Cell-culture system for continuous production of Toxoplasma gondii tachyzoites

The aim of this study was to identify a sustainable cell line and culture m ethod that could continuously provide a sufficient quantity of Toxoplasma g ondii tachyzoites to serve the needs of a general hospital laboratory. Thre e continuous cell lines (HeLa, LLC and Vero) and three cell-culture methods (culture in conventional flasks, culture in membrane-based flasks and an a utomated culture system) were investigated. In multiplicity of-infection an d time-course experiments, HeLa was the cell line of choice. Harvests from HeLa cells had significantly higher tachyzoite yields than those from LLC c ells (P < 0.00005) or Vero cells (P < 0.05). Membrane-based flasks gave hig her yields (6.15 x 10(6) tachyzoites/ml) than conventional flasks (1-2 x 10 (6) tachyzoites/ml) initially, but these were not sustained. The automated cell-culture system was unsuitable for parasite culture. Continuous passage in 25 cm(2) flasks was successful, yielding 1 x 10(6) tachyzoites/ml; viab ility exceeded 90% after 96-120 h of infection throughout 38 passes, during which time the viability improved and the time to harvest became more cons istent. Toxoplasma gondii grown in continuous culture in HeLa cells can pro vide a regular supply of viable tachyzoites. Demonstration that HeLa-derive d tachyzoites could be used for the dye test confirms the potential of this in vitro system for use in general hospital laboratories.