Comparative evaluation of Schistosoma mansoni, Schistosoma intercalatum, and Schistosoma haematobium alkaline phosphatase antigenicity by the alkaline phosphatase immunoassay (APIA).

To know if alkaline phosphatase (AP) from schistosomes other than Schistosoma mansoni can be used as diagnostic marker for schistosomiasis in alkaline phosphatase immunocapture assay (APIA), we comparatively tested n-butanol extracts of adult worm membranes from a Venezuelan (JL) strain of S. mansoni (Ven/AWBE/Sm); a Cameroonian (EDEN) strain of Schistosoma intercalatum (Cam/AWBE/Si) and a Yemeni strain of Schistosoma haematobium (Yem/AWBE/Sh). APIA was evaluated with sera of patients from Venezuela, Senegal, and Gabon infected with S. mansoni, from Gabon infected with S. intercalatum or S. haematobium, from Chine infected with Schistosoma japonicum and from Cambodian patients infected with Schistosoma mekongi. Results indicate that 92.5% (37/40) of Venezuela sera, 75% (15/20) of Senegal sera, 39.5% (17/43) of S. haematobium sera, and 19.2% (5/26) S. intercalatum sera were APIA-positive with the Ven/AWBE/Sm preparation. APIA with the Cam/AWBE/Si preparation showed that 53.8% of S. intercalatum-positive sera had anti-AP antibodies, and 51.2% S. haematobium-positive sera cross-immunocapturing the S. intercalatum AP. APIA performed with Yem/AWBE/Sh showed that 55.8% S. haematobium sera were positive. Only two out of nine S. japonicum sera were APIA-positive with the Ven/AWBE/Sm and Cam/AWBE/Si, and no reaction was observed with Cambodian S. mekongi-positive sera. AP activity was shown to be present in all the schistosome species/strains studied. The use of APIA as a tool to explore the APs antigenicity and the presence of Schistosoma sp. infections through the detection of anti-Schistosoma sp. AP antibodies in a host, allowed us to demonstrate the antigenicity of APs of S. mansoni, S. intercalatum, and S. haematobium.

Detection of the Sm31 antigen in sera of Schistosoma mansoni- infected patients from a low endemic area.

Schistosoma mansoni cathepsin B (Sm31) is a major antigen from adult worms that circulates in the blood of infected patients (Li et al., Parasitol Res 1996; 82: 14-18). An analysis of the Sm31 sequence (Klinkert et al., Mol Biochem Parasitol 1989; 33: 113-122) allowed the prediction of seven hydrophilic regions that were confirmed to be exposed on the surface of a 3D model of Sm31; the species specificity of these regions was checked using BLAST analysis. The corresponding peptides were chemically synthesized in polymerazible forms using the t-Boc technique. Rabbits developed a high humoral response against these peptides as tested by a multiple antigen blot assay; it recognized native Sm31 in crude S. mansoni extracts and as circulating antigen in sera of S. mansoni-infected patients by western blot. Relevant antigenic determinants were located at the N- and C-terminus sequences. Antibodies against these regions recognized the native enzyme in an ELISA-like assay called cysteine protease immuno assay in which the immunocaptured enzyme was revealed by the intrinsic cathepsin B hydrolytic activity of Sm31. The method successfully and specifically detected Sm31 in sera of infected individuals, most of them (83.3%) with light infections, offering a rationale for the development of parasite enzyme capture assays using anti-synthetic peptide antibodies for possible use in the diagnosis of schistoso,iasis.

Detection of schistosomiasis cases in low-transmission areas based on coprologic and serologic criteria The Venezuelan experience.

Low and very-low intensities of infection hinder the diagnosis of schistosomiasis. Therefore, new parameters should be established in order to more accurately identify active cases and true infection prevalence, for the adequate implementation of a control program. After the survey and analysis of the epidemiological characteristics of five Venezuelan communities, we propose three criteria for the definition of a "schistosomiasis case", based on different diagnostic methods: stool examination, ELISA-soluble egg antigen with sodium metaperiodate (SMP-ELISA), alkaline phosphatase immunoassay (APIA) and the circumoval precipitin test (COPT). Briefly, criterion I: persons with Schistosoma mansoni eggs in stools; criterion II: persons without eggs in stools, with positive COPT, without previous antischistosome chemotherapy in the last year; and criterion III: persons without eggs in stools, with negative COPT, with two positive immunoenzymatic tests (SMP-ELISA and APIA), and with no previous chemotherapy. The incorporation of serological tests to epidemiologic surveillance in areas of low-transmission tries to compensate the underestimation of prevalence based only on parasitological diagnosis.

