Cultivating the endogenous life cycle of Eimeria tenella in chicken intestinal organoids.

Coccidiosis, caused by Eimeria spp., leads to substantial economic losses in the poultry industry globally. These protozoan parasites invade the intestinal epithelium of birds, impairing nutrient absorption, causing diarrhoea and potentially leading to mortality. The complex endogenous life cycle of Eimeria spp., particularly the gametogony phase, presents significant challenges for in vitro cultivation. This study aimed to develop mature chicken intestinal organoids as an in vitro model capable of supporting the complete endogenous life cycle of Eimeria tenella. Two commercially available culture media, 3dGRO L-WRN conditioned medium (L-WRN) and IntestiCult™ Organoid Growth Medium (OGM), were evaluated for their ability to support chicken intestinal organoid development. The results demonstrated that basolateral-out organoids embedded in Matrigel and cultured in the L-WRN medium expanded more rapidly. In contrast, those apical-out organoids in the OGM developed more microvilli structures on enterocytes. Apical-out organoids, initially cultured in L-WRN medium and subsequently matured in OGM, were selected as the optimal host for the Eimeria infection model. Sporozoites of E. tenella successfully invaded the organoids and progressed through both the schizogony and gametogony phases. Moreover, the parasites produced a new generation of oocysts in this study. The presence of schizonts, gametocytes, and sporulated oocysts confirmed that the model can support the full endogenous life cycle of the parasite in vitro. This organoid-based infection model serves as a promising platform for studying host-pathogen interactions and developing novel interventions to control avian coccidiosis.