Comparison of different serotests for specific Toxoplasma IgM-antibodies (ISAGA, SPIHA, IFAT) and detection of circulating antigen in two cases of laboratory acquired Toxoplasma infection.

Two symptomatic Toxoplasma infections of laboratory personnel have been serologically followed up for 5.5 and 10 months, respectively. Results obtained by commonly used test systems (indirect fluorescent antibody tests for IgG and IgM antibodies, complement fixation test) were compared with those of two recently developed and improved tests for IgM detection (immunosorbent agglutination assay [ISAGA] and solid-phase indirect haemadsorption assay [SPIHA] as well as with those of a test designed for the detection of circulating antigen (cag-ELISA).

Efforts towards a vaccine against Toxoplasma gondii: a review.

In a review, past as well as present investigations carried out towards a vaccine against toxoplasmosis are outlined. A historical retrospect of the various immunization experiments is given, recent research projects intending the characterization of antigens that are relevant to host protective immunity are described, and a prospect to future problems and developments expected in the field is drafted.

Demonstration of a specific Echinococcus multilocularis antigen in the supernatant of in vitro maintained protoscolices.

Serodiagnosis of echinococcosis is still often met with difficulties resulting from unspecific reactions due to crude antigens which contain numerous host-derived proteins. In order to eliminate host protein contamination we produced Echinococcus multilocularis antigen by methods of in vitro technique: Evaginated protoscolices of Echinococcus multilocularis isolated from experimentally infected Mongolian gerbils (Meriones unguiculatus) were maintained in RPMI 1640 medium. Although no serum proteins were added, protoscolices could be kept alive for more than 2 weeks. The supernatants harvested from protoscolices cultures were tested for their immunoreactivity against sera of patients with confirmed alveolar or cystic echinococcosis, cysticercosis, schistosomiasis or fascioliasis by means of SDS-PAGE and immunoblotting. A polypeptide band at about 62,000 mol. mass was identified which proved to be specifically immunoreactive with sera from patients with alveolar echinococcosis, whereas sera from patients with cystic echinococcosis or with other helminthic infections as well as sera from healthy blood-donors did not react with this polypeptide band. Thus, the use of supernatant antigen in immunoblotting technique allows a serological differentiation between infections with Echinococcus multilocularis and those with Echinococcus granulosus and provides an accurate diagnosis of alveolar echinococcosis.

Higher yields and increased purity of in vitro grown Toxoplasma gondii.

An improved method for in vitro cultivation of Toxoplasma gondii is described. Commonly, confluent cell monolayers are infected with Toxoplasma; higher yields and an increased purity could be obtained instead when host cells were immediately infected after subculture: Depending on the initial parasite inoculum 2-3 fold increased multiplication rates were achieved and simultaneously the amount of host cell debris was reduced to 10-20 per cent compared to that deriving from infected cell monolayers. This may be ascribed to a facilitated invasion of host cells by Toxoplasma when the cell's glycocalyx is irritated by trypsinization and to a more far-reaching exploitation of young separated host cells in contrast to cells of monolayers. Moreover, the Toxoplasma trophozoites could be harvested from tissue culture already four days after infection without any change of medium, which makes the method very economical.

Cryptosporidiosis in Ogun State, south-west Nigeria.

During the period of one month in the rainy season in Ogun State in south-west Nigeria 479 stool samples (420 of which were diarrhoea stool samples) were examined for cryptosporidiosis. Oocysts were detected in 2.3% of all stools, in 2.6% of diarrhoeal stools and in 5.3% out of 150 children with diarrhoea. Cryptosporidium was the sole pathogen detected in six of 11 cases. In addition to cryptosporidia also Entamoeba histolytica, Blastocystis hominis, Giardia lamblia, Ascaris lumbricoides, and Trichuris trichiura could be found. Compared to other studies in tropical countries, the frequency of infection was lower in south-west Nigeria.

Experimental studies on circulating antigen of Toxoplasma gondii in intermediate hosts: criteria for detection and structural properties.

The present study was performed to demonstrate circulating antigen (cag) of Toxoplasma gondii in the sera of orally and intraperitoneally infected rabbits and swine, to determine the time of their appearance after infection, and to characterize the antigenic components of the cag by means of affinity chromatography, SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and electrophoretic blot onto nitrocellulose sheets. Cag, as detected by an indirect ELISA, was found in the sera of both rabbits and swine from week 5 to week 8 after infection. Electrophoretic separation of cag extracted from swine and human sera showed 5 and 8 distinct protein bands, respectively, the molecular weight of which ranged between 25 and 100 kD. After Western blot, 2 of the 5 protein bands of swine-cag (27 and 57 kD) and 3 of the 8 protein bands of human cag (27, 32, and 57 kD) reacted with the anti-Toxoplasma antibody used in the ELISA. On the basis of the data presented, the influence of the dose and mode of infection as well as that of the preparation method of antisera on cag detection is discussed.

A solid-phase indirect haemadsorption assay (SPIHA) for detection of immunoglobulin m antibodies to Toxoplasma gondii: application to diagnosis of acute acquired toxoplasmosis.

A solid-phase indirect haemadsorption assay (SPIHA) for the detection of immunoglobulin M (IgM) antibodies to Toxoplasma gondii is described. Polystyrol microtiter plates are coated with anti-human IgM (mu-chain-specific) antibodies and then sequentially allowed to react with patient's serum and sheep erythrocytes sensitized with soluble antigen of Toxoplasma gondii. A total of 111 sera were tested in fluorescent antibody test (FAT and IgM-FAT), complement fixation test (CFT), indirect haemagglutination assay (IHA), and in SPIHA. 47 sera were from individuals with a suspected or verified acute Toxoplasma infection. In most of these cases the SPIHA allowed a clear interpretation with respect to the status of the infection, even when the IgM-FAT was not conclusive. In contrast to IgM-FAT, rheumatoid factor or exceedingly high specific IgG antibodies did not interfere with results in SPIHA. A laboratory-acquired infection enabled us to demonstrate the course of immune response measured by SPIHA as well as by IgM-FAT, FAT, CFT, and IHA. The method proposed here is well appropriated to IgM detection, simple to perform, inexpensive, and thus representing an alternative to IgM-FAT, convenient for the routine laboratory.

[Immunoenzyme test with whole trophozoites of Toxoplasma gondii]. / Enzymimmuntest mit ganzen Trophozoiten von Toxoplasma gondii.

In a comparative study 151 human sera were tested for IgG and IgM antibodies against Toxoplasma gondii in the Fluorescent-Antibody-Test (FAT) as well as in the Enzyme-Linked Immunosorbent Assay (ELISA) using the same antigen (whole trophozoites of Toxoplasma gondii). A positive correlation was found between the results obtained in FAT and ELISA. The detection of specific IgG antibodies revealed a correlation of 100% and 97.5% in positive and negative sera, respectively. Only 8 out of 111 positive sera showed deviations of more than +/- 1 step within the titre ranges. While detecting specific IgM antibodies a highly significant correlation (99.3%) between FAT and ELISA was obtained for negative sera. 5 positive IgM sera showed titre end-points of 1:64 and 1 serum an end-point titre of 1:256 in FAT as well as in ELISA; however, a comparison was not done because of the small amount of positive IgM sera having been available.