Ectopic Expression of O Antigen in Bordetella pertussis by a Novel Genomic Integration System.
Ectopic Expression of O Antigen in Bordetella pertussis by a Novel Genomic Integration System.
复制标题
新型基因组整合系统中o抗原的异位表达。
DOI：
10.1128/msphere.00417-17
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发表时间：
2018-01
期刊：
影响因子：
4.8
通讯作者：
Horiguchi Y
中科院分区：
文献类型：
--
作者：
Ishigaki K;Shinzawa N;Nishikawa S;Suzuki K;Fukui-Miyazaki A;Horiguchi Y
一些细菌表型是通过位于一个大的遗传基因座内的许多基因的协同功能而出现的。为了将一种细菌的表型转移到另一种细菌，需要将大的遗传基因座引入受体细菌的手段。因此，我们开发了一个新的系统，结合细菌人工染色体载体和噬菌体衍生的基因整合机制的优点。在这项研究中，我们首次成功地将参与支气管败血波氏杆菌O抗原生物合成的基因位点引入到本质上缺乏O抗原的B.百日咳的染色体中，并使用该系统分析了所得到的B.百日咳突变株的表型改变。目前的结果表明，该系统成功地实现了上述目的。我们认为该系统适用于除博德特氏菌外的许多细菌。我们描述了一种新的基因组整合系统，使大至50 kbp的DNA片段导入受体细菌的染色体。该系统，命名为BPI，包括细菌人工染色体载体和噬菌体衍生的基因整合机制。我们使用BPI系统将O抗原生物合成所需的支气管败血波氏杆菌的wbm基因座引入本质上缺乏O抗原的B.百日咳的染色体中。在引入wbm基因座后，B百日咳杆菌在脂寡糖组分中呈现了一种额外的物质，该物质可被抗支气管炎B杆菌抗体特异性识别，但不被抗B抗体特异性识别。百日咳抗体，表明B。百日咳杆菌表达O抗原，与支气管炎B杆菌的O抗原相对应。O抗原表达B。百日咳杆菌对血清和多粘菌素B的杀菌作用的敏感性低于同基因亲本菌株。此外，体内竞争性感染试验表明，O抗原表达型B.百日咳菌在小鼠呼吸道的定殖优势超过了亲本菌株。这些结果表明，BPI系统提供了一种通过引入大的外源DNA片段来改变细菌表型的手段。一些细菌的表型是通过位于一个大的遗传位点内的许多基因的协同功能而出现的。为了将一种细菌的表型转移到另一种细菌，需要一种将大遗传基因座引入受体细菌的方法。因此，我们开发了一个新的系统，结合细菌人工染色体载体和噬菌体衍生的基因整合机制的优点。在这项研究中，我们首次成功地将参与支气管败血波氏杆菌O抗原生物合成的基因位点引入到本质上缺乏O抗原的B.百日咳的染色体中，并使用该系统分析了所得到的B.百日咳突变株的表型改变。目前的结果表明，该系统成功地实现了上述目的。我们认为该系统适用于除博德特氏菌外的许多细菌。
Some bacterial phenotypes emerge through the cooperative functions of a number of genes residing within a large genetic locus. To transfer the phenotype of one bacterium to another, a means to introduce the large genetic locus into the recipient bacterium is needed. Therefore, we developed a novel system by combining the advantages of a bacterial artificial chromosome vector and phage-derived gene integration machinery. In this study, we succeeded for the first time in introducing a gene locus involved in O antigen biosynthesis of Bordetella bronchiseptica into the chromosome of B. pertussis, which intrinsically lacks O antigen, and using this system we analyzed phenotypic alterations in the resultant mutant strain of B. pertussis. The present results demonstrate that this system successfully accomplished the above-described purpose. We consider this system to be applicable to a number of bacteria other than Bordetella. We describe a novel genome integration system that enables the introduction of DNA fragments as large as 50 kbp into the chromosomes of recipient bacteria. This system, named BPI, comprises a bacterial artificial chromosome vector and phage-derived gene integration machinery. We introduced the wbm locus of Bordetella bronchiseptica, which is required for O antigen biosynthesis, into the chromosome of B. pertussis, which intrinsically lacks O antigen, using the BPI system. After the introduction of the wbm locus, B. pertussis presented an additional substance in the lipooligosaccharide fraction that was specifically recognized by the anti-B. bronchiseptica antibody but not the anti-B. pertussis antibody, indicating that B. pertussis expressed O antigen corresponding to that of B. bronchiseptica. O antigen-expressing B. pertussis was less sensitive to the bactericidal effects of serum and polymyxin B than the isogenic parental strain. In addition, an in vivo competitive infection assay showed that O antigen-expressing B. pertussis dominantly colonized the mouse respiratory tract over the parental strain. These results indicate that the BPI system provides a means to alter the phenotypes of bacteria by introducing large exogenous DNA fragments. IMPORTANCE Some bacterial phenotypes emerge through the cooperative functions of a number of genes residing within a large genetic locus. To transfer the phenotype of one bacterium to another, a means to introduce the large genetic locus into the recipient bacterium is needed. Therefore, we developed a novel system by combining the advantages of a bacterial artificial chromosome vector and phage-derived gene integration machinery. In this study, we succeeded for the first time in introducing a gene locus involved in O antigen biosynthesis of Bordetella bronchiseptica into the chromosome of B. pertussis, which intrinsically lacks O antigen, and using this system we analyzed phenotypic alterations in the resultant mutant strain of B. pertussis. The present results demonstrate that this system successfully accomplished the above-described purpose. We consider this system to be applicable to a number of bacteria other than Bordetella.