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S R Smithers

Publications and source records attributed to S R Smithers.

At least 19 recordsLinked to original sources
PubMed1991

Sm25, a major schistosome tegumental glycoprotein, is dependent on palmitic acid for membrane attachment.

Sm25, a major antigen in the surface tegument of the parasitic helminth Schistosoma mansoni, is a 25 kDa N-glycosylated glycoprotein which co-purifies with isolated surface membranes and behaves as an integral membrane protein in Triton X-114 (TX-114). The deduced amino acid sequence of Sm25 shows a short C-terminal hydrophobic domain between residues 163 and 180, containing six uncharged polar amino acids and followed by a Lys181-Ser192 dipeptide. We were interested in whether or not this marginal C-terminal amphiphilic domain is responsible for the association of Sm25 with the membrane or whether a post-translational modification such as the addition of glycosyl phosphatidyl inositol (GPI) represents the membrane anchor for this molecule. We find that treatment with phosphatidyl inositol-specific phospholipase C, which cleaves many GPI anchors, does not reveal Cross Reacting Determinant (CRD) on Sm25, nor affect the association of this protein with membranes, providing no support for the addition of GPI. However, Sm25 is palmitoylated via a thioester bond to the single Cys residue, at position 168, which lies within the C-terminal hydrophobic domain. Removal of palmitate by reduction results in a marked decrease in the hydrophobicity of Sm25, as demonstrated by its partitioning into the aqueous rather than detergent phase of TX-114 and its quantitative release from membrane preparations. The hydrophobicity of several membrane proteins in addition to Sm25 is also decreased by reduction, raising the possibility that fatty acylation by thioester linkage is an important mechanism used by schistosomes to stabilize protein-membrane interactions.

Amino Acid Sequence↗
PubMed1991

Structure of Sm25, an antigenic integral membrane glycoprotein of adult Schistosoma mansoni.

Sm25 is the principal antigen recognised by antibodies from mice protectively vaccinated with isolated tegumental membranes of adult Schistosoma mansoni. The full-length amino acid sequence of this protein has been deduced from the sequence of two cDNAs, one isolated by screening a cDNA library and the other, including the 5' end of the gene, amplified directly from adult worm RNA using the polymerase chain reaction. The predicted sequence represents a nascent polypeptide of Mr 21,500. Following cleavage of a predicted signal sequence, the Mr of the resulting polypeptide is 17,600. The polypeptide contains 2 potential sites for N-linked glycosylation and a hydrophobic domain at the C-terminus that could facilitate membrane association. Analysis of the mature gene product confirmed that Sm25 is an N-glycosylated integral membrane protein and that the Mr of the deglycosylated polypeptide is between 15,000 and 20,000.

Amino Acid Sequence↗
PubMed1991

Molecular cloning and characterisation of the 22-kilodalton adult Schistosoma mansoni antigen recognised by antibodies from mice protectively vaccinated with isolated tegumental surface membranes.

A cDNA clone from an adult Schistosoma mansoni lambda gt11 expression library (A12) encoding an antigenic polypeptide of 22 kDa is described. A12 is 797 bp long and has one open reading frame encoding a protein of 190 amino acids which does not contain a signal sequence or membrane anchor motif and has no homologies with any sequences on the currently available data bases. Its product (sm22.6) is recognised by antibodies from mice protectively vaccinated with purified adult S. mansoni tegumental membranes and by serum from S. mansoni-infected Brazilians. It is present in all post-snail life cycle stages except the egg, is not sex-specific, and is found in 9 species of Schistosoma, but not in a range of other helminths. Data are presented which suggest that sm22.6 is a soluble, peripheral membrane protein.

Amidohydrolases↗
PubMed1990

Epitopes expressed on very low Mr Schistosoma mansoni adult tegumental antigens conform to a general pattern of life-cycle cross-reactivity.

The relationship between antigens associated with the surface of newly transformed schistosomula of Schistosoma mansoni and the tegumental surface membrane of adult S. mansoni worms has been further explored. Immunoprecipitation of detergent-solubilized 125I-tegumental surface membrane antigens of adult S. mansoni with antibodies from mice vaccinated with highly irradiated S. mansoni cercariae revealed major antigens of Mr 32, 20, 15 and 8K. The Mr 32 and 20K antigens have been previously demonstrated to be antigenically and electrophoretically identical to major antigens on the schistosomulum surface. The Mr 15 and 8K antigens, on the other hand, have not been identified by the immunoprecipitation of 125I-schistosomulum surface antigens, although a distinct schistosomulum surface antigen of Mr 15K is precipitated by antibodies from mice vaccinated with highly irradiated cercariae. Nevertheless, it was shown that antibodies to the Mr 15 and 8K antigens were specifically absorbed from vaccinated mouse serum by intact, live schistosomula, demonstrating that the Mr 15 and 8K antigens are exposed on or released from the schistosomulum surface. In contrast, absorption of the antiserum with eggs failed to remove antibody against any of the four tegumental membrane antigens examined. The Mr 15 and 8K antigens were shown to be recognized via polypeptide epitopes and not periodate-sensitive carbohydrate epitopes, further emphasizing the similarity of these to the well-characterized Mr 32 and 20K tegumental surface membrane antigens. A general relationship between schistosomulum surface, adult tegumental membrane and egg antigens was demonstrated by ELISA, using antibodies raised against the three antigenic fractions.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

