Figure 1. Experimental design. Procedures followed for each group of chickens included in the study.

Chickens were divided in two groups; the first consisted of 46 chickens that were inoculated with 10⁶ ELD₅₀ of H7N1 A/Chicken/Italy/5093/99 HPAI virus, while the second group of 18 chickens were used as non-infected control. In turn, the inoculated chickens were divided in three groups (A, B, and C), as well the non-infected chickens that were divided en two groups. Group A consists of 18 H7N1 inoculated chickens and 6 non-infected chickens that were perfused with EB at 6, 12, 18, 24, 36 and 48 hpi and their brains were frozen. Group B includes 16 H7N1 inoculated and 12 non-infected chickens that were perfused with PBS and their brains were frozen. Group C contains 12 chickens inoculated with H7N1 and which brains were fixed in formalin.