Use of larval, parasitic female and egg antigens from Strongyloides venezuelensis to detect parasite-specific IgG and immune complexes in immunodiagnosis of human strongyloidiasis

The aim of this study was to use larval, parasitic female and egg antigens from Strongyloides venezuelensis to detect parasite-specific IgG and immune complexes in human serum samples by enzyme-linked immunosorbent assay (ELISA). In total, 95 serum samples were analysed, consisting of 30 patients harbouring S. stercoralis larvae, 30 healthy subjects and 35 patients with other parasites. Sensitivity, specificity and diagnostic efficiency were calculated. A significant statistical difference was found in the detection of immune complexes and antibodies in patients harbouring S. stercoralis larvae from larval and eggs antigens, with higher positivity using larval antigen. The larval antigen showed the highest values for sensitivity, specificity and diagnostic efficiency in ELISA from detection of immune complexes. For the first time we used IgG anti-larvae, IgG anti-parasitic females or IgG anti-eggs for immune complex detection. We concluded that the association of antibody and immune complex detection could be used in the diagnosis of human strongyloidiasis.     

Filarial-specific IgG4 response correlates with active Wuchereria bancrofti infection

The filarial-specific humoral immune response of adult residents of two areas of Papua New Guinea, differing in transmission of Wuchereria bancrofti infection was compared. The majority of residents of the village of Bonahoi, in an area where transmission of filariasis had been interrupted by a 20-year insecticide spray program to control malaria, showed no parasitologic signs of active W. bancrofti infection and were negative for both circulating phosphorylcholine Ag and peripheral blood microfilariae. In contrast, adult residents of the village of Nanaha were in an area exposed to infection, and were phosphorylcholine-Ag- and microfilariae-positive. The antibody response of these two groups to both adult worm excretory/secretory (ES) Ag and somatic antigen extract was examined to determine which components of the filarial-specific immune response were dependent on active infection. Identification of these immune responses may point to immunologic methods to evaluate control programs for lymphatic filariasis. Adults from Bonahoi were found to have significant immune responses to [35S] methionine-labeled ES Ag by immunoprecipitation and to adult somatic antigen extracts by ELISA and by immunoblotting. This result is consistent with the fact that these individuals were previously exposed to and/or infected with W. bancrofti. Similarly, residents of the endemic village had detectable immune responses to these Ag irrespective of if they were microfilaremic. The most striking immunologic difference observed between the two groups was that residents of Bonahoi had a dramatically reduced filarial-specific IgG4 antibody response to both adult somatic Ag and adult ES Ag. These data suggest that longitudinal measurement of filarial-specific IgG4 levels may be a useful seroepidemiologic indicator of changes in W. bancrofti infection status.     

Separation and characterization of adult worm proteins and glycoproteins from the liver fluke Opisthorchis viverrini.

Detailed studies of liver fluke proteins and antigens are necessary to facilitate further investigation of the human immune responses to these parasites. Accordingly, Opisthorchis viverrini antigens were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. We initially encountered excessive background smearing, vertical streaking, and indistinct bands that were similar to problems previously described by investigators studying this and other trematodes including Schistosoma mansoni. These problems were especially evident with silver staining of proteins and occurred despite the extensive use of protease inhibitors. They were minimized by using mini (vs. large) SDS-PAGE and Coomassie blue protein staining. With the latter 2 techniques, adult worm somatic proteins and excretory-secretory products were separated and characterized. Immunoblots using rabbit anti-adult worm sera demonstrated that some of these proteins were antigens common to both the adult and metacercarial stages. Several of these antigens also corresponded (according to molecular weight) to glycoproteins, detected by concanavalin A blotting. These findings form a base for subsequent studies of the human immune response to liver fluke infection.     

