Cysts of Blastocystis ratti were produced in vitro
by culturing the parasite in Iscove's modified Dulbecco's medium
(IMDM) with increasing concentrations of horse serum. Yields up to 3×106
cysts/ml of culture medium were obtained
after 72 h. Encystation efficiency was time, strain and inoculum size dependent.
A viability of >70% was determined by
flow cytometry employing fluorescein diacetate and propidium iodide staining.
The presence of chitin as a cyst wall
component was demonstrated by Calcofluor White M2R staining with which
cystic stages showed blue fluorescence. The
changes in morphology during excystation were examined by transmission
electron microscopy. The cyst enlarged in size
and some vacuoles appeared within the condensed cytoplasm. The vacuoles
were full of inclusions and small glycogen
aggregates. Coalescence of the vacuoles led to central body formation.
Glycogen deposits were prominent throughout the
excystation process. Some cysts divided by binary fission before the completion
of the excystation.