Induction of Cytotoxic T-Cell Activity by the Protective Antigen of Schistosoma mansoni Sm28GST or its Derived C-Terminal Lipopeptide

In a previous work the authors demonstrated that immunization with Schistosoma mansoni 28-kDa glutathion-S-transferase (Sm28GST) was able to reduce hepatic damage in infected mice and that the adoptive transfer of Sm28GST-specific T cells reproduced the protective effect obtained with the recombinant molecule. In the present paper, the authors show that Sm28GST is also able to stimulate an antigen-specific, cytotoxic T-cell response against Sm28GST-pulsed P815 target cells in normal mice and that effector cells induced in vivo were classical Class I MHC-restricted CD8⁺ lymphocytes. The authors found no spontaneous CTL activity against Sm28GST-pulsed target cells during the course of the infection by S. mansoni although Sm28GST is expressed at different developmental stages of the parasite. It was observed, however, that immunization with Sm28GST is sufficient to elicit a significant level of CTL response for 6 weeks in infected mice. The role of these Class I MHC-restricted CD8⁺ lymphocytes in the protection observed precisely at the same period in immunized mice remains to be elucidated. The authors also observe that immunization with the lipopeptidic form of the C-terminal peptide of the molecule (190–211 peptide) led to a CTL activation comparable to that observed after immunization with the whole molecule demonstrating the feasibility of using a synthetic lipopeptide as immunogen for a CTL response against Sm28GST epitopes. Moreover, like Sm28GST-specific CTLs, 190–211 lipopeptide-specific cells were also Class I MHC-restricted lymphocytes.     

Comparison of the cloned genes of the 26- and 28-kilodalton glutathione S-transferases of Schistosoma japonicum and Schistosoma mansoni.

Both Schistosoma japonicum and S. mansoni contain 28- and 26-kDa glutathione S-transferases (GSTs). Despite their immunological cross-reactivity using rabbit antisera, the S. japonicum 28-kDa GST (Sj28) is weakly immunogenic relative to the S. mansoni protein (Sm28) in mouse immunization experiments using GSTs purified from adult worms. The difference in immunogenicity is also observed during schistosome infection in mice. Using surface-labeled living S. japonicum worms, evidence was obtained for a surface location of Sj28 comparable to that reported for the S. mansoni molecule. The nucleotide and deduced amino acid sequences of cDNA clones corresponding to Sj28 and Sm28 were compared. Despite obvious homology (77% identity), differences were found in regions known to contain T epitopes in the S. mansoni protein which may be an explanation for the striking differences in immunogenicity in regard to antibody production in mice. The 26-kDa GSTs of these two parasites (Sj26 and Sm26) are also closely related on the basis of nucleotide and deduced amino acid sequences, there being 82% identity in the putative coding regions. When the amino acid sequences of Sj28 and Sm28 were compared with those of Sj26 and Sm26, the overall sequence identity was approximately 20%. However, a relatively conserved region was identified in otherwise structurally different molecules which may participate in common properties of these enzymes.     

Molecular cloning and tissue distribution of a 26-kilodalton Schistosoma mansoni glutathione S-transferase

A Schistosoma mansoni cDNA library was constructed from the mRNA of adult worms in the expression vector lambda gt11 and screened with a rabbit antiserum raised against the 26-kDa S. mansoni glutathione S-transferase isoforms (Sm GST 26). Two clones were selected and the nucleotide sequences deduced. The predicted amino acid sequence, specified by these cDNAs, shows strong homology with a Schistosoma japonicum 26 kDa glutathione S-transferase and a lower level of homology with mammalian glutathione S-transferase class mu isoenzymes (EC 2.5.1.18). No significant homology score was found with a 28-kDa S. mansoni glutathione S-transferase (Sm GST 28). A study of the tissue distribution of the cloned Sm GST 26 by immunoelectron microscopy shows similarities to Sm GST 28 in that they are present in the tegument and in subtegumentary parenchymal cells. However, a major difference exists in the protonephridial region in which Sm GST 26 is present in the cytoplasmic digitations localized in the apical chamber delineated by the flame cell body, suggesting that Sm GST 26 may be actively excreted by adult worms.     

