This proposal aims to determine the genetic basis of acute virulence in a recombinant clone of the parasite Toxoplasma gondii. Most T. gondii isolates belong to one of three clonal "Types" with distinct disease outcomes in mice. Infection with a single Type I organism is lethal, while mice survive 1000-fold higher inoculations of Type II or III. Remarkably, a T. gondii clone (called S23) resulting from genetic recombination between type II and III strains is 1000 times more virulent than either parent, showing that loci from avirulent strains can interact to produce virulent phenotypes. I will dissect this unexpected, but epidemiologically important, observation by identifying the alleles responsible for the phenotype of clone S23. After developing an in vitro correlate for T. gondii virulence based on S23-specific host cell responses to infection, I will conduct backcrosses of S23 to its avirulent parents, and use quantitative trait loci (QTL) mapping to identify genomic segments linked virulence. Then, I will carry out allele exchanges between the avirulent parents of clone S23 different to identify the virulence alleles themselves. This work will improve our understanding of the evolution of T. gondii virulence under both laboratory and field conditions.