Characterization of antigens of the nematode Nippostrongylus brasiliensis by monoclonal antibodies

Hybridomas secreting monoclonal antibodies to Nippostrongylus brasiliensis antigens were generated by hybridization of IR983F myeloma cells with spleen cells from Lou/M/Wol rats infected with living third-stage larvae. Antibodies specific either for larval or worm antigens were identified by enzyme-linked immunosorbent assays with Nippostrongylus brasiliensis fragments, homogenates and secretions as antigens. The results demonstrate that all antibodies which recognized larval antigens (38 antibodies) also reacted with worm surfaces. Ten antibodies were specific only for worm antigens. Ten antibodies reacted with worm homogenate, three antibodies recognized components of worm secretion and 17 antibodies combined with acetylcholinesterase. The epitope specificity was investigated by the capacity of various glycosides, aminoacids, N-acetylneuraminic acid and phosphorylcholine to inhibit the binding to worm fragments. The analysis revealed that alpha-methylglucoside, alpha-methylmannoside, N-acetylglucosamine, N-acetylgalactosamine, fucose and the amino acids leucine, phenylalanine, tyrosine, serine, tryptophan did not combine with the antigen-binding sites of the antibodies. Proline, arginine and histidine, however, displayed inhibitory effects. With N-acetylneuraminic acid as inhibitor three groups of antibodies could be discriminated. At a concentration of 10-20 mM, phosphorylcholine was a potent inhibitor for all antibodies.     

Brugia malayi: detection of parasite antigen in sera from infected jirds

Sera from Brugia malayi-infected jirds were demonstrated to contain a heat-stable, 95- to 105-kDa parasite antigen by immunoblot with rabbit antibody to the parasite and with a monoclonal antibody that binds to phosphorylcholine. This antigen is a major component of B. malayi adult worm excretory/secretory antigen, and it is present in lavage fluid obtained from ip-infected animals. The antigen was detected by enzyme immunoassay in all sera collected from jirds 9-54 weeks after sc injection with 100 or 300 infective larvae (L3). Parasite antigen titers were higher in animals infected with the higher L3 dose. Antiphosphorylcholine antibodies were present in jird sera for the first 12 weeks after larval injection, but thereafter, antibody titers decreased to undetectable levels. Parasite antigen was not detected by immunoblot or enzyme immunoassay in sera from 21 human subjects with B. malayi microfilaremia. Antigen may be cleared from human sera by antiphosphorylcholine antibodies, which were present in all sera tested. The practical significance of B. malayi antigen detection in the jird is that it provides a sensitive means of noninvasively monitoring the status of infection in this important experimental filariasis model.     

Nippostrongylus brasiliensis: local humoral responses in the lungs and intestines of rats following vaccination with irradiated larvae

Groups of rats were infected with 2000 normal larvae of Nippostrongylus brasiliensis or larvae irradiated with 10 to 120 kR. On Day 10 after infection half the animals from each group were autopsied. The remainder were challenged with 5000 unirradiated larvae on Day 15 and killed ten days later. During the experiment enteric antibody levels were estimated by coproantibody measurement. At autopsy the worm burdens were determined and worm-specific antibodies evaluated in lung extracts and serum. It was found that the levels of coproantibody detected with adult worm metabolites were positively correlated with the number of adult nematodes recovered from the intestine after primary infection. The challenge induced a similar increase of these antibodies in all immunised rats which reflected a high immunity to reinfection of vaccinated animals. Preliminary immunochemical studies suggested that the coproantibodies had SIgA properties. In lung extracts of rats immunised with larvae irradiated at 40, 80, or 120 kR and in all animals after challenge, antibodies reacting with infective larval antigens were found. Their titres were negatively correlated with serum antibody levels. The significance of bronchial and enteric antibodies in conferring protection against challenge remains to be elucidated.     

Hemoglobin switching in the salamanderPleurodeles waltlii

Summary In order to study the cellular distribution of larval and adult hemoglobins during larval development ofPleurodeles waltlii a double specific immunofluorescent labelling technique was developed.Rabbit antibodies specific for larval and adult hemoglobin components were prepared and conjugated with tetramethyl-rhodamine isothiocyanate for the anti-larval antibodies and fluorescein isothiocyanate for the anti-adult hemoglobin antibodies.Both simultaneous and sequential staining with the two types of fluorescent antibodies indicated that larval and adult hemoglobins were never observed within the same erythrocyte during development. The results provide evidence that two distinct cell populations exist, one synthesizing exclusively larval hemoglobins which is progressively replaced by the other one synthesizing exclusively adult hemoglobins. It remains to be determined if these two populations arise from two distinct types of stem cells (adult and larval) or from the same stem cell type.     

