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. 2024 Jan 5:14:1310333.
doi: 10.3389/fmicb.2023.1310333. eCollection 2023.

A vesicular stomatitis virus-based African swine fever vaccine prototype effectively induced robust immune responses in mice following a single-dose immunization

Yunyun Ma  1   2 , Junjun Shao  1 , Wei Liu  1 , Shandian Gao  1 , Decai Peng  1 , Chun Miao  1 , Sicheng Yang  1 , Zhuo Hou  1 , Guangqing Zhou  1 , Xuefeng Qi  2 , Huiyun Chang  1
Affiliations

A vesicular stomatitis virus-based African swine fever vaccine prototype effectively induced robust immune responses in mice following a single-dose immunization

Yunyun Ma et al. Front Microbiol. .

Erratum in

Abstract

Introduction: African swine fever (ASF) is a highly contagious hemorrhagic fever disease in pigs caused by African swine fever virus (ASFV). It is very difficult to control and prevent ASF outbreaks due to the absence of safe and effective vaccines.

Methods: In order to develop a safe and effective ASF vaccine for the control and prevention of ASF, two ASFV recombinant vesicular stomatitis virus (VSV) live vector vaccine prototypes, containing the gene of p72, and a chimera of p30 and p54, were developed based on the replication-competent VSV, and named VSV-p72 and VSV-p35. The immune potency of VSV-p72 or VSV-p35 alone and in combination was evaluated in BALB/c mice via intramuscular and intranasal vaccination.

Results: The results indicated that whether administered alone or in combination, the two vaccine prototypes showed acceptable safety in mice and, more importantly, induced high-level specific antibodies against p72, p30, and p54 of ASFV and a strong cellular immune response 28 days after vaccination. The sera from mice vaccinated with the vaccine prototypes significantly inhibited ASFV from infecting porcine alveolar macrophages (PAMs) in vitro. Most notably, the immunized sera from a mixture of VSV-p35 and VSV-p72 inhibited ASFV from infecting PAMs, with an inhibition rate of up to 78.58%.

Conclusion: Overall, our findings suggest that ASFV recombinant VSV live vector vaccine prototypes may become a promising candidate vaccine for the control and prevention of ASF.