Immunogenicity of polymerizable synthetic peptides derived from a vaccine candidate against schistosomiasis: the asparaginyl endopeptidase (Sm32).

The asparaginyl endopeptidase (Sm32) is expressed in the gastrodermal cells of the schistosome gut and in the head glands of the cercariae. Possibly, Sm32 hydrolyzes pro-proteins involved in the degradation of host hemoglobin [Parasitol. Today 12 (1996) 125]. Preliminary evidences using an Sj32/Sm32 murine vaccine have shown a profound effect on oviposition and worm burden [Chin. J. Schist. Control. 7 (1995) 72; Bull. Human Med. Univ. 24 (1999) 225; Vaccine 20 (2002) 439]. The importance of Sm32 as a novel vaccine candidate is based on the possibility of preventing the maturation of other cathepsins and/or preventing schistosome skin invasion. We studied the immunogenicity of polymerizable peptides derived from Sm32 to select potential protective epitopes. Sm32 prediction of T and B epitopes and homology studies with human legumain were performed. Among the variety of factors that influence the antibody response, we specifically examined the effect of: (i) genetic background of mouse strain, inbred (C57BL/6) versus outbred (Swiss) mice; and (ii) vaccination with a single peptide versus pool of peptides. Swiss mice raised antibodies to three different regions of the Sm32, as tested by the Multiple Antigen Blot Assay (MABA): 182-215 (peptides IMT-70 and 72), 244-273 (IMT-64) and 336-355 (IMT-66). None of these regions were immunogenic for C57BL/6. On the contrary, other peptides, IMT-4 (21-40), IMT-12 (101-120) and IMT-26 (292-313) were highly immunogenic for this inbred strain. Only Swiss mice immunized with a single peptide (IMT-64 and 72) or with three different pools of IMT-peptides (Pool A-II: 14, 16, 18, 70, 72, 89; pool A-III: 22, 64, 24, 26, 28 and pool A-V: 64, 66, 28, 70, 72) recognized the original protein in a crude extract of the worm antigen by Western blot. Peptides IMT-64, 14 and 26 were responsible for this recognition. In general, the vaccination with pool of peptides was more immunogenic for both mouse strains. Predicted B cell epitopes, with hydrophilicity scores over +10 (IMT-12, 64, 26) were always immunogenic after either single or combined peptide vaccination. Sm32 sequences 41-80 (IMT-6 and 8), 141-160 (IMT-16) and 182-215 (IMT-70 and 72) were nearly identical to the corresponding human legumain regions and should be excluded from the human vaccine. We can conclude that the regions of Sm32 that were recognized by antibodies of mice immunized with polymerizable peptides depended on the mice strain and on the hydrophilicity score of the peptides.

Schistosomiasis mansoni in areas of low transmission: epidemiological characterization of Venezuelan foci.

Severe schistosomiasis is a rare event in Venezuela nowadays, after a successful national campaign by the Schistosomiasis Control Program. Unfortunately, this program has practically disappeared, and snail surveillance in field is not a priority, anymore. Thus, schistosomiasis has become a neglected disease in this country. However, surveys in different populations from the endemic area have shown particular epidemiological features described herein. In five communities we evaluated 2,175 persons and searched for the presence of Biomphalaria glabrata snails. Some markers were used for classifying schistosomiasis foci: mean age of the persons with Schistosoma mansoni eggs in the stools, serological tests, presence of B. glabrata snails, and intensity of infection. Places without B. glabrata snails and with few schistosomiasis cases were defined as "past transmission sites"; a site with abundant snails but few cases was defined as "potential risk"; "new transmission" foci were characterized by the presence of infected snails and young people passing eggs in the stools. A "re-emergent" focus has shared these last features, showing in addition a place where schistosomiasis had been reported before. Recent evidences of active transmission with the increasing dispersion of B. glabrata snails, point out the necessity for the re-establishment of the Schistosomiasis Control Program in Venezuela.

