset of separated plasma polypeptides, most prominently to a doublet of 52 and 54 kDa, and a complex of proteins with molecular masses greater than 150 kDa. Other polypeptides exhibiting weaker binding interactions included components of 34, 60, 64, 86, and 125 kDa. Although both M-line and 13-16-R1 snail strains contained a similar complement of ESP-binding plasma proteins, ESP binding to the 34- and 86-kDa proteins occurred at higher frequencies in susceptible M-line plasma. It is concluded that ESP, released during in vitro transformation of S. mansoni miracidia, contain carbohydrate-reactive proteins capable of selectively binding to components of snail plasma. Quantitative differences between snail strains in the occurrence of ESP-binding plasma molecules was documented.

Animals↗
PubMed1996

Schistosoma mansoni: use of a subtractive cloning strategy to search for RFLPs in parasite-resistant Biomphalaria glabrata.

A subtractive cloning strategy has been applied for the identification of two cDNA clones whose corresponding transcripts were elevated in Schistosoma mansoni-resistant (BS-90) compared to susceptible (M-line) snails. Clone pBS11 encoded a 1.9-kb transcript that was more elevated compared to a 500-bp transcript encoded by clone pBS12. Consequently, more attention was focused on the molecular characterization of clone pBS11. Results showed that the transcript encoded by this clone was expressed in the albumen gland and was developmentally regulated. Sequence analysis of pBS11 demonstrated the presence of an open reading frame that corresponded to a novel Biomphalaria glabrata albumen gland gene product. Comparative Southern analysis of the resistant and susceptible snail lines using pBS11 as probe indicated the presence of a BamHI and EcoRI RFLP between the two strains.

Animals↗
PubMed1997

Schistosoma mansoni: distribution patterns of miracidia among Biomphalaria glabrata snail as related to host susceptibility and sporocyst regulatory processes.

Parasite prevalences, miracidia developmental capacity, mother sporocyst mean intensities, sporocyst distribution patterns, and cercarial production levels were determined after individual exposure of Biomphalaria glabrata snails to increased doses of Schistosoma mansoni miracidia for two geographical strains (Brazilian, BRE, and Guadeloupean, GUA) of host and parasite. For a high level (100%) of host-parasite susceptibility and in the absence of mother sporocyst regulatory processes for the BRE combination, parasites were randomly dispersed among snail hosts with a frequency distribution conforming to a positive binomial. In contrast, for a moderate level (65%) of host-parasite susceptibility and in the presence of mother sporocyst regulatory processes for the GUA combination, parasites were overdispersed among snail hosts with a frequency distribution conforming to the negative binomial. Levels of cercarial production were found to be strain dependent, to be determined during early development of mother sporocysts, and to be correlated with the number of developed mother sporocysts. Results were analyzed in the general context of the infrapopulation dynamics of the intramolluscan stages of trematode and are discussed in terms of their consequences on the distribution of the genetic diversity of adult schistosomes among the definitive host population.

Animals↗
PubMed1997

Schistosoma japonicum: in vitro cultivation of miracidium to daughter sporocyst using a Biomphalaria glabrata embryonic cell line.

In vitro cultivation of Schistosoma japonicum miracidia to the mother sporocyst (MS) and then to the daughter sporocyst (DS) stage was achieved using the Biomphalaria glabrata embryonic (Bge) cell line as a coculture system. When comparing the effect of Bge cell and MS density on MS development, it was apparent that Bge cell density had a highly significant effect on both MS viability and growth. Viability and growth rate of MS cultured under high cell density conditions (350 cells/mm2) were almost 2 times greater than those of MS cultured under conditions of low cell density (60 cells/mm2). Growth under high cell density conditions corresponded to a 20 to 30 times increase in MS estimated volume within the first 9 weeks of cultivation. Emergence of fully formed motile DS was first observed after 11 weeks of cocultivation. A few DS lived for 14 weeks after emergence and attained a size of 770 +/- 100 microns in length and 48 +/- 13 microns in width. In contrast to what was observed in Bge cell/Schistosoma mansoni cocultures, Bge cells did not encapsulate S. japonicum MS. Our results show that, although the cellular interactions between Bge cells and schistosomes MS display some level of specificity, Bge cells apparently secrete soluble factors that permit excellent survival and can trigger advanced in vitro development of S. japonicum.

Animals↗
PubMed1998

Characterization of an intracellular receptor for activated protein kinase C (RACK) from the mollusc Biomphalaria glabrata, the intermediate host for Schistosoma mansoni.

