The enzyme-linked immunosorbent assay (ELISA) technique can play an important role in the early detection of fascioliasis. However, they have some diagnostic limitations, including cross-reaction with other helminths. It seems that the combination of recombinant parasite proteins as antigen can reduce these problems. Hence, the present study was aimed to design and confirm the antigenic recombinant multi-epitope (rMEP) construct of three protein epitopes (linear and conformational B-cell epitopes) of the parasite using immunoinformatic tools. For this purpose, the tertiary structures of Fasciola hepatica cathepsin-L1, saposin-like protein 2 and 16.5-kDa tegument-associated protein were predicted using the I-TASSER server. Validation of the modelled structures was performed by Ramachandran plots. The antigenic epitopes of the proteins were achieved by analysing the features of the IEDB server. The synthesized gene was cloned into the pET-22b (+) expression vector and transformed into the Escherichia coli BL21. Sodium dodecyl sulfate polyacrylamide gel electrophoresis was used to verify and analyse the expression of the rMEP protein. Western blotting was utilized to confirm rMEP protein immunogenicity in two forms, one using an anti-His tag antibody and the other with human pooled sera samples (fascioliasis, non-fascioliasis and negative control sera). Our results demonstrated that the rMEP designed for the three proteins of F. hepatica was highly antigenic, and immune-detection techniques confirmed the antigen specificity. In conclusion, the presented antigenic multi-epitope may be very helpful to develop serodiagnostic kits such as indirect ELISA to evaluate the proper diagnosis of fascioliasis in humans and ruminants.

Excretory/secretory (ES) products obtained by in vitro culture of infective-stage larvae of Trichinella spiralis and T. pseudospiralis were injected intramuscularly at various intervals into mice. Mini-osmotic pumps containing T. spiralis ES products were also implanted subcutaneously and intraperitoneally into rats. The introduction of ES materials into muscles elicited extensive lesions which included dissolution of myofibres, mobilization of mononuclear and polymorphonuclear leucocytes, angiogenesis, hypertrophy of myonuclei, myotube formation, mitosis, muscle bundles becoming rounded and separated from each other, disappearance of Z, I and A bands of sarcomeres, increase in endoplasmic reticulum and Golgi complexes, decrease in glycogen and relocation of mitochondria. These are considered as degenerative/regenerative changes of muscles to injury. Immunodominant epitopes of specific 45–53 kDa glycoproteins in ES antigens of T. spiralis could not be detected in hypertrophic nuclei of injected muscles by using polyclonal and monoclonal antibodies and immunocytochemical methods. ES products of T. spiralis failed to stimulate unsensitized lymphocytes in the lymphocyte transformation test. Infective-stage larvae of T. spiralis released from muscles were found capable of forming nurse cells after injection subcutaneously into rats. It is postulated that the invasion of muscles by trichinellids elicits two independent events, i.e. a general degenerative/regenerative response of muscles and a specific change in genomic expression of myonuclei. The two events are probably mediated by different effector molecules.

Sixteen monoclonal antibodies (MABs) were selected for their reactivity with adult schistosome tegument. The distribution of target epitopes in different tissues of the adult and in various developmental stages was investigated by indirect immunofluorescence. The distinct patterns of reactivity of the MABs permitted their classification into 9 groups. The distribution of epitopes in larvae, particularly 3 h schistosomula, generally mirrored that in adults. A change in distribution of epitopes coincided with transformation from cercaria to schistosomulum providing a marker for this process. Two MABs reacted with surface membranes of intact cercariae, and 5 with the surfaces of intact 3 h schistosomula. The target epitopes of these 5 MABs were present in the tegument of adults, but not accessible externally. These observations suggest masking of antigens, not shedding, in the course of development. Indeed, no MAB reacted with intact lung or liver worms unless they were damaged. Three MABs reacted with membranes of the tegument and most other tissues, implying properties in common. Evidence for shared functions between epithelia was provided by the reactivity of 2 MABs with the tegument and protonephridia. Eight MABs bound both to the tegument and its cell bodies. Since the latter contains the machinery for protein synthesis, it is difficult to explain why 5 MABs bound to the tegument alone. Little cross-reactivity was found with the tissues of the tapeworm Hymenolepis, rather more with the closely related Schistosoma japonicum. It was concluded that tegument antigens are to a degree tissue specific but definitely not stage specific.