Keywords: African swine fever virus; immune potency; safety; vaccine prototypes; vesicular stomatitis virus.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Construction of the recombinant viruses (VSV-p72 and VSV-p35). (A) The schematic representation of the recombinant viruses. (B) The identification of recombinant plasmids with Xho I and Nhe I restriction enzymes, 15 kb in size was the pVSVXN2-GFP vector fragment, and the other was the target gene fragment, 1980 bp and 1,233 bp in size, respectively. 1, 3, 5: the enzyme digestion plasmid; 2, 4, 6: the uncleaved plasmid. (C) PCR amplification of the target genes. (D) The target proteins expression of VSV-p35 and VSV-p72 in infected BHK-21 cells. (E) The morphology of the recombinant viruses under transmission electron microscopy; the scale bar was equivalent to 200 nm.
Figure 2
Figure 2
Genetic stability of recombinant viruses. (A) The growth properties of VSV-p35 and VSV-p72 were inoculated into BHK-21 cells at an MOI of 0.001. (B,C) RT-PCR amplification results in cell supernatants by serially passaging the recombinant viruses for 20 generations. (D) The results of IFA analysis on target proteins extracted from BHK-21 cells infected with VSV-p35 and VSV-p72 at the 20th passage.
Figure 3
Figure 3
Safety evaluation of recombinant viruses in BALB/c mice. (A) The survival rate of mice that had been inoculated with recombinant viruses. (B) The body weight change of mice that had been inoculated with recombinant viruses. (C,D) The relative expression of the VSV N gene in blood, heart, liver, spleen, lung, and kidney samples of BALB/c mice inoculated with recombinant viruses, respectively.
Figure 4
Figure 4
HE staining to analyze the histopathologic changes in the hearts, livers, spleens, lungs and kidneys of BALB/c mice inoculated with recombinant viruses at 28 dpi. VSV-rwt and PBS served as control groups (bar = 100 μm, 200×). Black arrow: central venous congestion; blue arrow: a small amount of granulocyte infiltration; HE, hematoxylin–eosin.
Figure 5
Figure 5
Immunohistochemical detection of VSV G protein in the hearts, livers, spleens, lungs, and kidneys of BALB/c mice inoculated with recombinant viruses at 28 dpi. VSV-rwt and PBS served as control groups (bar = 100 μm, 200×).
Figure 6
Figure 6
Blood routine analysis in mice that had been inoculated with recombinant viruses. (A) The total white blood cell count (WBC). (B–D) The percentage of neutrophils (NEUT), monocytes (MO), and lymphocytes (LY), respectively. (E–G) The total number of neutrophils, monocytes, and lymphocytes in turn.
Figure 7
Figure 7
Evaluation of immunogenicity in mice immunized with different doses of recombinant viruses. (A–C) The levels of IgG to p30, p54, and p72. (D) The proliferation of T lymphocytes from immunized mice stimulated with different doses (1 × 105, 2.5 × 105, and 5 × 105 TCID50/mL) of the recombinant viruses. The data are presented as the means ± SD. Statistical significance is denoted by ns = p > 0.05, *p < 0.05, **p < 0.01, and ***p < 0.001.
Figure 8
Figure 8
The antibody levels in sera. (A–C) The specific IgG levels against ASFV to p30, p54, and p72 in each group. (D,E) The levels of IgG1 and IgG2a in sera, respectively. (F) The calculated ratio of IgG2a/IgG1. The data are presented as the means ± SD. Statistical significance is denoted by ns = p > 0.05, *p < 0.05, **p < 0.01, and ***p < 0.001.
Figure 9
Figure 9
The cytokine secretion and T lymphocyte proliferation in spleen lymphocytes from mice immunized with the recombinant viruses or PBS. (A–D) The IL-2, IL-10, IFN-γ, and TNF-α levels secreted in spleen lymphocytes from i.m or i.n immunized mice. (E,F) The proliferation of T lymphocytes from i.m or i.n immunized mice stimulated by different doses (2.5 × 104, 1 × 105, 4 × 105 HAD50/well) of heat-inactivated ASFV. The data are presented as the means ± SD. Statistical significance is denoted by ns = p > 0.05, *p < 0.05, **p < 0.01, ***p < 0.001.
Figure 10
Figure 10
Spleen lymphocyte typing of immunized mice. (A,B) The percentage of CD4+ and CD8+ T in spleen lymphocytes from i.m or i.n immunized mice, respectively. (C,D) The statistical results of CD4+ and CD8+ T in spleen lymphocytes. The data are presented as the means ± SD. Statistical significance is denoted by ns = p > 0.05, *p < 0.05, **p < 0.01, ***p < 0.001.
Figure 11
Figure 11
Intracellular cytokine expression in spleen lymphocytes from immunized mice stimulated by inactivated ASFV or PBS. (A,C) The percentage and statistical results of IL-2+ CD8+ T cells in CD8+ T cells, respectively. (B,D) The percentage and statistical results of IFN-γ+ CD8+ T cells in CD8+ T cells. The data are presented as the means ± SD. Statistical significance is denoted by ns = p > 0.05, *p < 0.05, **p < 0.01, ***p < 0.001.
Figure 12
Figure 12
The ability of sera from immunized BALB/c mice to neutralize ASFV infection in vitro. ASFV CN/SC/19 virus strain pre-incubated with pre-immune sera as a control or immune sera collected at 28 dpi after vaccination was used to infect PAM cells. The copy number of the B646L gene was determined by qPCR to measure the neutralization efficiency of the recombinant viruses after being infected for 72 h. (A,B) The copy number of the B646L gene and neutralization rate of immunized BALB/c mice sera at different dilutions after incubation with ASFV. (C) The neutralization rate of immunized BALB/c mice sera at 1:8 diluting after incubation with ASFV.
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