A cDNA clone encoding part of the major 25,000-dalton surface membrane antigen of adult Schistosoma mansoni.

Immunoscreening of an adult Schistosoma mansoni cDNA expression library, using antibodies raised against purified adult worm tegumental surface membranes, identified a recombinant clone containing a 141-bp insert. Antibodies raised against the recombinant antigen bound specifically to the tegument of adult worms and immunoprecipitated the major 25,000-dalton surface membrane antigen as well as a 22,000-dalton nascent polypeptide generated by cell-free translation of adult S. mansoni mRNA. The mature 25,000-dalton antigen was found to be precipitated by antibodies from infected mice, rats and humans.

Amino Acid Sequence↗
PubMed1989

Antibody to carbohydrate and polypeptide epitopes on the surface of schistosomula of Schistosoma mansoni in Egyptian patients with acute and chronic schistosomiasis.

125I-Schistosoma mansoni schistosomulum surface antigens were immunoprecipitated with human antibodies from individual Egyptian patients diagnosed as being either acutely or chronically infected with S. mansoni. Both sets of patients were found to have IgG antibodies in their sera capable of immunoprecipitating the major Mr greater than 200, 38 and 32K antigens. However, the immunoprecipitation of the Mr greater than 200K antigen was found to constitute a significantly greater proportion of the total precipitate achieved with acute sera than with chronic sera. The Mr 38 and 32K antigens were more variably precipitated by the acute sera than the chronic sera but the proportion of the total precipitation that these two antigens constituted was not found to be significantly different between the two sets of sera. Immunoprecipitation with pooled antibodies absorbed with egg and adult worm homogenates which had been treated to remove either carbohydrate or polypeptide epitopes demonstrated that the Mr greater than 200K antigen was the principal target of egg-cross-reactive anti-carbohydrate antibody amongst the antigens detected. The Mr 38 and 32K antigens were found to be precipitated by antibodies to protease-sensitive and periodate-insensitive polypeptide epitopes. These results are consistent with egg-cross-reactive anti-carbohydrate IgG antibody making a greater contribution to schistosomulum surface recognition in acute infection than in chronic infection. Indeed the presence of a higher level of egg-cross-reactive and anti-carbohydrate antibody directed against schistosomulum surface epitopes in an acute serum pool than in a chronic serum pool was confirmed by measurement of antibody binding to whole schistosomula.

Acute Disease↗
PubMed1989

Variable species and stage specificity of schistosomulum surface epitopes recognized by mice vaccinated with highly irradiated cercariae.

Antibodies from mice vaccinated with highly irradiated Schistosoma mansoni or S. haematobium cercariae were used to characterize schistosomulum surface epitopes which were found to be diverse in their species and stage specificities. The epitopes recognized on the Mr greater than 200,000 and 15,000 schistosomulum surface antigens of S. mansoni and the Mr greater than 200,000 schistosomulum surface antigen of S. haematobium were found to be cross-specific whereas those on the Mr 38,000, 32,000 and 20,000 schistosomulum surface antigens of S. mansoni and the Mr 35,000, 30,000 and 24,000 schistosomulum surface antigens of S. haematobium were only immunoprecipitated by homologous antibody and are thus possible targets of the protective species-specific immunity stimulated by highly irradiated cercariae. The epitopes recognized on the Mr greater than 200,000 and 38,000 antigens of S. mansoni were shown to cross-react with both the egg and the adult worm whereas those on the Mr 32,000 and 20,000 antigens only cross-reacted with the adult worm, and those on the Mr 15,000 antigen cross-reacted with neither the adult worm nor the egg. In addition the epitopes on the Mr 38,000 and 32,000 antigens were demonstrated to be polypeptide in nature. Those on the Mr greater than 200,000, 20,000 and 15,000 antigens, on the other hand, could not be conclusively defined.

Animals↗
PubMed1989

Protective immunization of mice against Schistosoma mansoni with purified adult worm surface membranes.

