A SIMPLE AND RELIABLE METHOD OF PRODUCING IN-VITRO INFECTIONS OF CRYPTOSPORIDIUM-PARVUM (APICOMPLEXA)

A variety of techniques have been used to infect cell monolayers in cu lture with the protozoan, Cryptosporidium parvum. However, most of the se methods rely on the use of trypsin and/or bile salts to excyst spor ozoites in vitro, followed by washing sporozoites free of excystation solution prior to their addition to subconfluent monolayers. This meth od not only increases the amount of time required to establish infecti ons in vitro, but also results in prolonged exposure of free sporozoit es to environmental conditions. Here we report a simple, fast, and eff icient method of obtaining consistent infections of C. paruum in cell monolayers. This technique relies on the ability of the parasite to ex cyst at 37 degrees C but not at room temperature following pretreatmen t with sodium hypochlorite. By adding surface-sterilized oocysts direc tly to monolayers, sporozoites have access to host cells immediately u pon excystation.