Toxoplasma gondii: Bradyzoite Differentiation In Vitro and In Vivo
Toxoplasma gondii, a member of the Apicomplexa, is known for its ability to infect an impressive range of host species. It is a common human infection that causes significant morbidity in congenitally infected children and immunocompromised patients. This parasite can be transmitted by bradyzoites, a slowly replicating life stage found within intracellular tissue cysts, and oocysts, the sexual life cycle stage that develops in domestic cats and other Felidae. T. gondii bradyzoites retain the capacity to revert back to the quickly replicating tachyzoite life stage, and when the host is immune compromised unrestricted replication can lead to significant tissue destruction. Bradyzoites are refractory to currently available Toxoplasma treatments. Improving our understanding of bradyzoite biology is critical for the development of therapeutic strategies to eliminate latent infection. This chapter describes a commonly used protocol for the differentiation of T. gondii tachyzoites into bradyzoites using human foreskin fibroblast cultures and a CO2-limited alkaline cell media, which results in a high proportion of differentiated bradyzoites for further study. Also described are methods for purifying tissue cysts from chronically infected mouse brain using isopycnic centrifugation and a recently developed approach for measuring bradyzoite viability.
Key wordsToxoplasma Bradyzoite Tissue cyst Differentiation Stress In vitro In vivo
This work was supported by 1F31AI136401 (J.M.), R01AI134753 (L.M.W.), and R21AI123495 (L.M.W.).
- 18.Tomita T, Bzik DJ, Ma YF, Fox BA, Markillie LM, Taylor RC, Kim K, Weiss LM (2013) The Toxoplasma gondii cyst wall protein CST1 is critical for cyst wall integrity and promotes bradyzoite persistence, e1003823. PLoS Pathog 9Google Scholar
- 38.Tobin C, Pollard A, Knoll L (2010) Toxoplasma gondii cyst wall formation in activated bone marrow-derived macrophages and bradyzoite conditions. J Vis Exp 12(42):2091Google Scholar