Protective effect of rSm28GST-specific T cells in schistosomiasis: role of gamma interferon.

Immunization with a single dose of 50 micrograms of recombinant Schistosoma mansoni 28-kDa glutathione-S-transferase (rSm28GST) was able to induce a reduction in the worm burden, the number of eggs, and the degree of hepatic fibrosis as quantified by the measurement of collagen content in the liver of S. mansoni-infected mice. No relationship was found between anti-Sm28GST immunoglobulin G and immunoglobulin A titers and the levels of protection obtained. Adoptive transfers of Sm28GST-specific total, CD4+, or CD8+ T cells reproduced the protective effect obtained with the recombinant molecule. Moreover, experiments studying in vivo T-cell depletion demonstrated that anti-CD4- or anti-CD8-treated mice showed a significant decrease in the protective effect conferred, suggesting a role of the two T-cell subpopulations in the expression of Sm28GST-mediated protection against hepatic damage. Sm28GST-specific cells produced little interleukin-4 and high levels of gamma interferon. Treatment of immunized mice with anti-gamma interferon antibody totally suppressed the Sm28GST-induced protective effect and led to the rapid death of infected animals, suggesting a role for this cytokine in the expression of the protective immunity obtained after immunization with rSm28GST.     

A combined proteomic and immunologic approach for the analysis of Schistosoma mansoni cercariae and adult worm protein extracts and the detection of one of the vaccine candidates, Sm28GST, from a Venezuelan parasite isolate

Understanding the mode of Schistosoma mansoni larval invasion and the mechanism of immune evasion utilized by larvae and adult worms is essential for a rational development of vaccines or drugs to prevent or cure the disease. This parasite has a very complex molecular organization in all parasite stages, and identifying the major parasite proteins would give clues to schistosome metabolism and to the interaction of the parasite with the host immune system. Our goal was the evaluation of the protein parasite repertoire using a proteomic approach, and the characterization of protein extracts from two different parasite stages of a Venezuelan isolate, such as cercariae and adult worms, previously performed by other authors in some other strains. A comparison among authors was made. Besides, we aimed to identify different isoforms of one of the vaccine candidates, the gluthation-S-transferase protein (Sm28GST), by 2D SDS-PAGE and mass spectrometry, and to achieve its immunologic detection using sera from rabbits immunized with synthetic peptides derived from the Sm28GST protein. These techniques allowed the identification of some of the target molecules of the protective immune response that are being evaluated as potential members of a multi-component and multi-stage anti-S. mansoni vaccine and to clarify if the selected peptides induce antibodies that are able to recognize different isoforms of the Sm28GST.     

Expression of a 28-kilodalton glutathione S-transferase antigen of Schistosoma mansoni on the surface of filamentous phages and evaluation of its vaccine potential

A cloning and expression system that allows display of proteins on the surface of filamentous phages was exploited to display a 28-kDa glutathione S-transferase (Sm28GST) antigen of the human parasite Schistosoma mansoni. The phage-displayed Sm28GST (pdGST) was immunoreactive and was recognized by immune sera, suggesting that the Sm28GST protein displayed on the surface of phages potentially maintains native conformation. Subsequent immunization studies showed that mice can develop high titers of antibodies against pdGST and do not require any additional adjuvant for immunization. Isotype analysis suggested that the pdGST immunization predominantly induced immunoglobulin G2b (IgG2b), IgG3, and IgM anti-GST antibodies in mice. Furthermore, the pdGST immunization was found to confer about 30% protection after a challenge infection with 100 cercariae of S. mansoni in BALB/c mice. These findings suggest that phage display is a simple, efficient, and promising tool to express candidate vaccine antigens for immunization against infectious agents.     

