Prions consist mainly of PrP(Sc), a pathogenic conformer of host-encoded PrP(C). Prion populations with distinct phenotypes but associated with PrP(Sc), having the same amino acid sequence, constitute distinct strains. Strain identity is thought to be encoded by the conformation of PrP(Sc) and to be maintained by seeded conversion. Prion strains can be distinguished by the cell panel assay, which measures their ability to infect distinct cell lines. Brain-derived 22L prions characteristically are able to infect R33 cells (i.e., are "R33 competent"), as well as PK1 cells in the presence of the inhibitor swainsonine (i.e. are "swa resistant"). Here we report that 22L prions retained their characteristic cell tropism and swa resistance when transferred from brain to R33 cells. However, when transferred from the R33 cells to PK1 cells, they gradually became R33 incompetent and swa sensitive, unless the transfer was in the presence of swa, in which case swa resistance and R33 competence were retained. PrP(Sc) associated with swa-resistant/R33-competent and swa-sensitive/R33-incompetent prions had different conformational stabilities. When cloned R33-incompetent/swa-sensitive prions were again propagated in brain, their properties gradually reverted to those of the original brain-derived 22L prions. Our results support the view that 22L prion populations are heterogeneous and that distinct prion variants are selected in different cellular environments.