Guo Shanguang, Yan Weiwei, McDonough Sean P, Lin Nengfeng, Wu Katherine J, He Hongxuan, Xiang Hua, Yang Maosheng, Moreira Maira Aparecida S, Chang Yung-Fu
College of Veterinary Medicine, Cornell University, Ithaca, NY 14853, USA; College of Food Science, South China Agricultural University, Guangzhou 510642, China.
College of Veterinary Medicine, Cornell University, Ithaca, NY 14853, USA.
Vaccine. 2015 Mar 24;33(13):1586-95. doi: 10.1016/j.vaccine.2015.02.006. Epub 2015 Feb 16.
Clostridium difficile infection (CDI) causes nosocomial antibiotic-associated diarrhea and colitis in the developed world. Two potent cytotoxins, toxin A (TcdA) and toxin B (TcdB) are the virulence factors of this disease and can be a good vaccine candidate against CDI. In the present study, we genetically engineered Lactococcus lactis to express the nontoxic, recombinant fragments derived from TcdA and TcdB C-terminal receptor binding domains (Tcd-AC and Tcd-BC) as an oral vaccine candidate. The immunogenicity of the genetically engineered L. lactis oral vaccine delivery system (animal groups LAC and LBC or the combination of both, LACBC) was compared with the recombinant TcdA and TcdB C-terminal receptor binding domain proteins (animal groups PAC and PBC or the combination of both, PACBC), which were expressed and purified from E. coli. After the C. difficile challenge, the control groups received PBS or engineered L. lactis with empty vector, showed severe diarrhea symptoms and died within 2-3 days. However, both the oral vaccine and recombinant protein vaccine groups had significantly lower mortalities, body weight decreases and histopathologic lesions than the control sham-vaccine groups (p<0.05) except group LBC which only had a 31% survival rate after the challenge. The data of post infection survival showed that an average of 86% of animals survived in groups PAC and PACBC, 75% of animals survived in group LACBC, and 65% of animals survived in group LAC. All of the vaccinated animals produced higher titers of both IgG and IgA than the control groups (p<0.05), and the antibodies were able to neutralize the cytopathic effect of toxins in vitro. The results of this study indicate that there is a potential to use L. lactis as a delivery system to develop a cost effective oral vaccine against CDI.
艰难梭菌感染（CDI）在发达国家会引发医院内抗生素相关性腹泻和结肠炎。两种强效细胞毒素，毒素A（TcdA）和毒素B（TcdB）是该疾病的致病因子，可成为抗CDI的优良疫苗候选物。在本研究中，我们对乳酸乳球菌进行基因工程改造，使其表达源自TcdA和TcdB C端受体结合域的无毒重组片段（Tcd-AC和Tcd-BC），作为口服疫苗候选物。将基因工程改造的乳酸乳球菌口服疫苗递送系统（动物组LAC和LBC或两者组合，LACBC）的免疫原性与从大肠杆菌中表达并纯化的重组TcdA和TcdB C端受体结合域蛋白（动物组PAC和PBC或两者组合，PACBC）进行比较。在艰难梭菌攻击后，对照组接受PBS或带有空载体的工程化乳酸乳球菌，出现严重腹泻症状并在2 - 3天内死亡。然而，口服疫苗组和重组蛋白疫苗组的死亡率、体重下降和组织病理学损伤均显著低于对照假疫苗组（p<0.05），但LBC组在攻击后仅有31%的存活率。感染后存活数据显示，PAC组和PACBC组平均86%的动物存活，LACBC组75%的动物存活，LAC组65%的动物存活。所有接种疫苗的动物产生的IgG和IgA滴度均高于对照组（p<0.05），且这些抗体能够在体外中和毒素的细胞病变效应。本研究结果表明，有潜力将乳酸乳球菌用作递送系统来开发一种经济有效的抗CDI口服疫苗。
