Chick embryo tracheal organ: a new and effective in vitro culture model for Cryptosporidium baileyi

Abstract

In the present study, chick embryo tracheal organ (TOCs) was used to cultivate oocysts or sporozoites of Cryptosporidium baileyi. Approximately 5 × 10(4) sporozoites and oocysts mixture for group I; 5 × 10(5), 1 × 10(6), 2 × 10(6) purified sporozoites for group II, group III and group IV, respectively, were inoculated into respective chick embryo tracheal rings maintained in RPMI 1640 supplemented with 5% heat-inactivated FBS, and cultivated in each well of the 24-well culture plate at 40°C and 5% CO(2). The tracheal rings in four experimental groups (I-IV) were successfully infected with C. baileyi, and different stages of parasites were also observed under light and electron microscopy. Parasite infection and cytological alterations were noted as early as PI 72 h. The Cryptosporidium were seen attached to the edge of the tracheal epithelium, with more number of parasites in group I than that in group II, group III and group IV. The moderate nuclear swelling and chromatin margination were also detected, and the normal vertical orientation and basilar location of the nuclei of the epithelial cells were almost lost. C. baileyi that has been passed by TOCs exhibited similar immunity and molecular features with parasites before intratracheal inoculation. These results suggest that chick embryo tracheal organ is a new and effective in vitro culture model for C. baileyi and other respiratory pathogens.