D Bout

Identification and purification of the specific antigen of Paracoccidioides brasiliensis responsible for immunoelectrophoretic band E.

A new purified antigen (E2) of Paracoccidioides brasiliensis mycelial growth phase was isolated by immunoadsorption from a crude metabolic soluble extract of the fungus. The antiserum prepared in a rabbit by inoculation of E2 antigen developed only one immunodiffusion line with the crude metabolic extract. Findings on immunological analysis showed that E2 antigen is the antigenic component of immunoelectrophoretic band E. The isolated antigens did not possess detectable alkaline phosphatase activity. It reacted in immunodiffusion tests with all the sera (14/14) from P. brasiliensis infected patients containing precipitating antibodies.

Further observations on the specificity of antigen 5 of Echinococcus granulosus.

The presence of IgE antibodies to antigen 5 of Echinococcus granulosus was detected by means of radioimmunoelectrophoresis in the sera of two of six patients infected with E. multilocularis. Sera from three of these patients gave a precipitin band in gel diffusion tests identical to that produced by a monospecific rabbit anti-E. granulosus antigen 5 serum, when tested against whole hydatid fluid. Sera from 19 individuals infected with Fasciola hepatica, 20 with Schistosoma mansoni, and 5 with with Taenia saginata showed no detectable antibodies against antigen 5 of E. granulosus, The monospecific rabbit anti-E. granulosus antigen 5 serum did not react in immunodiffusion with homologous antigen when absorbed with either 4 mg/ml of whole hydatid fluid or with 200 mg/ml of a soluble E. multilocularis extract. Absorption of the monospecific antiserum with crude antigens of either F. hepatica, Onchocerca volvulus, S. mansoni, or T. saginata did not abolish the reaction with antigen 5. It appears, therefore, that antigen 5 can no longer be considered specific for E. granulosus, but is also present in E. multilocularis. In the light of this observation, some reevaluation of immunodiagnostic tests in hydatid disease will be necessary.

Etude in vitro des phenomenes immunologiques dans la schistosomiase humaine et experimentale—I. Etude comparative in vitro de l'activite lethale d'immunserums sur les formes immatures et sur les adultes de Schistosoma mansoni

Resume The abilitity to culture immature forms (schistosomules) and to maintain adult worms of Schistosoma mansoni has made possible a study in vitro of immunological phenomena in human and experimental schistosomiasis. The existence of a lethal factor leading to cytolysis of the schistosomules within a period of 4 days has been demonstrated in the serum of 74 per cent of patients with S. mansoni schistosomiasis. A significant relationship was found between the presence of the lethal factor and the hepatosplenic form of the disease. This lethal factor was also present in the rabbit and in the rat infected with S. mansoni cercariae and ultrastructural studies have revealed lesions in the surface of the schistosomules. The absence of lethal activity in sera from infections with S. haematobium appears to indicate a certain specificity of the action of the antibody. Moreover, this lethal factor seems of the stage specific, the same sera being without effect on adult schistosomes. Hyperimmunization of rabbits with soluble extracts of S. mansoni adults stimulated the formation of antibodies inactive on schistosomules but cytotoxic for adult schistosomes grown in vitro. The activity which was in the IgG fraction, depended on labile factors in fresh serum. This antibody is not directed against surface antigens. Ultrastructural studies of adult worms show that lesions do not affect the surface of the worm. These experiments seem to indicate that schistosomules are sensitive to antibodies produced in natural or experimental infection. While the adult worms are sensitive to the antibodies induced by immunization with extracts of adult worms, they are not affected by antibodies from natural or experimental infections. Though it does not appear that lethal antibody can be related to immunity, it may have some significance in immunopathology of schistosomiasis in man. The production of a cytotoxic antibody to adult schistosomes may represent a different approach of in vitro studies of the effects of antibodies on schistosomes.

Filaricidal effects of Cyclosporin-A against Dipetalonema viteae in Mastomys natalensis

Outbred male Mastomys natalensis were injected subcutaneously with 100 infective larvae of Dipetalonema viteae obtained from Ornithodorus tartakovskyi. Groups of five animals were treated with 30 mg/kg of the immunosuppressive drug Cyclosporin-A daily for five days (experimental) or Miglyol 812 (control). One group served as untreated controls. Contrary to expectations, 60% of the animals were completely protected against D. viteae and the remainder were partially protected. The mechanism remains unknown.