Trichinella spiralis: antifecundity and antinewborn larvae immunity in swine

The development of antifecundity and antinewborn larvae immunity in swine infected with Trichinella spiralis was investigated. In primary infections, adult female worm fecundity dropped sharply after 3 weeks, although adults could be recovered from the small intestine for at least 7 weeks after infection. In challenge infections of pigs infected previously, adult female worm fecundity was depressed up to 51% and the adults were expelled within 3 weeks. Since immune pigs are almost completely resistant to the secondary establishment of muscle larvae, this suggested the existence of immune effector mechanisms also acting on the newborn larvae. This was supported by observations, using an indirect fluorescent antibody assay, that pig antibody bound to the surface of the newborn larvae. Passive transfer of immune pig serum resulted in a large reduction in muscle larvae burden in both infected pig and rat recipients. Adult female worm fecundity in such immune serum recipients was reduced only by 20% and worm survival in the intestine was unaffected. These results indicate that immunity to the newborn larvae, in addition to antifecundity effects, are responsible for the high levels of acquired resistance to T. spiralis in swine.     

Detection of antibodies to Angiostrongylus cantonensis in serum and cerebrospinal fluid of patients with eosinophilic meningitis

Adult and young adult antigens of Angiostrongylus cantonensis were purified by immuno-affinity chromatography and used to detect antibody in serum and cerebrospinal fluid (CSF), by enzyme-linked immunosorbent assay (ELISA), in cases of human eosinophilic meningitis or meningoencephalitis. The levels of IgG, IgA, IgM and IgE antibodies to A. cantonensis in these patients were higher than levels in control subjects. Antibodies in patients detected against adult and young adult worm antigens of A. cantonensis did not differ significantly. Significantly higher IgM and IgE antibody levels were observed in serum compared with CSF from infected patients (Student's t-test, P less than 0.05). Both adult and young adult A. cantonensis antigens proved to be highly sensitive in ELISA for serum antibodies; however, the sensitivity was significantly lower in tests on CSF.     

Circulating antigens and antibodies in human and mouse dermatophytosis: use of monoclonal antibody reactive to phosphorylcholine-like epitopes

The presence of circulating antibodies in the sera of patients infected with either Trichophyton concentricum or Trichophyton rubrum, and in the sera of BALB/c mice chronically infected with Trichophyton quinckeanum, was determined by ELISA. High levels of antibody to dermatophyte cytoplasmic antigens were detected both in infected humans and in mice. Partial inhibition of this reaction was observed by pretreatment of the sera with the hapten phosphorylcholine (PC). Moreover, antibodies were shown to have some reactivity with PC when tested by ELISA against PC conjugated to bovine serum albumin. Significant levels of circulating antigen were detected in patients with T. concentricum and T. rubrum infections, but not in uninfected subjects, by an immunoradiometric assay using a monoclonal antibody, Tq-1, which reacts with the PC-like epitopes of dermatophytes. It is possible that this dermatophyte antigen may play a role in modulating the cell-mediated immune responses, which would appear to be defective in most patients with these chronic forms of dermatophytosis.     

Brugia pahangi: effects of maternal filariasis on the responses of their progeny to homologous challenge infection.

Granulomatous lesion formation and immune responses to Brugia pahangi infections were compared in age-matched male progeny of homologously infected and uninfected female jirds. Infections initiated in 2-week-old offspring yielded mean +/- SD adult worm recoveries of 6.0 +/- 5.7 and 4.2 +/- 5.4 in offspring from infected or uninfected mothers, respectively. Infections initiated in 4-week-old offspring resulted in an mean +/- SD recovery of adult worms of 11.3 +/- 11.3 and 10.2 +/- 5.8 in offspring from infected and uninfected mothers, respectively. The ratio of intralymphatic thrombi per intralymphatic worm was similar between infected offspring from infected or uninfected mothers within experiments. Areas of granulomas around B. pahangi antigen-coated beads embolized in the lungs were not significantly affected by maternal origin in infected or uninfected progeny. Offspring infected at 2 or 4 weeks of age from infected mothers exhibited significantly reduced titers of serum IgG antibodies to Brugia antigens at 5-8 weeks postinfection compared to infected offspring of uninfected mothers. Infected offspring from infected mothers also had significantly fewer splenic IgG plaque-forming cells to B. pahangi antigens at 5 weeks postinfection than similarly infected offspring from uninfected mothers. Western immunoblot analysis indicated qualitative and quantitative reductions in serum antibody reactivity to adult B. pahangi antigens in infected progeny of infected females compared to age-matched infected controls. Reduced homologous serum antibody responses in progeny exposed to maternal B. pahangi infection suggest that maternal immunoregulation to filarial antigens may occur. Reduced antibody responsiveness to B. pahangi antigens observed in infected offspring from infected mothers, however, had no demonstrable effect on adult worm burdens, microfilaremias, lymphatic lesion formation, or antigen-specific granulomatous inflammatory responses compared to infected progeny of uninfected mothers.     

