Improved excystation protocol for Eimeria nieschulzi (Apikomplexa, Coccidia)

Abstract

Large numbers of sporozoites are a crucial prerequisite for in vitro experiments with Eimeria species. There are no protocols to obtain high amounts of vital purified sporozoites of Eimeria nieschulzi; therefore, an improved excystation protocol is urgently needed. Most excystation procedures for Eimeria oocysts use a mechanical disruption method for the release of sporocysts, assuming that oocyst disruption of Eimeria does not require enzymes (proteases). However, rodent Eimeria oocysts are susceptible to pepsin digestion (Kowalik S, Zahner H (1999) Eimeria separata: method for the excystation of sporozoites. Parasitol Res 85:496-499). Here, we describe a method that combines enzymatic treatment of oocyst walls before the mechanical disruption with glass beads. Using this protocol, we achieved an up to fivefold increase of free viable sporozoites of E. nieschulzi and could significantly shorten the time of excystation. These results confirm the assumption that rodent Eimeria species, in contrast to Eimeria species of birds, possess protease sensitive oocysts.