Disruption of the Toxoplasma gondii bradyzoite-specific gene BAG1 decreases in vivo cyst formation

The bradyzoite stage of the Apicomplexan protozoan parasite Toxoplasma gond ii plays a critical role in maintenance of latent infection. We reported pr eviously the cloning of a bradyzoite-specific gene BAG1/hsp30 (previously r eferred to as BAG5) encoding a cytoplasmic antigen related to small heat sh ock proteins, We have now disrupted BAG1 in the T, gondii PLK strain by hom ologous recombination. H7, a cloned null mutant, and Y8, a control positive for both cat and BAG1, were chosen for further characterization. Immunoflu orescence and Western blot analysis of bradyzoites with BAG1 antisera demon strated expression of BAG1 in the Y8 and the PLK strain but no expression i n H7, All three strains expressed a 116 kDa bradyzoite cyst wall antigen, a 29 kDa matrix antigen and the 65 kDa matrix reactive antigen MAG1, Mice in oculated with H7 parasites formed significantly fewer cysts than those inoc ulated with the Y8 and the PLK strains. H7 parasites were complemented with BAG1 using phleomycin selection, Cyst formation in vivo for the BAG1-compl emented H7 parasites was similar to wild-type parasites, We therefore concl ude that BAG1 is not essential for cyst formation, but facilitates formatio n of cysts in vivo.