Vaccines That Reduce Viral Shedding Do Not Prevent Transmission of H1N1 Pandemic 2009 Swine Influenza A Virus Infection to Unvaccinated Pigs

Abstract

Swine influenza A virus (swIAV) infection causes substantial economic loss and disease burden in humans and animals. The 2009 pandemic H1N1 (pH1N1) influenza A virus is now endemic in both populations. In this study, we evaluated the efficacy of different vaccines in reducing nasal shedding in pigs following pH1N1 virus challenge. We also assessed transmission from immunized and challenged pigs to naive, directly in-contact pigs. Pigs were immunized with either adjuvanted, whole inactivated virus (WIV) vaccines or virus-vectored (ChAdOx1 and MVA) vaccines expressing either the homologous or heterologous influenza A virus hemagglutinin (HA) glycoprotein, as well as an influenza virus pseudotype (S-FLU) vaccine expressing heterologous HA. Only two vaccines containing homologous HA, which also induced high hemagglutination inhibitory antibody titers, significantly reduced virus shedding in challenged animals. Nevertheless, virus transmission from challenged to naive, in-contact animals occurred in all groups, although it was delayed in groups of vaccinated animals with reduced virus shedding.IMPORTANCE This study was designed to determine whether vaccination of pigs with conventional WIV or virus-vectored vaccines reduces pH1N1 swine influenza A virus shedding following challenge and can prevent transmission to naive in-contact animals. Even when viral shedding was significantly reduced following challenge, infection was transmissible to susceptible cohoused recipients. This knowledge is important to inform disease surveillance and control strategies and to determine the vaccine coverage required in a population, thereby defining disease moderation or herd protection. WIV or virus-vectored vaccines homologous to the challenge strain significantly reduced virus shedding from directly infected pigs, but vaccination did not completely prevent transmission to cohoused naive pigs.

Keywords: influenza A; pH1N1; pig; transmission; vaccine.

FIG 1

Study design. Groups of five pigs were prime-boost vaccinated on days 0 and 21 postvaccination (dpv) while cohoused during the vaccination phase. For the challenge phase, pigs were housed in separate groups and challenged with the pH1N1 strain A/swine/England/1353/2009 10 weeks after the prime vaccination. Pigs were monitored daily postinoculation (dpi) until postmortem (PM) sampling on 6 or 7 dpi. On 2 dpi, five naive unvaccinated contact pigs were introduced into each group and were monitored daily until 12 days postcontact (dpc) at the end of the study. Inoculated and unvaccinated pigs remained in contact for a total of 4 or 5 days.

FIG 2

Viral shedding. In each group (A to H), viral RNA shedding following challenge was assessed daily in nasal swabs by RRT-qPCR and is expressed as mean log₁₀ relative equivalent units REU. The mean REU (± SEM) is shown for the vaccinated and challenged pigs in each group (solid lines), as well as the individual shedding profiles for the naive pigs in these groups (dotted lines). Horizontal lines under the x axis denote the contact period.

FIG 3

Statistical modeling. Nasal shedding of viral RNA (A to C) from vaccinated, inoculated pigs (red) and naive contact pigs (blue) was modeled for all groups as follows: G1, Cont; G2, WIV_hom; G3, WIV_het; G4, S-FLU; G5, Cont_Ad/MVA; G6, Ad_hom/MVA_hom; G7, MVA_hom; and G8, MVA_het. For each parameter, circles represent the individual value, the gray squares show the mean, and the black error bars indicate the mean ± SD. Analysis is shown for (A) the area under the curve (AUC), which provides a measure of total viral RNA shedding, (B) the generation time (T_g), which is the time interval between the onset of virus shedding in a primary case and in its secondary case, and (C) the latent period, which is the interval between inoculation or contact and first detection of nasal shedding of viral RNA. The relationship between the latent period and the exposure to virus (D) is shown for each treatment group, with virus exposure computed as the sum of the AUC in log scale for the vaccinated pigs in a given treatment group.

FIG 4

Humoral immune response during the vaccination phase. Longitudinal serum samples were assessed by influenza A virus nucleoprotein (NP) competition ELISA, hemagglutination inhibition (HI), and virus neutralization (VN) assays. (A) Anti-NP antibody response is expressed as the mean (± SEM) of the inverse of the percentage of competition: 1 − (OD_sample/OD_negative) × 100%. Results below the 50% threshold (gray shading) are considered negative. Antibody levels in the WIV_hom-, WIV_het-, S-Flu-, and Ad_hom/MVA_hom-immunized groups above the 50% threshold are significantly higher (P < 0.0001) than those in the control groups (Cont and Cont_Ad/MVA). HI and VN titers against (B and D) the homologous antigen, pH1N1 (A/swine/England/1353/2009), and (C and E) the heterologous antigen, H1_avN1 (A/swine/England/453/2006), are shown as the geometric mean ratios (GMRs ± geometric SD) for the titers for each pig relative to the corresponding 0-dpv sample. **, P < 0.001, and ***, P < 0.0001, by the color corresponding to the group, as indicated under “Legend.”

FIG 5

Humoral immune response after challenge infection with pH1N1. Serum samples from 6 days postinfection (6 dpi) were assessed by influenza A virus nucleoprotein (NP) competition ELISA, Hemagglutination inhibition (HI) and virus neutralization (VN) assays. (A) Antibody response to NP is expressed as the inverse of the percentage of competition: 1 − (OD_sample/OD_negative) × 100%. Results over 50% are considered positive. Individual HI and VN titers against (B and D) the homologous antigen, pH1N1 (A/swine/England/1353/2009), and (C and E) the heterologous antigen, H1_avN1 (A/swine/England/453/2006), are shown relative to their corresponding 0-dpv sample. *, P < 0.05, and ***, P < 0.0001, significant difference from the control group, by the color corresponding to the group, as indicated under “Legend.”

FIG 6

Cellular immune response. Shown is the mean number (± SD) of IFN-γ producing PBMCs (spot-forming cells [SFC]/million cells) induced by (A) inactivated pH1N1 A/swine/England/1353/2009 antigen, (B) NP, or (C) M1 18-mer peptides for the virus-vectored vaccine groups. Responses were evaluated 1 week after boost vaccination (28 dpv [solid circles]) and after viral challenge (6 dpi [open circles]). (D) Comparison of SFC induced by either inactivated homologous antigen (pH1N1 [blue bars]) or heterologous antigen (H1_avN1 [red bars]) 2 weeks after boost. *, P < 0.05, significant difference from the control group.