Schistosomiasis mansoni in areas of low transmission. Epidemiological characterization of Venezuelan foci

Severe schistosomiasis is a rare event in Venezuela nowadays, after a successful national campaign by the Schistosomiasis Control Program. Unfortunately, this program has practically disappeared, and snail surveillance in field is not a priority, anymore. Thus, schistosomiasis has become a neglected disease in this country. However, surveys in different populations from the endemic area have shown particular epidemiological features described herein. In five communities we evaluated 2,175 persons and searched for the presence of Biomphalaria glabrata snails. Some markers were used for classifying schistosomiasis foci: mean age of the persons with Schistosoma mansoni eggs in the stools, serological tests, presence of B. glabrata snails, and intensity of infection. Places without B. glabrata snails and with few schistosomiasis cases were defined as "past transmission sites"; a site with abundant snails but few cases was defined as "potential risk"; "new transmission" foci were characterized by the presence of infected snails and young people passing eggs in the stools. A "re-emergent" focus has shared these last features, showing in addition a place where schistosomiasis had been reported before. Recent evidences of active transmission with the increasing dispersion of B. glabrata snails, point out the necessity for the re-establishment of the Schistosomiasis Control Program in Venezuela

Parasite and vertebrate host genetic heterogeneity determine the outcome of infection by Schistosoma mansoni.

Intraspecific variation in Schistosoma mansoni infection and modulation of its expression by vertebrate host genetics was studied by evaluation of some biological parameters of the infection in BALB/c and C57BL/6 mice infected with one Brazilian (BH) and two Venezuelan (YT and SM) laboratory strains of the parasite. Mice infected with 60 cercariae of each parasite strain were euthanized at 5, 6, 8, and 12 weeks. Parameters recorded included the number of adult worms recovered by portal perfusion (infectivity); the number of eggs in the feces, the intestine, and the liver; and the ability of the eggs to cross the intestine, expressed as a quotient of the number of eggs in the intestine versus the feces. Results showed that the parasite appeared to determine the infectivity, the sex ratio, the onset and timing of oviposition, the number of eggs produced, initial egg laying toward the liver, and the ability to cross the intestinal wall. In this sense the BH strain appeared to be the most efficient and the SM strain, the most delayed; the YT strain was intermediate, although closer to the SM strain. On the other hand, the host appeared to influence the susceptibility to infection, the fecundity, and the percentage of eggs distributed in the liver and in the intestine during the chronic stage. In this sense, although they have been shown to be less susceptible to infection than BALB/c mice, C57BL/6 mice permit more eggs to be produced and exhibit similar numbers of eggs in the intestine and the liver at certain time points. It appears from these results that parasite genetics is essential for the outcome of infection with S. mansoni, but some characteristics may be quantitatively modulated by host genetics.

Enzyme activities in Schistosoma mansoni soluble egg antigen.

Schistosoma mansoni egg antigens are mostly responsible for the granulomatous pathology in human intestinal schistosomiasis. Several previous studies have indicated that the induction of an immune response against some parasite enzymes may protect against pathology. The present work was designed to identify enzyme activities present in a standard soluble egg antigen (SEA) preparation. Simple colorimetric analyses were performed incubating SEA with 2-naphthyl, 2-naphthylamide (2NA), or p-nitrophenyl substrates at different pHs in the absence of added effectors. Results showed prominent acid phosphatase (pH 5.4), alkaline phosphatase (pH 8.5), and N-acetyl-beta-glucosaminidase (pH 5.4) activities. Relevant peptidase activities were also detected at pH 6.5-7.5 against 2NA derivatives of (1) aliphatic (alpha-Ala > beta-Ala > Leu > Met > S-benzyl-Cys), polar (Ser > Gln), basic (Arg > Lys > ornithine), and acidic (Glu) amino acids; (2) dipeptides: X-Ala (X = Gly > Leu > Lys > Asp), X-Arg (X = Ala > Arg > Phe > Gly > Pro > Asp), Ser-Met, and Phe-Pro; and (3) tripeptides (Ala-Phe-Pro > Phe-Pro-Ala). The data demonstrated that S. mansoni SEA contains a rich set of hydrolases with different specificities that might play a role in the egg physiology and possibly also in the host-parasite relationships.

Specificity of the solid phase alkaline phosphatase immunocapture assay for the diagnosis of human Schistosoma mansoni infection.