Kinetics of classes and sub-classes of total immunoglobulins and specific antibodies to Schistosoma mansoni during murine infection

During the course of Schistosoma mansoni murine infection there is a dramatic increase of some immunoglobulins and S. mansoni-specific antibodies. The most substantial response is initiated after 40 days of infection and results in a prolonged increase of total IgG1, IgM and IgA. The maximum increase is respectively 26, 14 and 3-fold the basic immunoglobulin level in control mice. Some anti-S. mansoni classes and sub-classes were studied by an original radio-immunoadsorbent test. Anti-S. mansoni IgG1 and IgM antibodies appear and increase at the same time as that of total IgG1 and IgM. Anti-S. mansoni IgA antibodies appear later (80th day) and correspond to a second peak of total IgA.

Echinococcus granulosus: The distribution of hydatid fluid antigens in the tissues of the larval stage: II. Localization of the thermostable lipoprotein of parasitic origin (antigen B)

Abstract By means of a rabbit monospecific antiserum, the localization of hydatid fluid antigen B, a thermostable lipoprotein was demonstrated in the larval tissues of Echinococcus granulosus . The antigen was present in the cuticular “membrane,” in the tegument of the protoscoleces, and in the substance contained inside the brood capsules. This distribution corresponds exactly to that of PAS-positive substances described previously in the hydatid cyst of E. granulosus . The localization of antigen B and the fact that it was not detected in the excretory system suggest that the sub-tegumental cells participate in its secretion.

Isolation of a specific antigen with alkaline phosphatase activity from soluble extracts of Paracoccidioides brasiliensis

A specific antigen of Paracoccidioides brasiliensis was isolated from a metabolic extract of the fungus. The extraction was made by specific adsorption to and subsequent elution from a column containing a cross linked polymer to which the antibodies of a monospecific rabbit serum had been covalently attached. The purity of the final product was demonstrated by immunodiffusion analysis of the eluate using immune serum produced in a sensitized rabbit. The purified antigen was shown to have cationic electrophoretic mobility and alkaline phosphatase activity.

Echinococcus granulosus: distribution of hydatid fluid antigens in tissues of the larval stage. I. Localization of the specific antigen of hydatid fluid (antigen 5).

Abstract The indirect immunofluorescent test employing a monospecific antiserum has been used to detect the tissue localization of Echinococcus granulosus specific antigen “5.” The antigen was revealed in the inner portion of the germinal “membrane” and in the parenchyma of the protoscoleces. In these stages, it was also demonstrated fixed to the walls of some collecting ducts. It is postulated that the synthesis of the antigen “5” may occur in specialized cells of both the germinal “membrane” and the protoscoleces of the hydatid cysts. The osmoregulatory system of E. granulosus larvae seems to be involved in the transfer of the substance to the cystic cavity.

Serum concentration of specific IgE antibody against Aspergillus fumigatus and identification of the fungal allergen.

Abstract The concentration of specific IgE antibody against whole Aspergillus fumigatus antigens was quantitatively measured in 60 sera from patients with anti-A. fumigatus precipitins and compared with total IgE level. Raised IgE levels were observed in 31 patients. By the use of an immunosorbent prepared with A. fumigatus, IgE antibody against the fungus was quantified. Mean concentration of 10.3 IU/ml of anti-Aspergillus IgE was observed. The mean ratio of anti-Aspergillus IgE to total IgE was 4.8%. Radioimmunoelectrophoresis allowed identification of the allergen in A. fumigatus extracts combined with the IgE antibody. This allergen was distinct from the specific antigen C2 with chymotryptic activity previously shown to be specific for the species A. fumigatus.

In vitro Killing of Entamoeba histolytica trophozoites by interferon-γ-activated mouse macrophages

The effect of murine interferon gamma (IFN-gamma) on macrophage activation for amoebicidal activity was examined. Peritoneal macrophages were harvested from C57BL/6 mice and preincubated with IFN-gamma and/or lipopolysaccharide (LPS). In vitro amoebicidal activity of these macrophages was determined by trypan blue exclusion test against a virulent strain of E. histolytica (IP:0682:1). It was found that in vitro amoebicidal activity was evident in macrophage monolayers treated with both IFN-gamma and LPS. Macrophages treated with IFN-gamma alone did not develop cytotoxic activity unless they were exposed to LPS as a second triggering signal. The ability of IFN-gamma to prime macrophages to respond to trigger signals of LPS and develop cytotoxicity increased with time of incubation, the highest response being observed after 24 h. There was a dose-dependent relationship between the concentrations of both IFN-gamma and LPS used to activate macrophages and the number of dead trophozoites. These data suggest that macrophages are important in host defense against amoebiasis.