Toxoplasma gondii is an important opportunistic pathogen in immunocompromised individuals. Successful propagation in an infected host by this obligate intracellular parasite depends on its ability to enter and exit host cells. Egress from the cell can be artificially induced by causing fluxes of calcium within the parasite with the use of calcium ionophores. While this ionophore-induced egress (IIE) has been characterized in detail, it is not known whether it mimics a normal physiological process of the parasite. This is underscored by the fact that mutants in IIE do not exhibit strong defects in any of the normal growth characteristics of the parasite in tissue culture. We have isolated and characterized a T. gondii mutant that along with a delay in IIE exhibits a severe defect in establishing a successful infection in vivo. In tissue culture this mutant displays normal ability to invade, divide within cells and convert into the latent encysted bradyzoite form. Nevertheless, mice infected with this mutant are less likely to die and carry less brain cysts than those infected with wild type parasites. Thus, our results suggest that normal response to calcium fluxes plays an important role during in vivo development of T. gondii.