Isolation of viable Toxoplasma gondii cysts from brain samples for oral infection

V.R. Puvanesuaran, N. Ibrahim, R. Noordin, V. Balakrishnan

Institute for Research in Molecular Medicine (INFORMM), Universiti Sains Malaysia, 11800 USM, Pulau Pinang, Malaysia. e-mail: venuinformm@gmail.com

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• Infectious Diseases

AIM: A method was developed to separate contaminant-free viable Toxoplasma gondii cysts from brain samples of infected mice for molecular biology studies and reinfection.

MATERIALS AND METHODS: The mice brains were homogenized and washed with phosphate buffered saline (PBS) – Tween 80 prior to fractionation using 19-22% dextran solution. Finally, the supernatant was purified by two-step membrane filtration (100-160 microm and < 10 microm) to obtain pure T. gondii cyst. The isolates were analyzed through microscopic observation, qPCR and by reinfection of new batch of mice.

RESULTS: T. gondii cysts were best isolated with 21% dextran solution and two step filtration.

CONCLUSIONS: The method was observed not to disrupt the integrity of the cysts containing bradyzoites. In addition, the isolated cysts in the filtrate were found to be contaminant-free, viable and able to infect healthy mice when introduced orally; which, mimics the natural infectivity pathway.

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