A receptor for activated protein kinase C (RACK) was characterized from the mollusc Biomphalaria glabrata, the intermediate host for the human parasite Schistosoma mansoni. This protein was shown to possess structural and functional characteristics of other RACK proteins from various cells and organisms. Its ability to bind mammalian PKCs also confirmed the conservation of PKC and RACK interactive domains throughout evolution. Results of immunolocalization indicated the presence of Bg RACK in the cytoplasm of mollusc hemocytes and B. glabrata embryonic (Bge) cells with a more intense staining around the nucleus. These results are in agreement with the association of RACK proteins with cytoskeletal elements.

Amino Acid Sequence↗
PubMed1993

Polypeptides synthesized in vitro by Biomphalaria glabrata hemocytes bind to Schistosoma mansoni primary sporocysts.

In vitro labeled polypeptides secreted by schistosome-susceptible and -resistant Biomphalaria glabrata were tested for their ability to bind the surface tegument of Schistosoma mansoni primary (= mother) sporocysts. Out of a complex pattern of SDS-PAGE-separated hemocyte polypeptides, only two (19 and 46 kDa) bound to sporocysts. The 19-kDa polypeptide consistently bound to both live and fixed sporocysts, although fixed larvae usually bound more than living sporocysts. The 46-kDa component bound primarily to fixed sporocysts. A similar pattern of binding was seen when sporocysts were exposed to polypeptides secreted into culture medium by both resistant and susceptible snail hemocytes. However, previous work indicates that sporocyst secretory products, in the presence of homologous snail plasma components, can differentially influence the quantity of these two polypeptides. Therefore, it is hypothesized that "regulation" of the interaction of snail hemocyte polypeptides with the parasite may lie in the sporocyst-mediated differential effect on synthesis and/or secretion of the 19- and 46-kDa polypeptides by hemocytes of these two snail strains. Thus, the hemocyte polypeptides reported in this study may represent potential mediators of self/non-self-recognition or hemocyte activation in the B. glabrata internal defense system.

Animals↗
PubMed1994

Surface membrane polypeptides associated with hemocytes from Schistosoma mansoni-susceptible and -resistant strains of Biomphalaria glabrata (Gastropoda).

Previous studies have shown that hemocytes from inbred susceptible and resistant strains of the snail, Biomphalaria glabrata, differentially react to primary sporocysts of the blood fluke, Schistosoma mansoni. It has been hypothesized that this differential reactivity may be due to differences in the expression of hemocyte surface determinants serving as parasite recognition and/or hemocyte activating factors. In order to begin addressing this hypothesis the hemodynamics and surface polypeptides of circulating hemocytes from susceptible and resistant B. glabrata strains were compared in two morphologically distinct cellular subpopulations that differed in their substrate adherence properties. When compared to a S. mansoni-resistant snail strain, two susceptible strains exhibited overall lower total circulating cell numbers and a consistently lower proportion of nonadherent hemocytes. Exposure of snails to S. mansoni miracidia induced significant increases in total circulating cell number in all strains by 2 days postexposure (PE). However, the proportion of adherent and nonadherent hemocytes remained constant in each snail strain at 2 and 4 days PE. A sensitive technique involving biotin labeling of polypeptides at the surface of living hemocytes, followed by nonreducing SDS-PAGE separation and electrotransfer to nitrocellulose, revealed a similar pattern of hemocyte surface polypeptides ranging in apparent molecular weights from 210 to 30 kDa in the three B. glabrata strains. One exception was the presence of a 66-kDa polypeptide expressed at the surface of both adherent and nonadherent hemocytes in the two susceptible snail strains, which was only weakly expressed or absent from hemocytes of the resistant 13-16-R1 B. glabrata strain.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Biomphalaria glabrata hemolymph lectins: binding to bacteria, mammalian erythrocytes, and to sporocysts and rediae of Echinostoma paraensei.

Polyclonal antibodies were raised in rabbits to two groups of diffusely staining M line Biomphalaria glabrata plasma polypeptides, of 150-210 and 70-120 kDa, designated as Group 1 molecules (G1M) and group 2 molecules (G2M), respectively. G1M and G2M are known to increase in abundance and to become more diverse following infection of B. glabrata with the digenetic trematode Echinostoma paraensei. These antibodies were used in conjunction with immunoblotting and slot blotting procedures to document binding of G1M/G2M from plasma of unexposed control snails (C plasma) or plasma from snails with 8-day infections of E. paraensei (I plasma) to various targets. Binding of G1M/G2M to both gram-positive and gram-negative bacteria, human A, B, and O and rabbit erythrocytes, and to sporocysts and rediae of E. paraensei was documented. Immunoblots revealed that erythrocytes are bound by a particular G2M band of 100-120 kDa present in I plasma that is in low concentration or lacking in C plasma. This explains previous results indicating that I plasma agglutinates all 4 erythrocytes examined whereas C plasma agglutinates only rabbit erythrocytes. More G1M/G2M binding to sporocysts occurred if I plasma rather than C plasma was used for incubations. Also, monosaccharide inhibition of G1M/G2M binding to sporocysts was observed in I plasma but not in C plasma. The results indicate that infection with E. paraensei induces production by B. glabrata of unique plasma polypeptides and that molecules present in I plasma can bind to the surfaces of non-self objects including E. paraensei larvae in a lectin-like fashion.

