Guan S, Verma N K
Division of Biochemistry and Molecular Biology, Faculty of Science, Australian National University, Canberra, Australia.
FEMS Microbiol Lett. 1998 Sep 1;166(1):79-87. doi: 10.1111/j.1574-6968.1998.tb13186.x.
Shigella flexneri SFL124 (serotype Y) is a promising live oral vaccine candidate, which has been shown to be safe and immunogenic in human volunteers. To change the serotype of this vaccine strain, we inserted a serotype conversion gene cluster into the chromosome of SFL124 by using a bacteriophage-based site-specific integration system. By cloning an integrase gene (int), an attachment site (attP) and a glucosyl transfer gene cluster from bacteriophage SfX into a suicide vector, and subsequently introducing this construct into S. flexneri SFL124, we obtained a S. flexneri strain (designated SFL1213) expressing the serotype X somatic antigen specificity. The strain retained other characteristics of the parent strain, such as colony shape, growth rate, and Congo red binding property. Stability test showed that the serotype X O-antigen specificity in SFL1213 was 100% stable after being cultured approximately 72 successive hours under non-selective condition. In a mouse pulmonary model, the recombinant strain elicited a significant level of humoral antibodies which recognized the lipopolysaccharide (LPS) of a wild-type S. flexneri serotype X strain. The site-specific insertion system will be useful when stable expression of a cloned single copy gene is desired in the chromosome of S. flexneri vaccine candidate, SFL124.
福氏志贺菌SFL124（血清型Y）是一种很有前景的口服活疫苗候选株，已在人类志愿者中显示出安全性和免疫原性。为了改变该疫苗株的血清型，我们使用基于噬菌体的位点特异性整合系统将血清型转换基因簇插入SFL124的染色体中。通过将来自噬菌体SfX的整合酶基因（int）、附着位点（attP）和糖基转移基因簇克隆到自杀载体中，随后将该构建体导入福氏志贺菌SFL124，我们获得了表达血清型X菌体抗原特异性的福氏志贺菌菌株（命名为SFL1213）。该菌株保留了亲本菌株的其他特征，如菌落形态、生长速率和刚果红结合特性。稳定性测试表明，SFL1213中的血清型X O抗原特异性在非选择性条件下连续培养约72小时后100%稳定。在小鼠肺部模型中，重组菌株引发了高水平的体液抗体，这些抗体识别野生型福氏志贺菌血清型X菌株的脂多糖（LPS）。当希望在福氏志贺菌疫苗候选株SFL124的染色体中稳定表达克隆的单拷贝基因时，位点特异性插入系统将很有用。
