DNA Vaccination Elicits Protective Immune Responses against Pandemic and Classic Swine Influenza Viruses in Pigs

A pseudotype lentiviral inhibition assay confirms anti-H1N1 neutralizing antibody responses in pooled sera. Neutralizing antibody responses against A/California/2009 and A/Ohio/2007 were confirmed by a pseudotype inhibition assay performed on pooled sera from each treatment group, with 10 pigs/group. Neutralizing antibody titers are presented as log₁₀ transformations of half-maximal inhibitory concentrations (IC₅₀). Data points are plotted for each treatment group with the same symbols used in Fig. 1.

DNA vaccination elicits IFN-γ responses measured by an ELISpot assay with PBMCs collected 1 week prior to H1N1 challenge. An ELISpot assay was used to detect cells secreting IFN-γ against A/California/2009 and A/Ohio/2007 in vaccinated animals. Results are presented as single data points in a group, with lines indicating the mean and standard error of the mean (SEM) for each group. A Mann-Whitney t test was used for statistical analysis comparing values between immunized and nonimmunized control animals. P values of less than 0.05 are indicated: * represents a P value between 0.05 and 0.001, while ** indicates a P value of ≤0.001. θ indicates a value that is nonsignificant when Bonferroni's adjustment is applied.

DNA vaccination reduces viral load and protects against H1N1 influenza virus challenge. Immunized and control animals were challenged with either H1N1 A/California/2009 or A/Ohio/2007. Postchallenge viral loads were assessed for up to 7 days using a plaque reduction assay. Viral titers are presented as the log of the TCID₅₀ for each animal, with bars indicating means and error bars indicating SEMs. Differences between immunized animals and controls were analyzed for statistical significance using a Mann-Whitney t test. P values of less than 0.05 are indicated: * represents a P value between 0.05 and 0.001, while ** indicates a value of ≤0.001. θ indicates a value that is nonsignificant when Bonferroni's adjustment is applied.

DNA vaccination elicits protection from lung disease induced by A/California/2009 challenge and prevents viral replication in the lung. Lungs were collected from 5 pigs per group at 5 dpc with A/California/2009. Hematoxylin and eosin staining was performed to assess histopathology, and immunohistochemistry analysis was performed to detect influenza virus antigens in the lung. Micrographs are representative for each treatment group (including monovalent NS and NF vaccine groups) at a magnification of ×20. See Results for lesion description. In panel A, the epithelium lining the bronchiole is irregular (arrow), and prominent peribronchiolar lymphocyte infiltration (*) is evident. In the control IHC photomicrograph in panel E, virus antigen in bronchiolar epithelial cells (arrow) appears as brown coloration of the nucleus and cytoplasm.