Toxoplasma gondii has an unusual population structure consisting of three clonal lineages that predominate in North America and Europe. This simple pattern has encouraged the use of only a few laboratory isolates that are representative of each lineage. Principle among these is the type I RH strain, originally isolated from a child with encephalitis some 70 years ago. Comparison of different passages of the RH strain that have been propagated differently over the intervening time period revealed that the commonly used clonal line called RH-ERP was not representative of natural isolates of the type I lineage. Notably, RH-ERP formed much larger plaques than other type 1 strains, including a separate, earlier derived isolate of the RH strain. The RH-ERP variant also showed enhanced extracellular survival, faster growth, and decreased differentiation compared to the prototype type I strain GT1. Comparison of gene expression differences in the RH-ERP line revealed that several ABC transporters were upregulated, which may provide a growth advantage in vitro. These findings illustrate that dramatic phenotypic changes can arise in laboratory strains, emphasizing the need for comparison with recent clinical isolates.