FIG 3.

The permissiveness of porcine 3D airway organoids to PRCoV infection. (A) The morphological changes of PRCoV-infected or mock-infected 3D AOs derived from 2-day-old piglets were observed under a bright-field microscope at 24 hpi and 48 hpi. Scale bars = 200 μm. (B) Detection of PRCoV infection in porcine 3D airway organoids by transmission electron microscopy (TEM). Porcine 3D AOs mock-inoculated or inoculated with PRCoV were harvested at 24 hpi, and then the 3D AOs were fixed in extracellular matrix (ECM) with 2.5% glutaraldehyde and analyzed by H-7650 electron microscopy (Hitachi, Japan). The red arrow points to cilia, the black arrow points to the vacuole, and the white arrows point to PRCoV particles. Scale bars = 500 nm. (C) Monitoring PRCoV infection in 3D AOs derived from a 2-day-old piglet (D2) by IFA. 3D AOs were infected with PRCoV at an MOI of 5, and the levels of infection at 24 hpi were determined by anti-NP IFA staining (scale bar = 20 μm). (D) The kinetics of PRCoV replication in 3D AOs. 3D AOs were infected with PRCoV at an MOI of 5, and samples were collected at the indicated time points for PRCoV viral load detection by RT-qPCR compared with mock AOs. Error bars denote the deviations indicated by *: *, P < 0.05; **, P < 0.01; ***; P < 0.001.