GM-CSF and TNF-alpha synergize to increase in vitro granuloma size of PBMC from humans induced by Schistosoma mansoni recombinant 28-kDa GST

The 28-kDa Glutathione S-transferase of Schistosoma mansoni (Sm28 GST) was described as a protective antigen capable of reducing female fecundity and the number of eggs in mice hepatic tissues. The role of GM-CSF and TNF-alpha in the in vitro granuloma reaction of peripheral blood mononuclear cells (PBMC) from chronic intestinal schistosomiasis patients before and after chemotherapy treatment to S. mansoni recombinant Sm28 GST was evaluated. Treatment of PBMC with recombinant Sm28 GST caused a significant increase in granuloma formation when compared to SEA or SWAP. Contrary to SEA or SWAP, Sm28 GST was not capable of inducing significant cellular proliferation. Moreover, recombinant Sm28 GST promoted a significant elevation in GM-CSF and TNF-alpha levels. However, we did not detect any significant IL-10 production. When Sm28 GST was applied in the presence of anti-GM-CSF or anti-TNF-alpha antibodies in cultures, we observed a significant decrease in granuloma size. Indeed, our results demonstrated that Sm28 GST was capable of promoting high in vitro granuloma index, and this event was associated with the balance of GM-CSF and TNF-alpha. These evidences suggest a role for GM-CSF as a major mediator in increasing granuloma reaction in human schistosomiasis. This event may contribute to exacerbate the pathology resulting from egg deposition in host tissues.     

A monoclonal antibody blocking the Schistosoma mansoni 28-kDa glutathione S-transferase activity reduces female worm fecundity and egg viability

The protective effects of two different monoclonal antibodies (mAb) raised against the Schistosoma mansoni 28-kDa glutathione S-transferase (Sm 28 GST) were investigated. Two mAb of the same isotype (IgM) have been selected according to the blocking effect on Sm 28 GST enzymatic activity (S13) or the lack of blockade (H12). When passively transferred into Fischer rats, both S13 and H12 significantly reduced the worm burden. In BALB/c mice clear effects on female worm fecundity and egg viability were observed when the S13 mAb was transferred; these effects included significantly reduced loads of intestinal eggs, reduced egg hatching rates and an increased proportion of non-living eggs. No effect on egg production and egg hatching was observed in H12-treated mice. In addition, worm pairs recovered from S13-but not H12-treated mice laid significantly fewer eggs in vitro, and normal worm pairs incubated in vitro with the S13 mAb produced significantly fewer eggs than those incubated with H12 mAb. The impairment of egg hatching ability was also reproduced in vitro by the S13 mAb. These data suggest the existence of two different effector mechanisms induced by immunization with Sm 28 GST. The effect on the schistosome worm burden appears to be independent of GST activity whereas the effect on S. mansoni female fecundity and egg viability seems to be significantly linked to the inactivation of the enzymatic site.     

Schistosoma mansoni antigens differentially recognized by resistant WEHI 129/J mice.

Mice of the strain WEHI 129/J are genetically resistant to chronic Schistosoma mansoni infection. Resistance is expressed in at least 50% of mice, with the remaining mice showing normal susceptibility to infection. The serum antibody specificities in the resistant proportion of WEHI 129/J were analyzed at various times after exposure to cercariae by using both Western blotting and immunoprecipitation. Comparisons with the susceptible proportion of WEHI 129/J and other permissive mouse strains revealed four antigens that were differentially recognized by resistant mice at various times of infection: Sm25, an Mr 25,000 integral membrane protein of adult worms that was better recognized by resistant mice 40 to 50 days after exposure; Sm67, an Mr 67,000 water-soluble antigen of adult worms that was better recognized by resistant mice at days 30 to 40; Sm120, an Mr 120,000 antigen expressed by cercariae and adult worms that was differentially recognized, although inconsistently, at days 20 to 40 postexposure; and Sm26, an Mr 26,000 glutathione S-transferase that was uniquely recognized by resistant mice at day 20 in two of three experiments. Analysis of antibody specificities in (BALB/c x WEHI 129/J)F1 x WEHI 129/J backcross mice indicated that high responsiveness to Sm25 at days 40 to 50 correlated with resistance. The candidacy of these four molecules as vaccines for schistosomiasis mansoni is discussed.     