The species specificity of the solid phase alkaline phosphatase immunocapture assay (APIA) for the immunological detection of human immunoglobulin G antibodies to the alkaline phosphatase of adult Schistosoma mansoni was evaluated. Sera from schistosomiasis patients from South America, West Africa, south-east Asia and uninfected control subjects were compared. Only the sera of patients infected with S. mansoni gave positive results. There was no apparent difference between 2 populations infected with S. mansoni, one from South America and the other from West Africa. The results with sera from various regions of West Africa were also indistinguishable. Although the APIA was not able to discriminate the geographical origin of the S. mansoni-infected subjects, the method appeared to be specific for S. mansoni and suitable for use in the immunodiagnosis of schistosomiasis mansoni, particularly in endemic areas where mixed infections of Schistosoma spp. occur.

Natural Schistosoma mansoni infection in wild rats from Guadeloupe: parasitological and immunological aspects.

Rattus rattus is the predominant rodent in the mangrove area of Guadeloupe. Between 1990 and 1991 we found 73 R. rattus and five R. norvegicus. Among the infected rats with Schistosoma mansoni, 59% for R. rattus and 80% for R. norvegicus, the comparison of the median of the worm load was not statistically different. Both species of infected rats showed adult worms and eggs in the lungs and 20% of them showed, at the same time, two and even three generations of worms. Neither adults nor eggs were seen in the intestinal wall or stools of R. norvegicus, instead R. rattus had eggs in the liver, in the intestinal wall and the stools. Therefore, R. norvegicus gets infection as well as R. rattus, but does not participate in the transmission of the schistosomiasis. In order to elucidate this difference, we looked at the humoral recognition of these two rats, to the molecular antigens of the three stages of the parasite: cercaria, adult worm (AWA) and egg (SEA). In general, R. norvegicus recognized cercarial antigens more frequently than R. rattus, 73, 81 and 172 kDa being statistically different. Regarding AWA, molecules 82, 86, 117 and 150 kDa were recognized more often by R. rattus as compared to R. norvegicus. The reverse was true for the 18, 33 and 61 kDa. Only the differences between 61 and 150 kDa molecules were statistically significant. With respect to SEA, R. norvegicus recognized more 28, 45, 47, 49, 64 and 92 kDa molecules than R. rattus, but the latter recognized the 140 kDa molecules of SEA to a higher degree (95 and 140 kDa were significantly different). It is plausible that the immune response to cercarial invasion is more effective in R. norvegicus in allowing the parasites to reach adulthood, but it does not let them live in the mesenteric veins and therefore to lay their eggs in the intestinal wall and feces.

Evaluation of alkaline phosphatase immunoassay and comparison with other diagnostic methods in areas of low transmission of schistosomiasis.

The alkaline phosphatase immunoassay (APIA) is an antibody detection technique which permits the diagnosis of schistosomiasis using a butanolic extract preparation from adult worms. APIA has demonstrated high sensitivity and specificity in previous reports with well characterized human sera. Its potential as a diagnostic tool for epidemiological surveillance was assessed in comparison with three other diagnostic tests: stool examination, ELISA with soluble egg antigen (SEA) and the circumoval precipitin test (COPT). APIA was 100% specific in an area without Schistosoma mansoni transmission and had 89% sensitivity in an endemic area where 69% of the infected subjects excreted less than 100 eggs g of faeces. It was found to be less sensitive in children under 5 years of age who were positive by the COPT test. APIA can be applied as an initial screening test, based on its high sensitivity, specificity, absence of cross-reactivity with intestinal parasites and the fact that it is a technique suitable for use in epidemiological surveillance.

Effect of chemotherapy on immune response to egg antigens of Schistosoma mansoni in chronically infected children from areas of low transmission.

In an attempt to identify antigenic molecules from Schistosoma mansoni eggs, a serological study was performed on children of a Venezuelan town (Caraballeda) in which the transmission of schistosomiasis had been interrupted two years prior to sera sampling. Infected children received treatment with Praziquantel and, based on the disappearance of eggs in the stools plus negativization of the circumoval precipitin test (COPT) one year after treatment, they were classified as either responders or non-responders to chemotherapy. Western blots of soluble egg antigen (SEA) with a very sensitive chemiluminescent substrate were performed. Sera from responder children recognized a 25 kDa band of SEA which diminished significantly after treatment. This was less frequent in non-responder children. When the sera of responder and non-responder children were compared before treatment, we found that the recognition of the 40 and 41 kDa proteins could be predictive of response to chemotherapy. All these antigens, used in ELISA-type techniques, might be of importance in the evaluation and follow-up of large scale schistosome control programmes.

