Abstract
This paper describes a standard method for in vitro excystation of sporocysts of S. capracanis, S. hircicanis, S. ovicanis and S. arieticanis. A standard pretreatment was performed by 20 minutes preincubation of sporocysts in aqueous sodium hypochlorite (NaOCl) solution of 6% or 8% at room temperature. Then, sporocysts were washed 5 times in distilled water and were incubated for one hour at 39℃ in excystation fluid consisting of RPMI 1640 medium, 10% FCS and 15% bovine bile. Additional sonication of pretreated sporocysts increased excystation rates as compared to not sonicated controls. Excystation rates for 1-9 months old sporocysts were 77% for S. capracanis, 77% for S. hircicanis, 72% for S. arieticanis and 92% for S. ovicanis. Sporozoite suspension as obtained by this method proved to be decontaminated and sporozoites were viable and able to invade Vero cells in tissue culture. Sporozoites were also subjected to cryopreservation in RPMI 1640 containing 10% FCS and 7.5% DMSO. After storage in liquid nitrogen, sporozoites proved to be infective for their intermediate hosts after intraperitoneal injection.