Bordetella pertussis Filamentous Hemagglutinin Enhances the Immunogenicity of Liposome-Delivered Antigen Administered Intranasally

In an attempt to increase the immunogenicity of mucosally delivered antigens, we incorporated the Bordetella pertussis filamentous hemagglutinin (FHA) adhesin into liposomes containing the glutathione S-transferase of Schistosoma mansoni (Sm28GST) as a model antigen. Outbred mice immunized twice intranasally with liposomes containing a constant suboptimal dose of Sm28GST and increasing doses of FHA produced anti-Sm28GST antibodies in a FHA dose-dependent manner. The addition of 3 μg of FHA to the liposomes induced more than 10-fold-higher anti-Sm28GST antibody titers, compared to those induced by liposomes without FHA. The presence of FHA did not alter the nature of the humoral immune response, and the sera contained anti-Sm28GST immunoglobulin G1 (IgG1), IgG2a, and IgG2b. However, anti-Sm28GST IgA was only detected when at least 3 μg of FHA was added to the preparation. These results show a promising potential for FHA to enhance the immunogenicity of mucosally administered antigens incorporated into liposomes.     

Inter-species variation of schistosome 28-kDa glutathione S-transferases.

The 28-kDa glutathione S-transferase from Schistosoma mansoni (Sm28GST) is a candidate vaccine antigen. To evaluate the antigenic and phylogenetic variations between the 28-kDa GSTs from 4 species of schistosome, we have cloned and sequenced the 28-kDa GSTs from Schistosoma haematobium (Sh28GST) and Schistosoma bovis (Sb28GST). Sb28GST and Sh28GST are more similar to each other (97%) than to Sm28GST (90%) and particularly to the 28-kDa GST from Schistosoma japonicum (Sj28GST, 77%). Antisera directed against the major Sm28GST epitopes revealed differences in the recognition of the 28-kDa GSTs from the other schistosome species suggesting that these regions have been subjected to evolutionary pressure. The consequences of such species-specific epitopes on the development of a multi-species anti-schistosome vaccine are discussed.     

Mice immunized with a chlamydial extract have no increase in early protective immunity despite increased inflammation following genital infection by the mouse pneumonitis agent of Chlamydia trachomatis.

We have determined that immunization with a detergent extract of the mouse pneumonitis agent of Chlamydia trachomatis fails to induce a protective inflammatory immune response following genital infection by C. trachomatis. We demonstrated that mice immunized with the detergent extract have increased cutaneous delayed-type hypersensitivity and increased splenic T-cell proliferation in response to the chlamydial extract. After genital infection by C. trachomatis, extract-sensitized mice had significantly increased genital inflammation (P = 0.044) compared with controls. The inflammation was characterized by significantly increased eosinophils in the genitalia (P < 0.0005) and increased genital edema (P < 0.0005). However, the increased genital inflammation of extract-sensitized mice provided no increase in protection against infection (P = 0.92).     

Immunization with Schistosoma mansoni 22.6 kDa antigen induces partial protection against experimental infection in a recombinant protein form but not as DNA vaccine

Schistosomiasis is a major public health problem that affects mainly developing countries. There are 200 million people worldwide infected with schistosomes resulting in more than 250,000 deaths per year. Although schistosomicidal drugs exist, the advent of an efficacious vaccine remains the most potentially powerful means for controlling this disease. In this study we isolated a cDNA clone encoding the Schistosoma mansoni lung-stage Sm22.6 protein, which is 100% and 79% identical with the 22.6 kDa adult worm tegument antigen of S. mansoni and S. japonicum, respectively. Further, we produced recombinant (r) Sm22.6 and constructed an Sm22.6 DNA vaccine. Western blot analysis confirmed the identity of purified MBP-Sm22.6 fusion protein using anti-MBP (maltose binding protein) and anti-rSm22.6 antibodies. Additionally, C57BL/6 mice were immunized and specific anti-Sm22.6 IgG responses were produced when both vaccination strategies were used. Importantly, only rSm22.6 vaccine provided levels of protection against challenge infection (34.5%). Mice immunized with rSm22.6 induced production of IgG1 and IgG2a and synthesis of IFN-gamma and IL-4 in cultured mouse splenocytes. Finally, rSm22.6 vaccination induced a Th0 type of immune response and protective immunity that suggests Sm22.6 as a potential candidate to compose an anti-schistosome vaccine.     

