Two IBV, 2575/98 and 2296/95, were isolated in Taiwan and maintained using serial passage through specific pathogen-free (SPF) eggs [29] . IBV were inoculated into the embryonic eggs at the E9 stage and harvested at the E11 stage from chorioallantoic fluid. The original viral titers of 2575/98 and 2296/ 95 are 10 7.4 EID 50 /0.1 mL and that of 2296 reached 10 9.6 /0.1 mL, respectively [14] . The applied viral titer of both IBV was 1 · 10 5 EID 50 /mL.