Immunity to Schistosoma mansoni in the mouse was induced by vaccination with adult worm surface membrane (mb-S). Of several adjuvants tested, including Freund's, BCG and alum, 50 micrograms of saponin per mouse given subcutaneously with the antigen was the easiest to administer, and gave consistent protection, approaching levels usually seen in our mouse model after exposure to irradiated cercariae. An antibody response to the schistosomular surface was detected in mice immunized with mb-S plus saponin which was predominantly anti-polypeptide, not anti-carbohydrate, and thus similar to the antibody response of mice exposed to irradiated cercariae. The level of antibodies to Mr 90,000 and 38,000 schistosomular surface antigens as well as to Mr 25,000 adult surface membrane antigen was significantly correlated with the presence of protection.

Adjuvants, Immunologic↗
PubMed1988

Analysis of the anti-Schistosoma mansoni surface antibody response during murine infection and its potential contribution to protective immunity.

Absorption of serum from chronically infected mice with homogenized schistosome eggs reduced antibody binding to the schistosomulum surface by 94%, indicating that almost all schistosomulum surface recognition during chronic infection is due to epitopes shared with the egg. Absorption of the serum with egg homogenate from which protein antigens had been removed by boiling and digestion with proteinase K resulted in a similar reduction of antisurface antibody demonstrating that all the shared epitopes that are recognized are carbohydrate in nature. Analysis of the time course of anticarbohydrate antibody production and the levels of antibody in mice infected with a single sex of schistosome indicated that eggs directly stimulated this response. Mouse mAb were identified that bound at very high levels to the schistosomulum surface and that recognized carbohydrate epitopes shared with the egg. Three of these had previously been demonstrated to passively transfer resistance, indicating that these surface carbohydrates are potential targets of protective immunity in the mouse. All the anticarbohydrate mAb also bound to the surface of schistosomula of other schistosome species. Thus, the strong immune response against these epitopes in chronic infection could account for the cross-specific immunity observed. Mice vaccinated with irradiated cercariae lacked high levels of anticarbohydrate antibodies and their recognition of the surface was largely due to antibody to species-specific polypeptide epitopes. With respect to the Mr greater than 200,000 and 38,000 antigens, it was demonstrated that these epitopes were present on the same antigens that bear the carbohydrate moieties recognized by antibodies from chronically infected mice. This specific polypeptide recognition is also reflected in the immunity generated by exposure to irradiated cercariae.

Animals↗
PubMed1988

Serum from CBA/Ca mice vaccinated with irradiated cercariae of Schistosoma mansoni protects naive recipients through the recruitment of cutaneous effector cells.

Passive transfer experiments showed that 76% of the resistance induced in CBA/Ca mice by exposure to radiation-attenuated cercariae of Schistosoma mansoni could be transferred to naive recipients by administration of donor serum. The level of protection achieved depended on the volume of serum administered and immunity was demonstrated most consistently with serum harvested from thrice vaccinated donors. The serum was twice as effective when given to the recipients at the time of cercarial challenge as compared to administration 5 days after challenge, a result which indicates that serum-dependent challenge elimination is probably accomplished in the cutaneous tissues. This view was confirmed by the observation that passive protection could be ablated by administration of a monoclonal antibody which we have shown elsewhere to deplete the cutaneous inflammatory reaction to cercarial penetration. Histopathological studies revealed that vaccine serum induced subdermal inflammatory reactions in challenged recipient mice which were identical both in induration and kinetics to those seen in conventionally vaccinated individuals; on days 4 and 5 post-challenge, the reactions comprised 60% mononuclear cells and 40% eosinophils. Challenge larvae, which had transformed from skin-stage to lung-stage parasites, became trapped within such reactions and eventually showed generalized vacuolation consistent with the onset of damage. Some foci were seen to contain degenerated leucocytes, free eosinophil granules and debris which is thought to represent the remnants of dead parasites. Small focal reactions were identified on occasion in naive challenged mice and in recipients of normal mouse serum, but these reactions comprised predominantly mononuclear cells and were rarely seen to encompass challenge parasites. These data show that serum-transferred resistance in the vaccinated CBA/Ca mouse model involves the induction of a cutaneous inflammatory response in the recipients.

Animals↗
PubMed1987

Characterisation of the structure and expression of the gene encoding a major female specific polypeptide of Schistosoma mansoni.