A Staphylococcus aureus capsular polysaccharide (CP) vaccine and CP-specific antibodies protect mice against bacterial challenge.

The efficacy of capsular polysaccharide (CP)-specific antibodies elicited by active immunization with vaccines composed of Staphylococcus aureus types 5 and 8 CP linked to Pseudomonas aeruginosa exoprotein A or with immune immunoglobulin G (I-IgG) obtained from vaccinated plasma donors was tested in lethal and sublethal bacterial mouse challenge models. A dose of 2 x 10(5) CFU of S. aureus type 5 CP per mouse administered intraperitoneally (i.p.) with 5% hog mucin was found to cause 80 to 100% mortality in BALB/c mice within 2 to 5 days. Mice passively immunized i.p. 24 h earlier or subcutaneously 48 h earlier with 0.5 ml of I-IgG showed significantly higher average survival rates than animals receiving standard IgG or saline (P < 0.01) following the bacterial challenge. Animals actively immunized with the monovalent type 5 CP-P. aeruginosa exoprotein A conjugate showed a survival rate of 73% compared with 13% in phosphate-buffered saline-immunized animals. The prechallenge geometric mean titer of type 5 CP antibodies in animals that died was significantly (P < 0.05) lower than that of animals which survived the challenge (95.7 versus 223.6 micrograms/ml, respectively). The IgG was further evaluated in mice challenged i.p. with a sublethal dose of 5 x 10(4) CFU per mouse. Serial blood counts were performed on surviving animals at 6, 12, 24, and 48 h. Surviving animals were sacrificed at 72 h, and bacterial counts were performed on their kidneys, livers, and peritoneal lavage fluids. Animals receiving I-IgG had lower bacterial counts in blood samples and lower bacterial densities in kidneys, livers, and peritoneal lavage samples than mice immunized with standard IgG (P < 0.05). These data suggest that S. aureus type 5 CP antibodies induced by active immunization or administered by passive immunization confer protection against S. aureus infections.     

Protection against Schistosoma mansoni achieved by immunization with sonicated parasite

Partial protection of (C57BL/6 X DBA/2)F1 mice against infection with Schistosoma mansoni has been achieved by immunization with small amounts (0.2 to 2.0 microgram) of crude cercarial sonicate adsorbed on aluminium hydroxide gel adjuvant (alum). A decrease of 34-90% in the adult worm burden, of the immunized mice, as compared to that of untreated mice or those injected with adjuvant alone, has been found in five experiments by liver perfusion six weeks after percutaneous challenge infection. High titers of anti-cercarial IgE antibodies have been found in the sera of the immunized mice by two independent techniques, radioimmunoassay and degranulation of rat basophilic leukemia cells (RBL-2H3), determined by 3H-serotonin release. By counting the live worms in the lungs of immunized and uniummized mice on days 4-7 after infection it was observed that the schistosomula were killed before they reached the lungs, probably at the skin. Mice immunized with the same amounts of cercarial antigen in Freund's complete adjuvant were not protected against infection with S. mansoni. These animals developed high titer of total anti-cercaria antibodies (determined by radioimmunoassay) but had low levels of antiparasite IgE. The possible role of IgE in protective immunity is discussed.     

Protective immunity against Salmonella typhimurium elicited in mice by oral vaccination with phosphorylcholine encapsulated in poly(DL-lactide-co-glycolide) microspheres.

