A Genetic Screen to Isolate Toxoplasma gondii Host-cell Egress Mutants

This video illustrates a new phenotypic screening method to isolate mutants with a temperature sensitive egress phenotype. Mutagen and parasites are applied to cultures of toxoplasma infected human foreskin fibroblasts. For two hours, the non invading parasites are removed and the flask is transferred to 40 degrees Celsius to induce phenotype development.

After 26 hours, the vacuoles contain eight to 16 parasites. Next cells are stimulated with egress inducing compounds. Mutant parasites with a defect in egress will remain intracellular, whereas wild type parasites will egress into the medium.