In vitro culture and synchronous release of Sarcocystis neurona merozoitesfrom host cells

The growth of Sarcocystis neurona, isolate UCD1, in continuous culture was examined in 10 cell lines to identify growth conditions and methods for the preparation of parasites free of gross host cell contamination for molecul ar studies. The unpredictable, slow release of merozoites in most cell line s prompted development of a method to synchronously release the parasites f rom infected host cells. The calcium ionophore A23187 at a concentration of 1 muM was found to release intracellular merozoites with a 40 min treatmen t at 37 degreesC. The release of merozoites en masse from attached host cel ls allowed for the rapid collection of relatively pure parasites from the c ulture supernatant. This release of merozoites occurred in five different h ost cell lines, The ionophore-released parasites were highly infectious for host calls and appeared to be morphologically identical to naturally relea sed merozoites, except that the treated merozoites had an increased number of micronemes when examined by electron microscopy. The ionophore did not e nhance the release of sporozoites from sporocysts, but freezing in the pres ence of 5% DMSO released sporozoites that were infectious to bovine monocyt es in in vitro culture. (C) 2001 Elsevier Science B.V. All rights reserved.