The cellular and humoral immune response to Schistosoma mansoni infections in inbred rats: I. Mechanisms during initial exposure

Fischer rats were infected with Schistosoma mansoni by exposure to cercariae. The effects of animal age at exposure and the size of that exposure were determined by quantitation of subsequent worm burdens. Optimal conditions for assay of protection mediated by cells or serum transferred from exposed donors were ascertained. The major initial protective immune response to infection was mediated by thymusdependent lymphocytes as demonstrated by discrete subpopulation cell transfer experiments. Simultaneously harvested serum enhanced worm survival and obviated the protective effects of transferred cells when serum and cells were given concomitantly. Subsquently, host immunity became dependent on antibody formation. In the chronic phase of infection, a small number of surviving parasites coexisted with antibody which specifically interacted with schistosomal antigens but did not protect in transfer experiments. These studies suggest a form of immunologic enhancement with antibody modulation of cellular immunity as possible components of the immune response to both acute and chronic schistosomiasis.     

Inhibition phenomena mechanisms in experimental obeliscoidosis in rabbits. II. Local and systemic antibody responses

A series of experiments was carried out using adult outbred Polish race rabbits of both sexes infected, during spring or autumn, with 10,000 larvae of Obeliscoides cuniculi, either fresh or stored at 4 degrees C. Extracts of mucosal proteins and bile were collected at postmortem 6 or 12 weeks after infection. Antibody levels were determined in antisera, bile and stomach mucosa by haemagglutination and precipitation tests. Local antibody responses were demonstrated in the stomach and bile, and reactions were obtained with the tissue fluids by haemagglutination and precipitation tests with worm antigens and ES products. Additionally, some specific immunological response was observed in the circulation during the primary infection. These results suggest a clear-cut relationship between increased levels of these antibodies and either larval inhibition or worm expulsion during O. cuniculi infections.     

Immune changes of Schistosoma japonicum infections in various rodent disease models

Rodent models for Schistosoma japonicum infections have demonstrated that these animals possess a degree of resistance to schistosome infections that may be both T and B lymphocyte-mediated. However, their exact role is not well-defined and other immune mechanisms are likely to also play a role in protecting against infection. Immunosuppressed and immunocompetent reed voles (Microtus fortis, Mf), rats and mice (n=24/group) were infected with S. japonicum, and animals were sacrificed 42 days later under anesthesia. Neither worms nor eggs were observed in infected immunosuppressed Mf or rats, with the exception of one rat that presented with few eggs. In immunosuppressed mice, changes in the number and size of the worms were not significantly different compared to immunocompetent mice, but worm fecundity was affected. The size and number of granulomas in immunosuppressed animals was also reduced. Analysis of serum antibodies specific to schistosome adult worm antigen at 3 weeks post-infection demonstrated that the levels of antibodies in the sera of rats were significantly higher than in Mf and mice. In addition, Mf serum levels of IL-4 and IL-12 were significantly higher than levels observed in rats and mice. Antibodies and cytokines in the sera of Mf peaked 3 weeks post-infection and then began to decrease, while antibody responses in rats and mice increased gradually between weeks 3-7 post-infection. It is possible that T and B cells have a dual role in both mediating protection and exacerbating disease outcomes.     

