Abstract：To construct Lactobacillus heat labile enterotoxin B subunit (LTB) expressing vector,and study the mucosal immune protecting effect of LTB on Clostridium difficile (C. difficile) toxoid vaccine, LTB gene was cloned and pSIP411-LT recombinant plasmid was prepared. The Lactobacillus LTB expressing vector was prepared by recombinant plasmid electrotransforming to Lactobacillus. After A toxin (TcdA) and B toxin (TcdB) of C. difficile were purified by dialysis and chromatography, the toxoid of TcdA and TcdB were obtained by formaldehyde treatment for vaccine candidate antigen. On 0 day, 14 days and 28 days, the mice were subcutaneously inoculated with Tcd toxoid and oral lactobacillus expressing LT adjuvant. On 0 day, 10 days, 24 days and 38 days of first inoculation, serum IgG and fecal IgA of anti-A-toxin and B-toxin were detected. On 38 days, the mice were infected by C. difficile by mouth, and the protective effect of the vaccine was observed. Adjuvant expressing vector of LTB Lactobacillus was constructed, and toxoid of TcdA and TcdB was prepared successfully. After infection with C. difficile, no diarrhea or weight loss occurred in all vaccine groups, while the diarrhea incidence was 100%, the mortality incidence was 42.86%, and the weight loss of 5 days was 0.83±0.11g in the control group. On the 10 days, 24 days and 38 days after first inoculation, the serum IgG and fecal IgA levels of anti-TcdA or TcdB in each vaccine group were significantly higher than those in the blank group (P<0.01), and the serum IgG and fecal IgA levels in the vaccine + LTB adjuvant group were significantly higher than those in the vaccine group (P<0.05). In conclusion, as vaccine candidate antigens, toxoid of TcdA and TcdB has good protective effect on infection of C. difficile. The adjuvant LTB expressing of Lactobacillus can improve antibody titer and enhance mucous membrane immunity in immune animals.
2019, Vol. 35 
