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SUMMARYSeveral recombinant clones expressing antigens from Echinococcus granulosus were isolated previously from a parasite cDNA library using cystic hydatid disease (CHD) patients' sera or rabbit hyperimmune antiserum against a lipoproteic fraction from bovine cyst fluid. Six of these antigens were expressed in Escherichia coli and the purified recombinant proteins were tested in enzyme-linked immunosorbent assay (ELISA) for specific IgG with a panel of sera from patients with surgically confirmed ( n = 58) or immunologically diagnosed ( n = 71) CHD. Sera from clinically normal individuals ( n = 203) and sera from individuals with other helminthic infections ( n = 65) were assayed for the assessment of specificity. A cut-off value was determined by receiver-operating-characteristic plots for each antigen. A recombinant antigen B subunit (AgB8/2) presented the highest sensitivity (93·1%), considering the group of sera from patients with CHD surgically confirmed, and specificity (99·5%) and is proposed as the basis for an immunodiagnostic test. The other recombinant antigens tested presented sensitivities between 58·6% and 89·7%, and three of them were considered of complementary value. In subclass-specific ELISA, different IgG isotypes showed dominance in the response for each of the recombinant antigens. There was a clear predominance of IgG4 response for all antigens tested, indicating that this would be the subclass of choice to be assessed for these recombinant proteins.

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The antibody response was followed weekly during 68 weeks in 17 Balb/c mice intraperitoneally (i.p.) infected with 2000 Echinococcus granulosus protoscoleces (PSC) and in three mice i.p. immunized with 2000 dead PSC. Antibodies against hydatid cyst fluid (HCFA) and its peptidic (periodate-resistant) and carbohydrate (periodate-sensitive) epitopes were titrated by ELISA. Avidity and the antigen recognition pattern of antibodies were also analysed during infection and immunization by ELISA and immunoblot, respectively. The antibody response of infected mice showed quantitative and qualitative variations during infection, since both titre as well as recognition of peptide and carbohydrate epitopes in HCFA depended on time post infection. No avidity maturation was evident during the course of infection. Sera from infected mice recognized the 38 kDa subunit of Ag5 but did not react with the 8 kDa subunit of AgB. On the contrary, the antibodies response of immunized mice showed only one peak of antibodies that recognized both peptidic and carbohydrate epitopes of HCFA. In addition, sera from these mice recognized mainly 60 and 110 kDa bands. Our results suggest that: a) avidity and antigen recognition patterns of antibodies in mice treated with live PSC are different from those treated with dead PSC; b) antibodies against HCFA glucidic or peptidic epitopes appear at different times post infection.

A carbohydrate enriched soluble fraction (CHP) was prepared by mild treatment of viable Echinococcus granulosus protoscoleces (PSC) with the enzyme endoglycosidase-F (endo-F) and characterized by SDS-PAGE, glycoside- inhibition ELISA, and immunoblotting. Three groups of four BALB/c mice were immunized with the CHP, with the remaining deglycosylated PSC (DGP) and with dead PSC (DPSC) to analyse the relative immunogenicity of carbohydrates on the surface of PSC. A fourth sentinel group was not immunized. The antibody response was analyzed during primary and hyperimmune responses. Specific antibody titres (IgG and IgM) against somatic PSC antigens (PSA) were evaluated by ELISA and their antigen recognition pattern by immunoblotting, discriminating carbohydrate and peptide specific antibody responses by periodate treatment of PSA. The avidity index of those antibodies and the titer of non-specific immunoglobulins during the whole protocol were evaluated by ELISA in vitro mitogenic activity of CHP was also evaluated. The results indicated: 1) immunodominance of surface carbohydrate epitopes from PSC, 2) predominant IgM and low avidity response against these epitopes, 3) a dramatic increase of non-specific antibody titres and 4) in vitro mitogenic activity of CHP.