Inhibition of specific antibody binding to adult male Schistosoma mansoni by adsorbed host serum components

The ability of anti-schistosome antibody to bind to adult male Schistosoma mansoni was studied, using fluoresceinated Staphylococcus aureus to detect specific antigen-antibody interaction at the parasite's surface. Both freshly perfused parasites and parasites which had had their adsorbed host antigens removed by elution were employed in a series of experimental manipulations to ascertain under what conditions specific antibody binding occurs and what conditions or factors are necessary for the parasite to reconstitute its surface so that specific binding is precluded. Neither normal mouse serum nor normal mouse IgG bound in a specific manner to either fresh or eluted worms. Slight binding was noted with immune mouse serum on both fresh and eluted worms, while immune IgG produced weak binding on fresh worms, but strong binding to eluted worms. This strong binding was reduced to the level seen on fresh worms by pre-incubation of the eluted worm in normal IgG prior to incubation in immune IgG and binding was completely negated by pre-incubation in either normal mouse serum or normal mouse serum minus IgG. The binding of immune IgG to eluted worms was not diminished by pre-incubation in mouse albumin, bovine albumin, or fetal bovine serum. These studies demonstrate that adsorbed host serum components can inhibit specific antigen-antibody interaction at the parasite's surface and suggest that a degree of specificity exists in what the parasite adsorbs from the host. These data further suggest that the protective serum factor or factors may include, but are not limited to, host IgG.     

Trichinella spiralis: light microscope monoclonal antibody localization and immunochemical characterization of phosphorylcholine and other antigens in the muscle larva

A panel of monoclonal antibodies was used to examine the structure of the muscle larva of Trichinella spiralis under the light microscope. Immunofluorescence and, in some cases, immunoperoxidase staining were used. All four antibodies reacted with the cuticle of the organism, although differences in the staining pattern were observed for some of these. Interestingly, all the antibodies also reacted with the stichosome. One of the antibodies (Ts2Ab) is specific for the hapten, phosphorylcholine. In a binding assay, this antibody also reacted with extracts of Trichuris suis, Ascaris suum, and Fasciolopsis buski, but not with extracts derived from Cysticercus cellulosae, Candida albicans, Salmonella typhi, or Escherichia coli. This crossreactivity was confirmed microscopically in which the cuticle, oviduct and eggs of T. suis, the cuticle, muscle cells, and eggs of A. suum, and the cuticle and vitelline glands of F. buski were seen to be clearly stained by the antibody. In addition, Ts2Ab also reacted with the cuticle and stichosome of the adult T. spiralis worm. In Western blot analysis, Ts2Ab recognized a 43-kDa antigen from T. spiralis muscle larvae extracts, while a previously studied antibody (7C2C5Ab) identified four major antigens (48.5, 47, 43, and 39 kDa) in this preparation. Similar results were obtained when the 24-hr excretory-secretory (ES) antigens of T. spiralis were immunoblotted with the antibodies, although the reactivity shown by Ts2Ab was relatively weak. With the 72-hr ES material, on the other hand, major antigens of lower mol wt (44, 28, and 25 kDa) were revealed by 7C2C5Ab, and no reactivity was seen with Ts2Ab. However, this antigen preparation reacted well with both antibodies in an enzyme-linked immunoassay. Taken together, the findings suggest that the 72-hr ES antigens probably result from extensive degradation of material originally secreted or excreted by the worm. Similar binding studies on the 24-hr ES preparation indicated that this source may be relatively rich in 7C2C5Ab-reactive epitopes and relatively poor in the antigen identified by Ts2Ab. Other studies performed demonstrated that the antigens recognized by these two antibodies were distinct and physically unassociated.     

Specific suppression of the antibody response in vitro by serum from paralyzed mice.

BALB/c mice immunized with either the whole vaccine or the C-polysaccharide obtained from the R36A strain of pneumococcus produce antibody to phosphorylcholine. Mice injected i.v. with a single high dose of the C-polysaccharide are specifically unresponsive to immunization to phosphorylcholine for many months and are considered paralyzed. The induction of paralysis does not eliminate cells reactive to phosphorylcholine; however, serum from paralyzed mice specifically suppresses the response of cultures of normal spleen cells to phosphorylcholine. Paralyzed mice have an early low antibody response to phosphorylcholine and to the receptor for phosphorylcholine as indicated by plaque-forming cell assays. The factor or factors present in serum which may suppress cultures, and, by presumption, be responsible for paralysis are complexes of antigen, antibody, and antibody to the receptor for phosphorylcholine.     

