In vitro encystation and excystation of Blastocystis ratti

Cysts of Blastocystis ratti were produced in vitro by culturing the parasit e in Iscove's modified Dulbecco's medium (IMDM) with increasing concentrati ons of horse serum. Yields up to 3 x 10(6) cysts/ml of culture medium were obtained after 72 h. Encystation efficiency was time, strain and inoculum s ize dependent. A viability of > 70 % was determined by flow cytometry emplo ying fluorescein diacetate and propidium iodide staining. The presence of c hitin as a cyst wall component was demonstrated by Calcofluor White M2R sta ining with which cystic stages showed blue fluorescence. The changes in mor phology during excystation were examined by transmission electron microscop y. The cyst enlarged in size and some vacuoles appeared within the condense d cytoplasm. The vacuoles were full of inclusions and small glycogen aggreg ates. Coalescence of the vacuoles led to central body formation. Glycogen d eposits were prominent throughout the excystation process. Some cysts divid ed by binary fission before the completion of the excystation.