Induced protection from a CCHFV-M DNA vaccine requires CD8+ T cells

Abstract

Crimean-Congo hemorrhagic fever (CCHF) is a World Health Organization prioritized disease because its broad distribution and severity of disease make it a global health threat. Despite advancements in preclinical vaccine development for CCHF virus (CCHFV), including multiple platforms targeting multiple antigens, a clear definition of the adaptive immune correlates of protection is lacking. Levels of neutralizing antibodies in vaccinated animal models do not necessarily correlate with protection, suggesting that cellular immunity, such as CD8⁺ T cells, might have an important role in protection in this model. Using a well-established IFN-I antibody blockade mouse model (IS) and a DNA-based vaccine encoding the CCHFV M-segment glycoprotein precursor, we investigated the role of humoral and T cell immunity in vaccine-mediated protection in mice genetically devoid of these immune compartments. We found that in the absence of the B-cell compartment (µMT knockout mice), protection provided by the vaccine was not reduced. In contrast, in the absence of CD8⁺ T cells (CD8⁺ knockout mice) the vaccine-mediated protection was significantly diminished. Importantly, humoral responses to the vaccine in CD8⁺ T-cell knockout mice were equivalent to wild-type mice. These findings indicated that CD8⁺ T-cell responses are necessary and sufficient to promote protection in mice vaccinated with the M-segment DNA vaccine. Identifying a crucial role of the cellular immunity to protect against CCHFV should help guide the development of CCHFV-targeting vaccines.

Keywords: CD8+ T cells; Correlates of protection; Crimean-Congo hemorrhagic fever; DNA vaccine; Glycoproteins; M-segment.

Fig. 1

Immune response in CCHFV-MAfg-09 vaccinated T-cell and B-cell knockout IS mice. Groups of 15 mice were vaccinated three times with 50 µg of CCHFV-M_Afg-09 or empty vector by IM-EP at three-week intervals. The splenocyte T-cell response of five mice per group was analyzed by ELISpot (B6 empty group n = 4, all other groups n = 5). Splenocytes from individual mice were restimulated with two pools of peptides derived from the CCHFV strain Afg09-2990 M-segment. Anti-CCHFV-M specific (A) IL-2+and (B) IFN-γ+ T cells were quantified by ELISpot. Data are the group mean averages ± SD. *p < 0.05; **p < 0.01; ***p < 0.001; ***p < 0.0001. P-values were determined by two-way ANOVA with Sidak's multiple comparison test with a 95% confidence interval. (C) anti-G_C antibody responses of vaccinated mice to be challenged was measured in the sera at three weeks after the final vaccination by Magpix.

Fig. 2

CCHFV challenge of vaccinated T- and B-cell deficient mice. Groups of 8-10 mice were challenged with 100 PFU of CCHFV strain Afg09-2990, µMT empty (n = 10), µMT CCHFV-M_Afg09 (n = 9), TCRα/β^−/− empty (n = 10), TCRα/β^−/− CCHFV-M_Afg09 (n = 9), B6 empty (n = 8), and B6 CCHFV-M_Afg09 (n = 10). Survival and weight change from baseline on day 0. ***p = 0.0003 and ****P < 0.0001, comparison of CCHFV-M_Afg09 to empty vector groups for each mouse species by log-rank test.

Fig. 3

Humoral immune responses in CD8^−/− and B6 wild-type mice. The anti-GP38 and anti-G_C antibody responses of vaccinated mice to be challenged was measured in the sera at three weeks after final vaccination by MAGPIX.

Fig. 4

Survival and antibody response of CCHFV-M_Afg-09 vaccinated CD8⁺ T-cells IS mice. Groups of 10 mice were vaccinated three times with 50 µg of CCHFV-M_Afg-09 or empty vector by IM-EP at three-week intervals. Groups of 8–10 mice were challenged with 100 PFU of CCHFV strain Afg09-2990. Survival and weight change from baseline on day 0. **p < 0.01, comparison of CCHFV-M_Afg09 to empty vector groups for each mouse species by log-rank test.