Rhie Gi-Eun, Jung Hae-Mi, Park Jungchan, Kim Bong-Su, Mekalanos John J
Division of High-risk Pathogen Research, Center for Infectious Diseases, National Institute of Health, Seoul, Korea.
FEMS Immunol Med Microbiol. 2008 Jan;52(1):23-8. doi: 10.1111/j.1574-695X.2007.00346.x. Epub 2007 Dec 7.
The most widely used oral whole-cell-recombinant B subunit cholera vaccine contains the nontoxic cholera toxin B subunit (CTXB) and either heat- or formalin-killed Vibrio cholerae O1 strains. Vibrio cholerae O1 strains in the vaccine provide antibacterial immunity, and CTXB contributes to the vaccine's efficacy by stimulating production of anti-CTXB antibody. Various attempts have been made to increase CTXB production. In this study, the mariner-FRT transposon delivery system developed by Chiang and Mekalanos was used to place the ctxB gene under the control of a strong chromosomal promoter in a nontoxigenic V. cholerae El Tor strain, M7922. The expression level of CTXB in transposon insertion mutant clones was screened by ganglioside-dependent enzyme-linked immunosorbent assay. Among CTXB-producing V. cholerae clones that were isolated, M7922-C1 produced the highest amount of CTXB (3.17+/-1.69 microg mL(-1)). M7922-C1 harbors a single insertion of ctxB into VC0972, which encodes a putative porin protein. Although the level of CTXB expression in this strain was not exceptionally high, this study indicates the possibility of using this delivery system to construct vaccine strains that overexpress specific antigens.
使用最广泛的口服全细胞重组B亚单位霍乱疫苗包含无毒霍乱毒素B亚单位（CTXB）以及经热灭活或甲醛灭活的霍乱弧菌O1菌株。疫苗中的霍乱弧菌O1菌株提供抗菌免疫力，而CTXB通过刺激抗CTXB抗体的产生来提高疫苗效力。人们已进行了各种尝试来增加CTXB的产量。在本研究中，采用Chiang和Mekalanos开发的水手-FRT转座子递送系统，将ctxB基因置于非产毒霍乱弧菌El Tor菌株M7922中一个强染色体启动子的控制之下。通过神经节苷脂依赖性酶联免疫吸附测定筛选转座子插入突变体克隆中CTXB的表达水平。在分离得到的产生CTXB的霍乱弧菌克隆中，M7922-C1产生的CTXB量最高（3.17±1.69微克/毫升）。M7922-C1在编码一种假定孔蛋白的VC0972中存在单个ctxB插入。尽管该菌株中CTXB的表达水平并非特别高，但本研究表明使用这种递送系统构建过表达特定抗原的疫苗菌株的可能性。