A study of auto-anti-idiotypes to BSA

In order to study the idiotypic relationships between the antibody populations produced in different species during normal immune responses to ordinary protein Ag, we raised immune sera in mice and chickens by using three protein Ag: BSA, keyhole limpet hemocyanin, and diphtheria toxoid. An avidin-biotin ELISA was used to measure idiotypic binding between antibody populations from these sera. We found that the chicken sera contained auto-anti-idiotypes (AAI) against Ag-specific antibodies that were present in the same serum and that co-purified with those antibodies on Ag-Sepharose columns. These AAI were present in secondary response chicken anti-BSA serum at levels comparable with those of the anti-BSA antibody. The chicken AAI also react specifically with Id in mouse anti-BSA serum. The mouse anti-BSA serum completely inhibits the binding between the chicken Id and AAI. This similarity between the Id of whole populations of antibodies produced in two distantly related species, in the absence of any manipulation with idiotypic or anti-idiotypic reagents, suggests that the AAI detected in this way are internal image antibodies. It indicates there is positive selection for such AAI to be internal images.     

Detection of circulating immune complexes by the enzyme-linked immunosorbent assay in sera from cattle infected with Fasciola hepatica

Polyethylene glycol-precipitated immune complexes (PIC) from the sera of 5 calves with Fasciola hepatica worm burdens ranging between 27 and 70 flukes were examined for parasite antigen content at 2, 4, 6, 8, 10, and 16 wk postinfection (PI) by the enzyme-linked immunosorbent assay (ELISA). Three assays were devised using an affinity-processed rabbit antibody to worm excretory/secretory (FhES) antigens. The PIC plate assay detected parasite antigen by adherence of anti-FhES antibody to PIC incubated overnight on ELISA plates, and tests were visualized using anti-rabbit peroxidase-linked antibody. The serum complex and PIC capture assay utilized the anti-FhES immunoglobulin as an antigen capture antibody linked to the solid phase. The attached complexes were then detected by the adhering bovine antibody, either soluble complexes in serum or as PIC. All assays showed circulating immune complex (CIC) values elevated at 6-8 wk PI, which generally coincided with increased host circulating antibody to FhES antigens. The greatest detection rate for all of the immune complex (IC) detection assays occurred with the PIC capture assay. It detected antigen in almost 90% of sera tested at 6 and 8 wk PI. Both the serum complex and PIC capture assay detected greatest amounts of CIC in those animals with the largest worm burdens, whereas the PIC plate assay showed no such trend. This study shows that F. hepatica antigen detection in CIC can be used to aid immunologic diagnosis of fascioliasis.     

Macrophages as effector cells of protective immunity in murine schistosomiasis. VI. T cell-dependent, lymphokine-mediated, activation of macrophages in response to Schistosoma mansoni antigens

Macrophages from Schistosoma mansoni-infected mice kill significant numbers of skin stage schistosomula and murine fibrosarcoma cells in vitro. In order to determine whether the macrophage tumoricidal and larvicidal activation observed in mice as a result of S. mansoni infection are mediated through T cell-dependent (lymphokine) or B cell-dependent (antibody or immune complex) mechanisms, the development of macrophage populations with cytotoxic activity against schistosome larvae or tumor cells was monitored in S. mansoni-infected nude or mu-suppressed mice. Whereas peritoneal cells from S. mansoni-infected congenitally athymic mice had no activity in either assay, cells from mu-suppressed S. mansoni-infected mice showed cytotoxic activity equivalent to that of cells from untreated S. mansoni-infected counterparts. Cells from mu-suppressed uninfected mice were not activated. The mu-suppressed animals had no detectable nonspecific IgM or specific antischistosome IgM, IgG, or IgE antibodies and showed a 90% reduction in numbers of splenic IgM+ cells upon fluorescence activated cell sorter analysis. These results indicate that antibody is not required for in vivo activation of macrophages during S. mansoni infection. Further experiments showed that lymphoid cells from S. mansoni infected mice respond in culture with various specific antigens (such as living or dead whole schistosomula or soluble adult worm antigens) by production of factors capable of activating macrophages from uninfected control mice to kill schistosomula or tumor cells in vitro. Macrophage-activating factors were produced by T cell-enriched, but not T cell-depleted or B cell-enriched, populations from spleens of schistosome-infected mice in response to schistosome antigen. Similar lymphokines may be responsible for the macrophage activation observed during chronic murine schistosomiasis. These observations emphasize the potential contribution of T cell-mediated immune mechanisms in resistance to S. mansoni infection.