Characterization of mouse monoclonal antibodies against common and repeated epitopes of different developmental stages of Schistosoma mansoni.

Eight IgM monoclonal antibodies (mAbs) obtained from a Schistosoma mansoni chronically infected mouse immunized twice with adult worms butanolic extract (BE) and soluble egg antigen (SEA) were characterized by immunochemical methods. An intense cross-reactivity between different developmental stages was observed by indirect immunofluorescence assay (IIFA) with five anti-SEA mAbs. These mAbs appeared to recognize glycosidic residues, as suggested by 1) the inhibition of their reactivity by periodate oxidation of SEA, 2) the multiple polypeptide recognition in radioimmunoprecipitation and immunoblot assays and 3) reactivity with KLH. Anti-SEA mAbs were able to mediate in vitro killing of schistosomula but they were not consistently able to mediate passive transfer immunity in vivo. Three of anti-SEA mAbs were suitable for the performance of a sandwich ELISA for antibody detection in S. mansoni infected patients, allowing an increase in the signal to noise ratio as compared to the direct ELISA SEA method. Three anti-BE mAbs showed a more stage restricted pattern of antigen recognition by IIFA. Only one out of three seemed to be directed against glycan residues, but the other mAbs showed a plural pattern of polypeptide recognition on BE immunoblot, suggesting that repeated epitopic motifs are also present in different proteins within the same parasite developmental stage.

Schistosoma mansoni: surface membrane isolation with lectin-coated beads.

Lectins from Lens culinaris and Arachis hypogaea immobilized on polyacrylamide beads were used for selective isolation of glycosylated surface membrane domains of adult Schistosoma mansoni worms, and the method was compared with the membrane isolation procedure developed with polycationic (Affi-Gel) beads. The lentil lectin proved to be suitable for interaction with surface membrane components: an increment in the specific activities of tegumental phosphohydrolases was observed in the bound fraction with respect to that observed in a total worm homogenate. A characteristic polypeptide pattern on gel electrophoresis was also seen, more restricted than that obtained with the bound Affi-Gel fraction. Immobilized peanut lectin was not successful as a method for isolating membrane material from the tegument of adult worms. Solubilization and dissociation of the lentil lectin-bound enzyme markers was achieved after addition of detergent and competing sugars. Glycosylation of the solubilized enzymes was further confirmed by affinity chromatography with fresh lentil lectin-coated beads. These results, together with histochemical evidences, suggest that the active sites of some of these enzymes are located within or close to the cytoplasmic leaflet of the surface tegumental membranes, and allow us to propose a model for the double surface membrane complex where some proteins may be crossing the two bilayers.

Isoenzyme studies in one Brazilian and two Venezuelan strains of Schistosoma mansoni.

1. Enzyme polymorphism, analyzed by starch gel electrophoresis, was found to be zero for acid phosphatase, phosphoglucomutase, phosphoglucose isomerase, glucose 6-phosphate dehydrogenase, lactate dehydrogenase, malate dehydrogenase and malic enzyme, in one Brazilian and two Venezuelan strains of Schistosoma mansoni. 2. All loci studied were monomorphic within strains, but the isoenzymic patterns were, however, different among the strains. 3. Results suggest a drastic loss of the genetic variability usually found in natural populations.

Immunocapture assay for quantification of human IgA antibodies to parasite antigenic enzymes. Application with the alkaline phosphatase of Schistosoma mansoni.

Conditions are described for using solid phase adsorbed jacalins in an immunocapture assay for IgA antibodies to the alkaline phosphatase of Schistosoma mansoni. Microtiter plates were activated with polylysine and jacalins were covalently adsorbed by means of glutaraldehyde. From three different jacalins, the one purified from seeds of Artocarpus tonkinensis showed the lowest non-specific adsorption and was used for further studies. Comparing solutions of bovine serum albumin, ovalbumin and Tween 20, it was shown that the latter was most successful in blocking non-specific adsorption. Low serum dilutions resulted in a less efficient IgA capture by the adsorbed jacalin than higher dilutions. Under optimal working conditions, a high correlation could be shown between the presence of specific anti-alkaline phosphatase antibodies of IgA isotype and IgG isotype.

