Exogenous nitric oxide stimulates early egress of Eimeria tenella sporozoites from primary chicken kidney cells in vitro

Egress plays a vital role in the life cycle of apicomplexan parasites including Eimeria tenella, which has been attracting attention from various research groups. Many recent studies have focused on early egress induced by immune molecules to develop a new method of apicomplexan parasite elimination. In this study, we investigated whether nitric oxide (NO), an immune molecule produced by different types of cells in response to cytokine stimulation, could induce early egress of eimerian sporozoites in vitro. Eimeria tenella sporozoites were extracted and cultured in primary chicken kidney cells. The number of sporozoites egressed from infected cells was analyzed by flow cytometry after treatment with NO released by sodium nitroferricyanide (II) dihydrate. The results showed that exogenous NO stimulated the rapid egress of E. tenella sporozoites from primary chicken kidney cells before replication of the parasite. We also found that egress was dependent on intra-parasitic calcium ion (Ca2+) levels and no damage occurred to host cells after egress. The virulence of egressed sporozoites was significantly lower than that of fresh sporozoites. The results of this study contribute to a novel field examining the interactions between apicomplexan parasites and their host cells, as well as that of the clearance of intracellular pathogens by the host immune system.

The V617I Substitution in Avian Coronavirus IBV Spike Protein Plays a Crucial Role in Adaptation to Primary Chicken Kidney Cells

The naturally isolated avian coronavirus infectious bronchitis virus (IBV) generally cannot replicate in chicken kidney (CK) cells. To explore the molecular mechanism of IBV adapting to CK cells, a series of recombinant viruses were constructed by chimerizing the S genes of CK cell-adapted strain H120 and non-adapted strain IBYZ. The results showed that the S2 subunit determines the difference in cell tropism of the two strains. After comparing the amino acid sequences of S protein of CK cell-adapted strain YZ120, with its parental strain IBYZ, three amino acid substitutions, A138V, L581F, and V617I, were identified. Using YZ120 as the backbone, one or more of the above-mentioned substitutions were eliminated to verify the correlation between these sites and CK cell tropism. The results showed that the CK cell tropism of the YZ120 strain depends on the V617I substitution, the change of L581F promoted the adaptation in CK cells, and the change at 138 position was not directly related to the CK cell tropism. Further validation experiments also showed that V617I had a decisive role in the adaptation of IBV to CK cells, but other areas of the virus genome also affected the replication efficiency of the virus in CK cells.