| Autor: |
Cheng Z; College of Veterinary Medicine, Southwest University, Chongqing 402460, China., Huang H; College of Veterinary Medicine, Southwest University, Chongqing 402460, China., Cao S; College of Veterinary Medicine, Southwest University, Chongqing 402460, China., Shi H; College of Veterinary Medicine, Southwest University, Chongqing 402460, China., Gao J; College of Veterinary Medicine, Southwest University, Chongqing 402460, China., Li J; College of Veterinary Medicine, Southwest University, Chongqing 402460, China. |
| Abstrakt: |
To construct a recombinant Bacillus subtilis strain expressing SpaA and CbpB of Erysipelothrix rhusiopathiae for oral administration, we constructed the recombinant plasmid pDG1730-CBJA by fusion PCR and seamless cloning. The plasmid was introduced into B . subtilis KC strain by natural transformation, and the recombinant strain KC- spaA - cbpB was screened out on the plate containing spectinomycin ( spe r ) and confirmed by PCR and starch degradation test. The SpaA and CbpB expressed by KC- spaA - cbpB were detected by Western blotting and indirect immunofluorescence assay, and the genetic stability of the recombinant strain in mice was determined. The plasmid pMAD-∆ spe r with knockout of spe r was constructed and transformed into KC- spaA-cbpB . The spe r -deleted mutant strain KC- spaA - cbpB : : ∆ spe r was screened and identified, and its immunogenicity in a mouse model was evaluated by oral immunization. The results showed that the recombinant strain KC- spaA - cbpB was stable in mice, expressing SpaA on the cell surface and CbpB on the spore surface. KC- spaA - cbpB : : ∆ spe r expressed SpaA and CbpB. The mice vaccinated with the spores of KC- spaA - cbpB : : ∆ spe r had higher levels of SpaA and CbpB-specific IgG in the serum that those vaccinated with the wild-type spores 42 days after vaccination by gavage ( P < 0.01). The protective rate of mice immunized with the recombinant spores was 67.5%. The results indicated that a recombinant B . subtilis strain expressing SpaA and CbpB of E . rhusiopathiae was successfully constructed, and the recombinant strain laid a foundation for the development of oral live vector vaccines for swine erysipelas. |
