Maternal Autogenous Inactivated Virus Vaccination Boosts Immunity to PRRSV in Piglets

Abstract

Maternal-derived immunity is a critical component for the survival and success of offspring in pigs to protect from circulating pathogens such as Type 2 Porcine Reproductive and Respiratory Syndrome Virus (PRRSV-2). The purpose of this study is to investigate the transfer of anti-PRRSV immunity to piglets from gilts that received modified-live virus (MLV) alone (treatment (TRT) 0), or in combination with one of two autogenous inactivated vaccines (AIVs, TRT 1+2). Piglets from these gilts were challenged with the autogenous PRRSV-2 strain at two weeks of age and their adaptive immune response (IR) was evaluated until 4 weeks post inoculation (wpi). The systemic humoral and cellular IR was analyzed in the pre-farrow gilts, and in piglets, pre-inoculation, and at 2 and 4 wpi. Both AIVs partially protected the piglets with reduced lung pathology and increased weight gain; TRT 1 also lowered piglet viremia, best explained by the AIV-induced production of neutralizing antibodies in gilts and their transfer to the piglets. In piglets, pre-inoculation, the main systemic IFN-γ producers were CD21α⁺ B cells. From 0 to 4 wpi, the role of these B cells declined and CD4 T cells became the primary systemic IFN-γ producers. In the lungs, CD8 T cells were the primary and CD4 T cells were the secondary IFN-γ producers, including a novel subset of porcine CD8α^-CCR7^- CD4 T cells, potentially terminally differentiated CD4 T_EMRA cells. In summary, this study demonstrates that maternal AIV vaccination can improve protection of pre-weaning piglets against PRRSV-2; it shows the importance of transferring neutralizing antibodies to piglets, and it introduces two novel immune cell subsets in pigs-IFN-γ producing CD21α⁺ B cells and CD8α^-CCR7^- CD4 T cells.

Keywords: CD4 TEMRA cells; IFN-γ producing B cells; PRRSV; T cells; autogenous inactivated vaccine; cell-mediated immune response; humoral immune response; maternal vaccination; swine; transfer of immunity.

Figure 1

Study design: The study included gilts that were either vaccinated with a modified live vaccine only (MLV, treatment (TRT) 0), or an MLV boosted by a series of four vaccinations using one of two autogenous inactivated virus (AIV) vaccines (TRT 1+2). One week before farrowing, blood from gilts was collected to assess the systemic humoral and cell-mediated adaptive immune response. Piglets from three gilts with high humoral and T-cell responses within each treatment group (TRT 0, 1, 2) were included in the second part of the study—the challenge part at the North Carolina (NC) State Laboratory Animal Resources (LAR). These piglets were intranasally inoculated with the autogenous PRRSV 1-7-4 strain at two weeks of age (0 weeks post inoculation, wpi). Pigs were also bled, as illustrated, to assess the immune response to Porcine Reproductive and Respiratory Syndrome Virus (PRRSV). To assess lung pathology and study the local immune response, half of the pigs were sacrificed at 2 wpi and half at 4 wpi. PBMC: peripheral blood mononuclear cells.

Figure 2

Maternal AIV vaccination boosts immunity to PRRSV-2 in 2-week old piglets: Viremia (A) and weekly body weight gains (B) were analyzed at different weeks post inoculation (wpi). For both, data were analyzed by mixed-effects model (REML) with Dunnett’s multiple comparisons test. (C) Percentage of macroscopic lung lesions, and (D) lung interstitial pneumonia histology scores at 2 weeks post inoculation. Data in (C) and (D) were analyzed by one-way ANOVA with Dunnett’s multiple comparison. All differences are shown within time point and compared to the TRT 0 control group – ** p < 0.01 and * p < 0.05.

Figure 3

Autogenous inactivated boost vaccinations induce homologous neutralizing antibodies that are transferred to piglets. (A) Maternal serum neutralizing antibody (NA) titers to the homologous 1-7-4 strain after vaccination. (B) Piglet serum NA titers to 1-7-4 prior to inoculation (0 weeks post inoculation, wpi) and weekly after inoculation. (C) Pearson correlation matrix for maternal and piglet NA titers to 1-7-4 at -3 and 0 wpi, respectively. Data were analyzed via repeated-measures two-way ANOVA with Dunnett’s multiple comparisons. Significant differences are shown between treatments and within time point; they are designated as ** p < 0.01 and * p < 0.05.

Figure 4

The systemic IFN-γ response to PRRSV 1-7-4 in gilts and their offspring: (A) shows the gating hierarchy starting with a forward/ side scatter (FSC/SSC) gate on lymphocytes, the exclusion of dead cells and doublets to focus the analysis of IFN-γ production on single living lymphocytes (SLLs). Subgates on B cells, NK cells, CD4, CD8 and TCR-γδ T cells were drawn on the whole SLL population (in gray) and overlaid by IFN-γ producing cells (in red). (B) shows the IFN-γ production as a percentage of all SLLs (left column) and the percentage of each immune cell subsets contributing to this IFN-γ production (all other columns). Data were analyzed via one-way ANOVA with Dunnett’s multiple comparisons. Significant differences between treatments are designated as * p < 0.05.

Figure 5

Differentiation of IFN-γ⁺ T-cell subsets in blood: (A) shows the gating used to differentiate CD4 naïve (T_naïve, CD8α⁻CCR7⁺), central memory (T_CM, CD8α⁺CCR7⁺), and effector memory (T_EM, CD8α⁺CCR7⁻) T cells; it also shows the distinction of CCR7⁺ lymph node homing and CCR7⁻ peripheral tissue homing CD8 T cells. Additionally, it illustrates the different TCR-γδ T-cell subsets—naïve CD8α⁻CCR7⁻, and antigen experienced CD8α⁺CCR7⁻ and CD8α⁺CCR7⁺ cells. (B) shows the frequencies of these differentiation and homing defined subsets within IFN-γ producing CD4 T cells (left), CD8 T cells (middle), and TCR-γδ T cells (right) in gilts at one week before farrowing (upper row), piglets at 0 wpi (2nd row), 2 wpi (3rd row) and 4 wpi (4th row). Data were analyzed via one-way ANOVA with Dunnett’s multiple comparisons. No significant differences (p < 0.05) between groups were observed.

Figure 6

The local IFN-γ response to PRRSV 1-7-4 in bronchoalveolar lavage, lungs, and tracheobronchial lymph nodes of piglets. IFN-γ production as a percentage of all SLLs (left column) and the percentage of each immune cell subsets contributing to this IFN-γ production (all other columns) is shown within tracheobronchial lymph nodes (TrBr LN), lung tissue, and bronchoalveolar lavage (BAL) at 2 wpi (A) and 4 wpi (B). Data were analyzed via one-way ANOVA with Dunnett’s multiple comparisons. Significant differences between treatments are designated as * p < 0.05.

Figure 7

Differentiation of IFN-γ⁺ T-cell subsets in tracheobronchial lymph nodes, lung tissue and bronchoalveolar lavage (BAL): These overlays show the CCR7 (x-axis) and CD8α (y-axis) expression of CD4, CD8, and TCR-γδ T-cell subsets (gray) and the respective IFN-γ⁺ T-cell subsets (red) from all animals in tracheobronchial lymph nodes (LN), lung tissue, and BAL at 2 wpi (three left columns) and 4 wpi (three right columns). No statistical analysis was performed.