Animals↗
PubMed1998

Comparison of several types of allografts in Biomphalaria glabrata (Mollusca: Pulmonata).

Interpretation of prior studies on molluscan allografts is complicated by the variety of experimental methods used and recurring reports of acute rejection of digestive gland implants. In this study, allografts of adult digestive gland, saccular kidney, mantle, albumin gland, brain, ovotestis, heart, or amoebocyte-producing organ (APO) were implanted into the hemocoel of Biomphalaria glabrata snails, and their histological condition at 60 days post implantation (DPI) was compared with that of preimplantation tissues. Also, digestive gland tissue, still surrounded by body wall, was implanted from immature donors and examined at 120 DPI. All 60-day allografts maintained some degree of normal cell and tissue structure, and most displayed additional evidence of viability, i.e., contraction (heart), hyperplasia (APO), gametogenesis (ovotestis), secretion (albumin gland, digestive gland, and kidney), shell deposition (mantle), or neuroma formation (brain). Recipient hemocytic reaction was minimal or absent, except that in three of six ovotestis implants and five of seven digestive gland implants, variable numbers of acini were undergoing phagocytosis by hemocytes or had been reduced to granulomas. Among seven implants examined at 120 DPI, digestive gland acini were found in five (with one instance of hemocytic encapsulation). Thus, in this study, acute rejection of allografts did not occur. Although the observed hemocytic reactions could be viewed as evidence of chronic graft rejection, the inconsistent and protracted nature of these responses suggests instead that they represent resorption of degenerating tissue damaged by nonimmunological effects, e.g., mechanical trauma, heterotopic location, or autolysis.

Animals↗
PubMed1998

Susceptibility of Biomphalaria glabrata to infection with Echinostoma paraensei: correlation with the effect of parasite secretory-excretory products on host hemocyte spreading.

Susceptibility of the snail Biomphalaria glabrata to infection with the digenetic trematode Echinostoma paraensei was correlated with the ability of secretory-excretory products (SEP) derived from sporocysts of this parasite to interfere with the spreading behavior of host hemocytes in an in vitro assay. In 15 separate experiments, the prevalence of infection achieved by miracidia was positively correlated (C.C. = 0.779, P < 0.001) with the ability of in vitro-transformed sporocysts derived from the same batches of miracidia to produce SEP that inhibited host hemocyte spreading. Under assay conditions reflecting differences in hemolymph volumes between juvenile snails (susceptible to infection) and adult snails (relatively refractory to infection), SEP had a significantly greater effect on hemocytes from juvenile snails. No significant differences in response to SEP were noted when equivalent numbers of adult and juvenile hemocytes were used in the assay. Snails of the M line strain of B. glabrata are significantly more susceptible to infection with E. paraensei than 13-16-R1 strain snails. Exposure to SEP significantly increased the number of unspread hemocytes for both strains. However, significantly more 13-16-R1 than M line hemocytes remained spread following SEP treatment. Echinostoma paraensei sporocyst SEP effects on host hemocyte spreading mirror observed patterns in both age- and strain-related susceptibility of B. glabrata to this parasite. The results suggest that the number of hemocytes available to a particular snail influences its vulnerability to infection with E. paraensei.

Animals↗
PubMed1999

Internal defenses of the snail Biomphalaria glabrata.

We describe three distinct types of cells among Biomphalaria glabrata hemocytes: large cells with a tubulo-vesicular compartment, a component of the endocytic system, and with numerous mitochondria and large aggregates of glycogen particles; medium-size cells poor in organelles and glycogen; and small cells with organelles and few secretory granules. Other small hemocytes can be interpreted as juvenile cells. B. glabrata hemocytes contain few enzymes and do not show specific secretory granules, except for a subpopulation of large cells richer in acid phosphatase vesicles. Hemocytes have different aspects corresponding to different physiological states and their transitions: in quiescent hemocytes, the cell cortex is narrow and organelles are scattered in the cytoplasm, both in circulating cells characterized by thin-folded filopods and large macropinocytic vacuoles and in sedentary cells in which extended filopods connect to the extracellular matrix. In stress-activated hemocytes, the cortical region is thickened by polymerization of actin, and organelles are gathered around the nucleus. Fixed phagocytes are components of the connective tissue; the presence of numerous lysosomes and residual bodies and of acid phosphatase and peroxidase activities suggests a high phagocytic activity.

Acid Phosphatase↗