Table 1.
Sequence variation in vaccine and challenge virus epitopes.
Table 2.
Peptides used in this study.
Figure 1.
Anti-NP serum antibody titers.
Anti-NP IgG levels in the serum of immunized mice at various timepoints following immunization. BALB/c (A) or B6 (B) mice were immunized with 1×1010 TCID50 of NP+M2-rAd or B/NP rAd, 2×105 TCID50 of A/Alaska ca+A/Hong Kong ca (Cold-adapted) or B/AA ca, or left unvaccinated. Serum was obtained at 3 weeks (solid bars) or 5 weeks (open bars) post-vaccination and tested for anti-NP IgG as described in Materials and Methods. Shown are the mean endpoint titers ±SD; n = 5 per group. * P≤0.05 versus naïve, † P≤0.05 versus ca; One Way ANOVA with Student-Newman-Keuls post-test.
Figure 2.
Anti-M2e serum antibody titers.
Anti-M2e IgG levels in the serum of immunized mice at various timepoints following immunization. BALB/c (A and B) mice were immunized as in Figure 1 or left unvaccinated. Serum was obtained at 3 weeks (A) or 5 weeks (B) post-vaccination and tested for anti-M2e IgG antibody. Anti-M2e titers were determined using the PR8 peptide sequence (solid bars) and the ma-CA/04 peptide sequence (open bars). Shown are the mean endpoint titers ±SD; n = 5 per group. * P≤0.0001 versus naïve, † P≤0.05 versus ca; One Way ANOVA with Student-Newman-Keuls post-test.
Figure 3.
IFN-γ responses of lung T cells following stimulation with peptides of vaccine sequences or challenge virus sequences. BALB/c (A) or B6 (B) mice were immunized as in Figure 1, or left unvaccinated. Three weeks after immunization, T cell responses were determined by IFN-γ ELISPOT as described in Materials and Methods. Pooled lung cells were tested using cells from 5 (BALB/c) or 7 (B6) mice per group. Shown are the mean of triplicate measurements of IFN-γ secreting cells per million ±SD.
Figure 4.
Vaccine effectiveness in protection against challenge with ma-CA/04. Groups of BALB/c (A and B) or B6 (C and D) mice were immunized as in Figure 1, or left unvaccinated. Three weeks after immunization, animals were challenged with 5 MLD50 of ma-CA/04 intranasally and monitored for survival (A and C) and weight loss (B and D). Weight loss graph shows average of n = 9 (BALB/c) or n = 12 (B6) mice ±SD. * P≤0.0001 versus naïve; log rank test (A and C); or P≤0.01 versus naïve; two-way repeated measures ANOVA with Holm-Sidak post-test (B and D). No statistically significant differences were found between NP+M2-rAd and ca vaccine.
Figure 5.
Viral replication in the lungs following ma-CA/04 challenge. BALB/c (A) or B6 (B) mice were immunized as in Figure 1, or left unvaccinated. Three weeks after immunization, animals were challenged with 5 MLD50 of ma-CA/04 intranasally. Lung viral titers were determined on day 3 (solid bars) and day 6 (open bars) by TCID50 assay as described in Material and Methods. Shown are the mean ±SD; n = 4 per group. * P≤0.05 versus naïve; Kruskal-Wallis test with Student-Newman-Keuls post test. No statistically significant differences were found between NP+M2-rAd and ca vaccine.