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. 2017 Jun 24;13(1):194.
doi: 10.1186/s12917-017-1115-3.

Immunogenicity and protective efficacy of a Salmonella Enteritidis sptP mutant as a live attenuated vaccine candidate

Zhijie Lin  1 , Peipei Tang  1 , Yang Jiao  1 , Xilong Kang  1 , Qiuchun Li  1 , Xiulong Xu  1   2   3 , Jun Sun  1   4 , Zhiming Pan  5 , Xinan Jiao  6
Affiliations

Immunogenicity and protective efficacy of a Salmonella Enteritidis sptP mutant as a live attenuated vaccine candidate

"VSports app下载" Zhijie Lin et al. BMC Vet Res. .

Abstract

Background: Salmonella enterica serovar Enteritidis (S. Enteritidis) is a highly adaptive pathogen in both humans and animals VSports手机版. As a Salmonella Type III secretion system (T3SS) effector, Salmonella protein tyrosine phosphatase (SptP) is critical for virulence in this genus. To investigate the feasibility of using C50336ΔsptP as a live attenuated oral vaccine in mice, we generated the sptP gene deletion mutant C50336ΔsptP in S. Enteritidis strain C50336 by λ-Red mediated recombination and evaluated the protective ability of the S. Enteritidis sptP mutant strain C50336ΔsptP against mice salmonellosis. .

Results: We found that C50336ΔsptP was a highly immunogenic, effective, and safe vaccine in mice. Compared to wild-type C50336, C50336ΔsptP showed reduced virulence as confirmed by the 50% lethal dose (LD50) in orally infected mice. C50336ΔsptP also showed decreased bacterial colonization both in vivo and in vitro V体育安卓版. Immunization with C50336ΔsptP had no significant effect on body weight and did not result in obvious clinical symptoms relative to control animals treated with phosphate-buffered saline (PBS), but induced humoral and cellular immune responses at 12 and 26 days post inoculation. Immunization with 1 × 108 colony-forming units (CFU) C50336ΔsptP per mouse provided 100% protection against subsequent challenge with the wild-type C50336 strain, and immunized mice showed mild and temporary clinical symptoms as compared to those of control group. .

Conclusions: These results demonstrate that C50336ΔsptP can be a live attenuated oral vaccine for salmonellosis V体育ios版. .

Keywords: Immune protection; Immunogenicity; Salmonella Enteritidis; SptP; Vaccine VSports最新版本. .

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Figures

Fig. 1
Fig. 1
Construction of sptP deletion mutant in S. Enteritidis strain C50336. a PCR verification of C50336ΔsptP and C50336-CmR. The wild type strain harbors the complete sptP gene, with a PCR product length of 2509 bp, whereas the PCR product from C50336ΔsptP has a length of 1061 bp, and the product from the C50336-CmR has a length of 1991 bp (right). b Growth curves of wild-type S. Enteritidis C50336 and C50336ΔsptP. Bacteria were grown in liquid LB medium at 37 °C for 12 h with agitation, and the OD600 values of triplicate cultures in LB medium were determined in 1-h intervals
Fig. 2
Fig. 2
Bacterial colonization in cells. Salmonella adhesion and invasion in the Human epithelial Caco-2 BBE cells (a), and mouse macrophage RAW264.7 cells (b). The number of Salmonella was determined and presented as 102 CFU/mL. Data are expressed as the mean ± SEM. **P ≤ 0.01
Fig. 3
Fig. 3
Body weight (a) and bacterial colonization in mouse organs post immunization. Salmonella colonization and persistence in the liver (b), spleen (c), and mLN (d) of mice following oral immunization with 1 × 107 CFU or 1 × 108 CFU of C50336ΔsptP. Control mice received 100 μl of PBS and were negative for Salmonella in the liver, spleen, and mLN. The number of Salmonella was determined and represented as Log10 CFU/g
Fig. 4
Fig. 4
Immunogenicity of C50336ΔsptP. Procedure for immunization (a). Determination of serum IgG levels (b) and IgG subtype levels (c) at 12 and 26 days post immunization. Stimulation index (SI) of mice lymphocytes determined by splenic lymphocyte proliferation assay using soluble antigen SEAgP (d) or lymphocyte mitogen ConA as a positive control. The vaccinated group received 1 × 107 CFU or 1 × 108 CFU of C50336ΔsptP orally, and the control group received 100 μl of PBS. Data are expressed as the mean ± SEM. *P ≤ 0.05, **P ≤ 0.01
Fig. 5
Fig. 5
Survival curve (a) and bacterial colonization in challenged mice. Bacterial colonization in the liver (b), spleen (c) and cecum (d) of challenged mice (Table 2, group A, immunized with C50336ΔsptP and then challenged with 1 × 108 CFU of C50336; group B, immunized with PBS and then challenged with 5 × 105 CFU of C50336). All samples from the blank control group were negative for Salmonella (Table 2, group C). The number of bacteria was determined and expressed as log10 CFU/g. Data are expressed as the mean ± SEM, **P ≤ 0.01
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