FIG 6.

Protection efficacy of Ms, NSs, and M/NSs PR8 viruses in mice. Six- to 8-week-old female C57BL/6 mice (n = 6) were mock vaccinated (with PBS) or vaccinated intranasally with the indicated MLD₅₀ (0.01 or 0.1) of the Ms or Ms/NSs PR8 virus or PR8 ts LAIV. At 2 weeks postvaccination, mice were challenged with 1,000 MLD₅₀ of WT PR8 virus. The replication of WT PR8 virus in the lungs of challenged mice was evaluated at days 2 and 4 postinfection using the immunofocus assay, and titers are given as the number of FFU per milliliter. Symbols represent data for individual mice (n = 3). Bars, geometric mean lung viral titers; dashed line, the limit of detection (200 FFU/ml). Since the MLD₅₀ of the Ms/NSs virus was higher than 10⁵ FFU, 10⁵ FFU was defined to be 0.1 MLD₅₀. Likewise, 10⁴ FFU was defined to be 0.01 MLD₅₀ for the Ms/NSs PR8 virus. *, P < 0.05 using Student's t test from Microsoft Excel software. For the mice vaccinated with 0.01 MLD₅₀ of the Ms/NSs virus, we detected the presence of WT PR8 virus at day 2 postchallenge in only two out of the three mice. Likewise, we detected WT PR8 virus in only one out of the three mice vaccinated with 0.1 MLD₅₀ of PR8 ts LAIV at day 2 postchallenge.