引用

肉毒杆菌和炭疽芽孢杆菌产生的强效毒素可引起人类严重疾病。这两种病原体都需要新的和改进的疫苗。对于使用乳酸菌的粘膜疫苗输送，抗原的染色体表达优于基于质粒的表达系统，因为染色体表达避免了质粒的不稳定性和抗生素压力的需要。在本研究中，我们构建了三株从染色体上表达(i)肉毒杆菌血清型A神经毒素(BoNT/A- hc)重链的无毒宿主受体结合域(ii)炭疽保护抗原(PA)和(iii) BoNT/A- hc和PA的嗜酸乳杆菌NCFM。BoNT/A- hc疫苗盒被设计成含有嗜酸乳杆菌s层蛋白A的信号肽和树突状细胞靶向肽。选择lba0889下游的一个编码高表达烯醇酶基因的染色体区域插入疫苗盒。Western blot分析证实了质粒和染色体位置的两种抗原的异源表达。稳定性分析表明，在没有抗生素维持的情况下，表达质粒中的疫苗片段丢失，而染色体整合片段保持稳定。RNA测序显示每种抗原的高表达，插入疫苗盒对染色体中其他基因的转录几乎没有影响。本研究表明，当靶向高表达染色体区域时，染色体整合重组株是很有前途的疫苗运输载体。 Levels of expression match high copy plasmids, and eliminate requirements for antibiotic selective maintenance of recombinant plasmids.,Clostridium botulinum and Bacillus anthracis produce potent neurotoxins that pose a biochemical warfare concern, therefore effective vaccines are required against these bacteria. Chromosomal expression of antigens is preferred over plasmid based expression systems since expressing antigens from a chromosomal location confers an advantage to the vaccine strains by eliminating antibiotic maintenance required for plasmids and negates issues with plasmid instability that would result in loss of the antigen. Lactic acid bacteria, including Lactobacillus acidophilus, have shown potential for mucosal vaccine delivery as L. acidophilus is bile and acid tolerant, allowing transit through the gastrointestinal tract where cells interact with host epithelial and immune cells including dendritic cells. In this study we successfully expressed C. botulinum and B. anthracis antigens in the probiotic L. acidophilus NCFM. Both antigens were highly expressed individually or in tandem from the chromosome of L. acidophilus.