A previously described cDNA clone, pSF10, of Schistosoma mansoni encoding the very dominant female specific polypeptide (FSP) has been used to characterize the gene and its expression. The gene is detectable in different isolates of S. mansoni and is estimated to be present in 3 copies per haploid genome. The gene is not sex linked and exhibits neither amplification nor rearrangement concomitant with expression. Expression of the gene by parasites maturing in hamsters is first detected after 5 weeks when the RNA is present at 1/10 the level of that of 6 week worms. Although the FSP gene is specifically and highly expressed by egg laying female worms a corresponding polypeptide produced by the cell-free translation of RNA is not detectable. It was confirmed, however, that pSF10 does indeed encode a mRNA by DNA sequence analysis. The sequence demonstrated a mRNA containing a poly(A) tail and two open reading frames. One reading contains no methionine but is very high (47%) in glycine. This amino acid composition could account for the inability to detect the gene product by cell-free translation in the presence of [35S]methionine.

Animals↗
PubMed1987

Resistance to reinfection with Schistosoma haematobium in Gambian children: analysis of their immune responses.

The relationship between reinfection with Schistosoma haematobium and immunological parameters was studied in a group of Gambian children aged from 8 to 13 years. Each individual's exposure to infection was assessed from observations of water contact, cercarial densities and infected snail densities at water contact sites. Eosinophil counts were made and responses to egg antigen (SEA) and adult worm antigen (WWH) measured by ELISA. Low levels of reinfection were associated with a high eosinophil count, high levels of antibodies against WWH and SEA, increased age and low exposure. In a multiple regression analysis of the association of reinfection with eosinophil count, antibody levels, exposure, age and sex, the effects of eosinophil count and exposure were still very significant after allowing for all the other variables. The effects of the antibody levels were close to significance after allowance for exposure and eosinophil count (for WWH: P = 0.09; for SEA: P = 0.07), although the evidence was less clear after additional allowance was made for age and sex. The ability of sera from the children to recognize different parasite antigens was also examined by immunoprecipitation of labelled schistosomulum surface, WWH, SEA and S. haematobium adult worm mRNA in vitro translation products. Schistosomulum surface antigens were recognized by all the sera and there was little variation in this response. There was more variation in their responses to SEA and WWH and a marked heterogeneity in the response to in vitro translation products. However, the pattern of antigen recognition appeared unrelated to susceptibility to reinfection.

Adolescent↗
PubMed1987

Surface antigens of and cross-protection between two geographical isolates of Schistosoma mansoni.

Two isolates of Schistosoma mansoni from Puerto Rico and Egypt were examined to determine if there were differences in surface antigens of the schistosomulum and to assess the ability of the two isolates to induce protection against one another in vivo. Immune mouse and human patient antisera recognized the same antigens on the schistosomulum surface of both isolates. However, mice immunized with schistosomula-released products from the Egyptian isolate recognized an additional antigen of Mr 13K on the Egyptian schistosomulum surface which was not present in the Puerto Rican isolate. In quantitative radioimmunoassay, sera from mice vaccinated with irradiated Egyptian cercariae bound more strongly to Egyptian schistosomula than to Puerto Rican parasites. Both isolates cross-protected against each other, but mice were less immune to challenge with Egyptian cercariae after being immunized with Puerto Rican irradiated cercariae. There was no difference in immunity to challenge when Egyptian irradiated cercariae were used to immunize. Although this evidence suggested some heterogeneity within the Egyptian isolate, cloned cercariae of the Egyptian isolate did not vary in their ability to cross-protect against each other. Furthermore, antisera from mice immunized with clones of Egyptian cercariae recognized the same schistosomulum surface antigens. The results reported here indicate that although there were small differences between the two isolates the major surface antigens are conserved.

Animals↗
PubMed1987

Surface and species-specific antigens of Schistosoma haematobium.

Of the surface antigens identified by radio-iodination, two-dimensional gel analyses showed no similarities between those of Schistosoma haematobium and Schistosoma mansoni, thus providing a basis for the species specificity of these antigens described previously (Simpson, Knight, Hagan, Hodgson, Wilkins & Smithers (1985) Parasitology 90, 499-508). The surface antigens of S. haematobium were glycosylated and comprised an acidic polypeptide of Mr 17,000 as well as a complex set of polypeptides of approximate pI 6-7, which resolved in the Mr range 20,000-30,000. At least one of the lower Mr forms of this complex is also present in the adult worm. Limited cross-reaction was observed with S. mansoni infection sera and this may be due to a shared carbohydrate epitope. In contrast, extensive cross-reaction was observed using sera from mice immunized with S. bovis. This pattern parallels the species-specificity of vaccine-induced immunity. Extensive cross-reaction was also observed within cell-free translation products of m-RNA from adult worms of S. haematobium and S. mansoni by use of heterologous human infection sera. The few antigens which were species-specific may represent surface antigens.

Animals↗