Twenty-one monoclonal antibodies, obtained after immunization of mice with erythrocytic stages of Plasmodium falciparum, produced a double dot image in IFA. Immunoelectronmicroscopy indicated that the mAbs reacted with the rhoptries. Rhoptries are pear-shaped apical organelles, believed to be involved in invasion of the host cell by the parasite. The mAbs all immunoprecipitated the high molecular weight antigen complex. Some mAbs recognized on immunoblots only 1 protein of this complex, whereas others reacted with RhopH1 and RhopH3, or RhopH2 and RhopH3 or with the 3 proteins. An additional antigen of 52 kDa was also recognized by some of the mAbs. The epitopes defined by the mAbs were present in most of the 40 P. falciparum strains or isolates studied by IFA. Interestingly, the mAbs also reacted with high titres on P. vivax and P. ovale, but produced images that did not indicate an apical location. The mAbs failed to react on the non-human malaria parasites studied, P. cynomolgi and P. inui. On P. berghei or P. chabaudi parasites, only 5 mAbs gave a positive reaction, labelling a large network outside the parasite. Finally, the mAbs did not react with P. falciparum sporozoites, indicating that the rhoptries of merozoites and sporozoites, the two invasive stages of the malaria life-cycle are equipped with distinct sets of proteins.

The relationship between antigens associated with the surface of newly transformed schistosomula of Schistosoma mansoni and the tegumental surface membrane of adult S. mansoni worms has been further explored. Immunoprecipitation of detergent-solubilized 125I-tegumental surface membrane antigens of adult S. mansoni with antibodies from mice vaccinated with highly irradiated S. mansoni cercariae revealed major antigens of Mr 32, 20, 15 and 8K. The Mr 32 and 20K antigens have been previously demonstrated to be antigenically and electrophoretically identical to major antigens on the schistosomulum surface. The Mr 15 and 8K antigens, on the other hand, have not been identified by the immunoprecipitation of 125I-schistosomulum surface antigens, although a distinct schistosomulum surface antigen of Mr 15K is precipitated by antibodies from mice vaccinated with highly irradiated cercariae. Nevertheless, it was shown that antibodies to the Mr 15 and 8K antigens were specifically absorbed from vaccinated mouse serum by intact, live schistosomula, demonstrating that the Mr 15 and 8K antigens are exposed on or released from the schistosomulum surface. In contrast, absorption of the antiserum with eggs failed to remove antibody against any of the four tegumental membrane antigens examined. The Mr 15 and 8K antigens were shown to be recognized via polypeptide epitopes and not periodate-sensitive carbohydrate epitopes, further emphasizing the similarity of these to the well-characterized Mr 32 and 20K tegumental surface membrane antigens. A general relationship between schistosomulum surface, adult tegumental membrane and egg antigens was demonstrated by ELISA, using antibodies raised against the three antigenic fractions. It was shown that both the egg and adult tegumental membrane antigens cross-react with the schistosomulum surface, but that the egg and adult membrane antigens exhibit very little, if any, mutual cross-reactivity. This antigen divergence possibly enables the host to dissociate pathological, anti-egg responses from potentially protective anti-membrane responses during the course of natural infection. It also suggests that adult membrane antigens could be used in an anti-schistosome vaccine without the possible complication of inducing pathological responses.

The major surface protease (msp or gp63) of Leishmania plays a major role in the host–parasite interaction. We analysed here the structure of the msp gene locus in Leishmania (Viannia) braziliensis and compared it to results obtained in other species. Physical mapping of cosmid contigs revealed a minimum of 37 genes per haploid genome and at least 8 different msp gene families. Within the same organism, these genes showed a nucleotide sequence varying in certain stretches from 3 to 34%, and a mosaic structure. From an evolutionary point of view, major differences were observed between subgenera Viannia and Leishmania, both in terms of msp gene number and sequence. Within subgenus Viannia, phenetic analysis revealed three clusters in which sequence variants of L. (Viannia) braziliensis and L. (Viannia) guyanensis were interspersed. Functional implications of our results were explored from predicted L. (Viannia) braziliensis protein sequences: regions encoding the msp catalytic site showed a conserved sequence, while regions encoding surface domains possibly involved in the host–parasite interaction (macrophage adhesion sites and immunodominant B-cell and T-cell epitopes) were variable. We speculate that this would be an adaptive strategy of the parasite.