Experimental chemotherapy of Schistosoma mansoni with praziquantel and oxamniquine: differential effect of single or combined formulations of drugs on various strains and on both sexes of the parasite.

The susceptibility of two Venezuelan (YT and SM) and one Brazilian (BH) strain of Schistosoma mansoni to single oral doses of praziquantel (Pz; 250 or 500 mg/kg), oxamniquine (Ox; 40, 60, or 100 mg/kg) or to low-dose combinations of both drugs (33 mg/kg Pz and 25 mg/kg Ox; 66 mg/kg Pz and 12.5 mg/kg Ox; 250 mg/kg Pz and 40 mg/kg Ox) was experimentally evaluated in mice. At lower doses of either drug, adult worms of the SM isolate were less susceptible than those of the BH and YT isolates. However, no difference in liver or intestinal egg counts (IECs) could be detected among the isolates after this treatment. At such doses, Pz was better than Ox at reducing IECs. In spite of lowered IECs, eggs continued to accumulate in the liver after Ox treatment. At higher individual doses or following treatment with low-dose combinations of both drugs, no difference in susceptibility could be detected among the parasite isolates. Under such conditions, oviposition was drastically reduced in all three isolates. We confirm that Ox preferentially kills male parasites and present for the first time evidence for the preferential killing of female worms by Pz. We propose that the synergistic effect obtained in the present study and in other investigations using low-dose combinations of both drugs may be due to the preferential cytotoxicity of each drug against a different parasite sex.

New approaches for the control and eradication of schistosomiasis in Venezuela.

Schistosomiasis in America with the exception of Brazil, behaves as a chronic mild disease with few clinical manifestations due to low parasite burden. These features restrict the clinical and parasitological diagnosis. The most commonly used stool examination method, Kato-Katz, becomes insensitive when the majority of individuals excrete less than 100 eggs/g of feces. In view that antigen-detecting techniques have not been able to reveal light infections, the antibody detecting assays remain as a very valuable diagnostic tool for epidemiological surveillance. The Venezuelan Schistosomiasis Research Group (CECOICE) has designed a mass chemotherapy strategy based on sero-diagnosis. Since blood sampling is one of the important limiting factors for large seroepidemiological trials we developed a simple capillary technique that successfully overcame most of the limitations of blood drawing. In this sense, ELISA seems to be the most adequate test for epidemiological studies. Soluble egg Schistosoma mansoni antigen (SEA) has been largely used in Venezuela. The sensitivity of ELISA-SEA in our hands is 90%, moreover its specificity reach 92% when populations from non-endemic areas but heavily infected with other intestinal parasites are analyzed. The Schistosomiasis Control Program is currently carrying out the surveillance of endemic areas using ELISA-SEA as the first screening method, followed by the Circumoval Precipitin test for validation assay. The results with these two serological techniques allowed us to defined the criteria of chemotherapy in populations of the endemic areas. On the search of better diagnostic technique, Alkaline Phosphatase Immunoenzyme Assay (APIA) is being evaluated in field surveys.

Parasite enzymes as a tool to investigate immune responses.

Previous evidences reported by us and by other authors revealed the presence of IgG in sera of Schistosoma mansoni-infected patients to immunodominant antigens which are enzymes. Besides their immunological interest as possible inductors of protection, several of these enzyme antigens might be also interesting markers of infection in antibody-detecting immunocapture assays which use the intrinsic catalytic property of these antigens. It was thus thought important to define some enzymatic and immunological characteristics of these molecules to better exploit their use as antigens. Four different enzymes from adult worms were partially characterized in their biochemical properties and susceptibility to react with antibodies of infected patients, namely alkaline phosphatase (AKP, Mg2+, pH 9.5), type I phosphodiesterase (PDE, pH 9.5), cysteine proteinase (CP, dithiothreitol, pH 5.5) and N-acetyl-beta-D-glucosaminidase (NAG, pH 5.5). The AKP and PDE are distinct tegumental membrane-bound enzymes whereas CP and NAG are soluble acid enzymes. Antibodies in infected human sera differed in their capacity to react with and to inhibit these enzyme antigens. Possibly, the specificity of the antibodies related to the extent of homology between the parasite and the host enzyme might be in part responsible for the above differences. The results are also discussed in view of the possible functional importance of these enzymes.