Histopathological Lung Assessment in Hamsters

Animal handling, hamster infections, NTZ preparation and oral administrations were performed as described above. The anatomo-histological study was implemented as previously described.²⁴ (link)^,26 Briefly, lungs were collected after intratracheal instillation of 4% (w/v) formaldehyde solution, and then fixed 72h at room temperature with a 4% (w/v) formaldehyde solution before being embedded in paraffin. Tissue sections of 3.5µm were stained with hematoxylin-eosin (H&E) and blindly analyzed by a certified veterinary pathologist. Microscopic examination was done using a Nikon Eclipse E400 microscope. Different anatomic compartments were examined (1) for bronchial and alveolar walls, a score of 0 to 4 was assigned based on severity of inflammation; (2) regarding alveoli, a score of 0 to 2 was assigned based on presence and severity of hemorrhagic necrosis; (3) regarding vessel changes (leucocytic accumulation in vascular wall or in endothelial compartment), absence or presence was scored 0 or 1 respectively. A cumulative score was then calculated and assigned to a grade of severity (see Supplementary Table 4).