Encapsulation of vaccines in biodegradable microspheres provides excellent mucosal immunogens with a high potential for immunization against bacterial infections. We tested the protective immunity elicited by intragastric vaccination with phosphorylcholine (PC) encapsulated in poly(DL-lactide-co-glycolide) (DL-PLG) microspheres against Salmonella typhimurium in a mouse model of invasive intestinal infection. We chose PC as the antigen because it was found to elicit an immune response after intestinal exposure of mice to PC-bearing S. typhimurium and because anti-PC immunity protects mice against Streptococcus pneumoniae, another PC-bearing microorganism. Mice were primed intragastrically on days 1, 2, and 3 and boosted on days 28, 29, and 30 with PC (280 microg) coupled to porcine thyroglobulin (PC-thyr) encapsulated in DL-PLG microspheres, free PC-thyr, or blank microspheres. A significant rise in anti-PC immunoglobulin A (IgA) titers, as measured by an enzyme-linked immunosorbent assay, was observed in the intestinal secretions after immunization with PC-loaded microspheres, compared to the titers of mice immunized with free PC-thyr or blank microspheres. This antibody response peaked 14 days after the last boost and correlated with a highly significant resistance to oral challenge by S. typhimurium C5 (P < 10(-3)). Control mice were primed intraperitoneally on day 1 with 15 microg of PC in complete Freund's adjuvant and boosted on days 10, 14, and 20 with the same dose without adjuvant but via the same route. In these mice, the levels of anti-PC IgA in intestinal secretions were equivalent to those of the mice intragastrically immunized with PC-loaded microspheres, but protection was significantly weaker, suggesting that either the IgAs were not functional or that other immune mechanisms are important in protection. Taken together, our results highlight the potential of antigen encapsulation in DL-PLG microspheres for eliciting protective immunity against invasive intestinal bacterial diseases and suggest that a similar strategy could be used against diseases caused by other PC-bearing microorganisms.     

Immunization with an acellular vaccine consisting of the outer membrane complex of Chlamydia trachomatis induces protection against a genital challenge.

The ability to induce protection against a genital challenge was studied in BALB/c female mice with three Chlamydia trachomatis mouse pneumonitis (MoPn) major outer membrane protein (MOMP) preparations as well as an acellular vaccine consisting of the chlamydial outer membrane complex (COMC). The MOMP preparations were extracted with three different types of detergents, sodium dodecyl sulfate (SDS), n-octyl-beta-D-glucopyranoside (OGP), and Zwittergent 3-14 (Z3-14). A positive immunization control consisted of mice inoculated intranasally with 10(4) C. trachomatis MoPn inclusion-forming units (IFU). Mice inoculated with ovalbumin served as a negative control. Furthermore, a sham-immunized, nonchallenged group was included as a fertility control. Two weeks after the last immunization, the mice were challenged in the left ovarian bursa with 10(5) C. trachomatis MoPn IFU. Vaginal swabs were collected for culture, vaginal and serum samples were assayed for chlamydial-specific antibodies, and splenocytes were collected to determine the lymphoproliferative response. At 42 days after the challenge, the mice were mated with proven male breeder mice. Animals that were considered to be pregnant (as determined by weight) were killed, and the embryos were counted. A significant humoral and cell-mediated immune response was observed in all the groups of mice inoculated with chlamydial antigens. Antibodies to variable domain (VD)1 of the MOMP were detected in serum samples from all the immunized groups. However, antibodies to VD3 and VD4 were detected only in the groups immunized with the Z3-14-MOMP and the COMC. Mice immunized with COMC developed significant immunoglobulin A chlamydia-specific antibodies in the vagina, while mice immunized with the detergent-extracted MOMPs had low antibody titers. Following the intrabursal challenge, a significant decrease in the intensity and duration of vaginal shedding was noted in the mice immunized with COMC and a moderate decrease was noted in the group immunized with OGP-MOMP. No protection against the infection was noted in the groups of animals immunized with SDS- and Z3-14-MOMP. Furthermore, of the mice immunized with the COMC preparation, only 25% (4 of 20) shed C. trachomatis, as determined by vaginal culture, while 83% (40 of 48) of the control mice inoculated with ovalbumin were culture positive (P < 0.05). In addition, after mating, the mice inoculated with COMC were found to have fertility rates comparable to those of the control sham-immunized, nonchallenged animals (70% [14 of 20] versus 81% [17 of 21], respectively [P > 0.05]), and there were no significant differences between the average number of embryos per mouse in the two groups (5.1 versus 5.9, respectively [P > 0.05]). In contrast, mice immunized with the purified MOMP preparations were not protected against infertility. In summary, a preparation of the COMC protected mice against infection and infertility, supporting the feasibility of the development of an acellular vaccine against C. trachomatis infections.     

