Rani D B, Bayer M E, Schifferli D M
Department of Pathobiology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Clin Diagn Lab Immunol. 1999 Jan;6(1):30-40. doi: 10.1128/CDLI.6.1.30-40.1999.
The strong immunogenicity of bacterial fimbriae results from their polymeric and proteinaceous nature, and the protective role of these immunogens in experimental or commercial vaccines is associated with their capacity to induce antiadhesive antibodies. Fimbria-mediated intestinal colonization by enteropathogens typically leads to similar antibody responses. The possibility of taking advantage of these properties was investigated by determining whether enteroadhesive fimbriae, like the 987P fimbriae of enterotoxigenic Escherichia coli, can serve as carriers for foreign antigens without losing their adhesive characteristics. Random linker insertion mutagenesis of the fasA gene encoding the major 987P subunit identified five different mutants expressing wild-type levels of fimbriation. The linker insertion sites of these mutants were used to introduce three continuous segments of viral surface glycoproteins known to be accessible to antibodies. These segments encode residues 11 to 19 or 272 to 279 of herpes simplex virus type 1 (HSV-1) glycoprotein D [gD(11-19) and gD(272-279), respectively] or residues 379 to 388 of the transmissible gastroenteritis virus (TGEV) spike protein [S(379-388)]. Studies of bacteria expressing fimbriae incorporating mutated FasA subunits alone or together with wild-type FasA subunits (hybrid fimbriae) indicated that foreign epitopes were best exported and displayed on assembled fimbriae when they were inserted near the amino terminus of FasA. Fimbriated bacteria expressing FasA subunits carrying the HSV gD(11-19) or the TGEV S(379-388) epitope inserted between the second and third residues of mature FasA elicited high levels of foreign epitope antibodies in all rabbits immunized parenterally. Antibodies against the HSV epitope were also shown to recognize the epitope in the context of the whole gD protein. Because the 987P adhesive subunit FasG was shown to be present on mutated fimbriae and to mediate bacterial attachment to porcine intestinal receptors, polymeric display of foreign epitopes on 987P offers new opportunities to test the potential beneficial effect of enteroadhesion for mucosal immunization and protection against various enteric pathogens.
细菌菌毛的强免疫原性源于其聚合和蛋白质性质，这些免疫原在实验性或商业疫苗中的保护作用与其诱导抗黏附抗体的能力相关。肠道病原体通过菌毛介导的肠道定植通常会引发类似的抗体反应。通过确定肠黏附菌毛（如产肠毒素大肠杆菌的987P菌毛）是否可以作为外源抗原的载体而不丧失其黏附特性，来研究利用这些特性的可能性。对编码主要987P亚基的fasA基因进行随机接头插入诱变，鉴定出五个表达野生型菌毛形成水平的不同突变体。这些突变体的接头插入位点用于引入已知可被抗体识别的病毒表面糖蛋白的三个连续片段。这些片段分别编码单纯疱疹病毒1型（HSV-1）糖蛋白D的第11至19位或第272至279位残基[分别为gD(11-19)和gD(272-279)]，或传染性胃肠炎病毒（TGEV）刺突蛋白的第379至388位残基[S(379-388)]。对单独表达含有突变FasA亚基的菌毛或与野生型FasA亚基一起表达（杂交菌毛）的细菌的研究表明，当外源表位插入FasA的氨基末端附近时，它们在组装好的菌毛上的输出和展示效果最佳。表达携带HSV gD(11-19)或TGEV S(379-388)表位的FasA亚基且表位插入成熟FasA的第二个和第三个残基之间的菌毛化细菌，在所有经胃肠外免疫的兔子中引发了高水平的外源表位抗体。还显示针对HSV表位的抗体在整个gD蛋白的背景下也能识别该表位。由于987P黏附亚基FasG在突变菌毛上存在并介导细菌与猪肠道受体的附着，因此在987P上对外源表位进行多聚体展示为测试肠黏附对黏膜免疫和抵御各种肠道病原体的潜在有益作用提供了新机会。
