Toxoplasma gondii: the biochemical basis of resistance to emimycin.

Emimycin was a potent and selective inhibitor of the growth and nucleic acid synthesis of Toxoplasma gondii in human fibroblasts. An emimycin-resistant mutant of T. gondii lost the pyrimidine salvage enzyme uracil phosphoribosyltransferase, the same enzyme absent in parasites resistant to fluorodeoxyuridine. The mutant resistant to emimycin was completely cross-resistant to fluorodeoxyuridine. Emimycin was as good a substrate as uracil for the uracil phosphoribosyltransferase of T. gondii. [3H]Emimycin supplied in the medium of cultures with actively growing intracellular parasites was converted to emimycin riboside-5'-phosphate in the soluble pool of T. gondii. All other emimycin analogs of uracil-containing nucleotides were also formed but little emimycin riboside diphosphate-N-acetylhexosamine was found. [3H]Emimycin was not converted to analogs of the cytidine nucleotides. When intracellular T. gondii were treated with a concentration of [3H]emimycin that partially inhibited parasite RNA synthesis, much less [3H]emimycin was incorporated into RNA than would be predicted by the amount of intracellular [3H]emimycin riboside triphosphate.

Production of gamma interferon by cultured human lymphocytes stimulated with a purified membrane protein (P30) from Toxoplasma gondii.

Purified P30, the principal iodinatable membrane protein of Toxoplasma gondii, induced proliferation of peripheral blood mononuclear cells from seropositive individuals but not from seronegative individuals. Culture supernatants from stimulated cells of seropositive individuals blocked the growth of T. gondii in human fibroblasts, whereas those from antibody-negative individuals failed to do so. The anti-toxoplasmic effect of culture supernatants correlated with the induction of indoleamine 2,3-dioxygenase and the destruction of tryptophan, as previously described for fibroblasts treated with recombinant gamma interferon (IFN-gamma). The anti-toxoplasmic effect was blocked by monoclonal antibody to IFN-gamma. The protective effect correlated with the amount of IFN-gamma in the culture supernatant, as measured by inhibition of viral CPE. Thus, the level of IFN-gamma appears to be an important immune factor in protection against toxoplasmosis in humans.

Toxoplasma gondii: in vivo and in vitro studies of a mutant resistant to arprinocid-N-oxide.

The anticoccidial drug arprinocid and arprinocid-N-oxide, a metabolite produced in vivo, blocked the growth of Toxoplasma gondii in human fibroblasts. The more potent arprinocid-N-oxide inhibited growth by 50% at 20 ng/ml while arprinocid inhibited at 2 micrograms/ml. For both drugs, the host cell was less sensitive than was the parasite. Hypoxanthine did not reverse the antitoxoplasma activity of either drug. We isolated a parasite mutant, R-AnoR-1 that was 16- to 20-fold more resistant to arprinocid-N-oxide than was the wild type RH T. gondii. This mutant was not resistant to arprinocid in vitro. Arprinocid in a daily oral dose of 136 micrograms regularly protected mice against an otherwise fatal infection with RH T. gondii and 55 micrograms had some protective effect. However, all mice infected with R-AnoR-1 and treated with 360 micrograms arprinocid per day died. Since this mutant is fully sensitive to arprinocid, the form of the drug that is therapeutically active in vivo cannot be arprinocid and is likely to be arprinocid-N-oxide.