Rydzewski Kerstin, Tlapák Hana, Niehaus Indra P, Dabrowski Piotr W, Grunow Roland, Heuner Klaus
Cellular Interactions of Bacterial Pathogens, Centre for Biological Threats and Special Pathogens, ZBS 2, Robert Koch Institute, Berlin, Germany.
Centre for Biological Threats and Special Pathogens, Highly Pathogenic Viruses, ZBS 1, Robert Koch Institute, Berlin, Germany.
Int J Med Microbiol. 2015 Dec;305(8):874-80. doi: 10.1016/j.ijmm.2015.08.037. Epub 2015 Sep 8.
Recently, we identified a putative prophage on a genomic island (GI) within the genome sequence of Francisella hispaniensis isolate AS0-814 (Francisella tularensis subsp. novicida-like 3523) by the analysis of the CRISPR-Cas systems of Francisella. Various spacer DNAs within the CRISPR region of different F. tularensis subsp. novicida strains were found to be homologous to the putative prophage (Schunder et al., 2013, Int. J. Med. Microbiol. 303:51-60). Now we identified the GI (FhaGI-1) as a mobile element which is able to form a circular episomal structure. The circular episomal form of FhaGI-1 is generated by F. hispaniensis, and the excision of the island is an integrase-dependent and site-specific process. Furthermore, we could demonstrate that the excision of the island is also possible in other bacterial species (Escherichia coli). In addition, we could show that a genetically generated small variant of the island is also functional and, after its electroporation into strain F. tularensis subsp. holarctica LVS, the GI was stable and site-specifically integrated into the genome of the transformants. The integrase is sufficient for the integration and excision of the small variant into and from the DNA backbone, respectively. Thus, the element may be suitable to be used as a genetic tool in F. tularensis research. Furthermore, we identified the tRNA(Val) gene of Francisella as an integration site for GIs. Genomic island FphGI-1 was identified in Francisella philomiragia ATCC 25016. We were not able to detect the episomal form of this GI, probably due to a mutated attR site. However, we could demonstrate that integrative GIs are present in Francisella and that they may allow horizontal gene transfer between different Francisella species.
最近，通过对土拉弗朗西斯菌的CRISPR-Cas系统进行分析，我们在西班牙弗朗西斯菌分离株AS0-814（类土拉弗朗西斯菌亚种新凶手亚种3523）的基因组序列中的一个基因组岛（GI）上鉴定出一个假定的前噬菌体。不同土拉弗朗西斯菌亚种新凶手亚种菌株的CRISPR区域内的各种间隔DNA被发现与假定的前噬菌体同源（Schunder等人，2013年，《国际医学微生物学杂志》303:51-60）。现在我们将该GI（FhaGI-1）鉴定为一种能够形成环状附加体结构的移动元件。FhaGI-1的环状附加体形式由西班牙弗朗西斯菌产生，该岛的切除是一个依赖整合酶且位点特异性的过程。此外，我们能够证明该岛在其他细菌物种（大肠杆菌）中也可以切除。另外，我们可以表明该岛的一个基因产生的小变体也是有功能的，并且在将其电穿孔导入土拉弗朗西斯菌亚种全北区LVS菌株后，该GI是稳定的并且位点特异性地整合到转化体的基因组中。整合酶分别足以将小变体整合到DNA主链中以及从DNA主链中切除。因此，该元件可能适合用作土拉弗朗西斯菌研究中的遗传工具。此外，我们将弗朗西斯菌的tRNA(Val)基因鉴定为GIs的整合位点。在嗜肺弗朗西斯菌ATCC 25016中鉴定出基因组岛FphGI-1。我们无法检测到该GI的附加体形式，可能是由于attR位点发生了突变。然而，我们能够证明整合性GIs存在于弗朗西斯菌中，并且它们可能允许不同弗朗西斯菌物种之间进行水平基因转移。
