Neuraminidase (NA) activity assay
NA activity assays of F/98 virus and recombinant virus rF/HAS145N were analyzed using a Neuraminidase Assay Kit (Beyotime Institute of Biotechnology, Jiangsu, China) as previously described (Qin et al., 2018). Each virus was calibrated with the same TCID50 titer by diluting the virus sample. Then, 10 μL was mixed with 70 μL detection buffer, 10 μL NA fluorogenic substrate, and 10 μL double-distilled water. Fluorescence was monitored using a multifunctional microplate reader (Tecan). NA activities are shown as the fluorescence intensity above the background values for allantoic fluid without virus.