Obtention of a human primary humoral response against schistosome protective antigens in severe combined immunodeficiency mice after the transfer of human peripheral blood mononuclear cells

Peripheral blood mononuclear cells (PBMC) from healthy donors were injected into C.B.-17 severe combined immunodeficiency (scid) mice which were subsequently immunized with crude Schistosoma mansoni adult worm antigenic preparation (SWAP) or with recombinant S. mansoni 28-kDa glutathione transferase (r-Sm-28-GST) antigen. PBMC from a S. mansoni-infected patient were also transferred. The specific human anti-SWAP and anti-Sm-28-GST antibody responses were monitored. The presence in both cases of human specific antibodies in scid mouse sera was determined by enzyme-linked immunosorbent assay and Western blotting techniques using anti-human immunoglobulin reagents. No antibodies were detected in these sera using anti-mouse immunoglobulin antisera. These antibodies were functional since a cytotoxic activity against schistosomula was observed when monocytes were incubated with scid mouse sera positive for anti-Sm-28-GST antibodies.     

T-cell responsiveness towards various synthetic peptides of the P28 antigen in rat and mouse models during Schistosoma mansoni infection.

It has recently been demonstrated that the Schistosoma mansoni P28 antigen can induce a strong protective immunity after direct immunization in various experimental models. T lymphocytes from Fischer rats immunized with the recombinant P28 antigen were cultured in vitro in the presence of seven synthetic peptides derived from the amino acid sequence of the P28. The most significant and reproducible proliferation was obtained with the 24-43 and 115-131 synthetic peptides. In order to analyze whether these located determinants were also exposed to the host's immune system during the natural S. mansoni infection or after immunization with crude antigenic extracts of various development stages of the parasite, the T-cell responsiveness of infected or immunized Fischer rats and BALB/c mice was tested towards these synthetic peptides. The results showed that, in both permissive (mouse) and non-permissive (rat) hosts, 24-43 and 115-131 synthetic peptides are recognized during the course of infection and that there is a dynamic variation of this recognition. These peptides are also recognized by T cells educated against crude antigenic extracts of different developmental stages of the parasite which contained the native form of the P28 molecule. Taken together, the results indicated that these synthetic peptides derived from the recombinant P28 antigen can activate T lymphocytes educated against the native P28 molecule during the development and maturation of the parasite in their hosts. Therefore, they might be useful for the construction of synthetic vaccines against schistosomiasis.     

Expression of a 28-Kilodalton Glutathione S-Transferase Antigen of Schistosoma mansoni on the Surface of Filamentous Phages and Evaluation of Its Vaccine Potential

A cloning and expression system that allows display of proteins on the surface of filamentous phages was exploited to display a 28-kDa glutathione S-transferase (Sm28GST) antigen of the human parasite Schistosoma mansoni. The phage-displayed Sm28GST (pdGST) was immunoreactive and was recognized by immune sera, suggesting that the Sm28GST protein displayed on the surface of phages potentially maintains native conformation. Subsequent immunization studies showed that mice can develop high titers of antibodies against pdGST and do not require any additional adjuvant for immunization. Isotype analysis suggested that the pdGST immunization predominantly induced immunoglobulin G2b (IgG2b), IgG3, and IgM anti-GST antibodies in mice. Furthermore, the pdGST immunization was found to confer about 30% protection after a challenge infection with 100 cercariae of S. mansoni in BALB/c mice. These findings suggest that phage display is a simple, efficient, and promising tool to express candidate vaccine antigens for immunization against infectious agents.