Characterization of mobile genetic elements from clostridium difficile and development of molecular tools for genetic manipulations

艰难梭菌目前被认为是北美和欧洲医院感染的最重要病原体之一，尤其是在老年人中。C.艰难梭菌主要依赖于两种毒素（TcdA和TcdB）以及在环境中形成孢子和持续存在的能力。本研究对一株C. difficile揭示了几个移动的遗传元件，包括一个质粒和两个完整的原噬菌体。因此，水平基因转移很可能是该病原体进化的重要机制。然而，C.艰难梭菌的研究很少，根据它们在其他病原体毒力中的作用，需要更好地表征它们并评估它们对C.很难此外，目前还存在着用于C. difficile仍然非常有限，这阻碍了进一步的基础研究。我们的长期目标是表征来自C.艰难梭菌，并开发其遗传操作的分子工具，以深入了解这种细菌的生态，多样性，毒力和进化。将通过分子技术对约1，000种菌株进行分类，并在微生物和分子水平（包括全基因组测序）鉴定和表征这些菌株基因组中的前噬菌体。 还将对质粒进行分离、测序和充分表征。原噬菌体和质粒在C.艰难梭菌的收集将被确定，这将有望为我们提供线索，他们的作用和影响C。艰难的生态学和进化。最后，这些质粒将用于开发分子工具（例如载体）来操纵C。difficile基因组这项研究将首次提供大量C.艰难菌株我们还将获得有关C的基本数据。艰难梭菌质粒及其潜在功能。在这些移动的遗传元件中有望发现新的毒力因子或适应因子，这将有助于我们更好地了解C.很难

Clostridium difficile is now considered as one of the most important pathogen responsible for nosocomial infections in North America and Europe, especially among the elderly. Virulence of C. difficile relies mainly on two toxins (TcdA and TcdB) and on the ability to sporulate and persist in the environment. The recent genome sequencing from a virulent strain of C. difficile revealed several mobile genetic elements, including a plasmid and two complete prophages. Thus, horizontal gene transfer is most likely an important mechanism for the evolution of this pathogen. However, prophages and plasmids of C. difficile have only been poorly studied so far and according to their role in virulence of other pathogens, there is a need to better characterize them and assess their impact on the biology and evolution of C. difficile. In addition, efficient molecular tools for the genetic manipulation of C. difficile remain very limited, which blocks further fundamental studies. Our long-term objective is to characterize prophages and plasmids from C. difficile and to develop molecular tools for its genetic manipulation in order to get insight into the ecology, diversity, virulence and evolution of this bacterium. A collection of ~1,000 strains will be classified by molecular techniques, and prophages within the genome of these strains will be identified and characterized at the microbiological and molecular level, including whole genome sequencing. Plasmids will also be isolated, sequenced and fully characterized. The distribution of prophages and plasmids across our C. difficile collection will be determined, which will hopefully give us clues about their role and impact on C. difficile ecology and evolution. Finally, these plasmids will be used to develop molecular tools (e.g. vectors) to manipulate the C. difficile genome. For the first time, this study will provide data about prophage diversity in a large collection of C. difficile strains. We will also gain fundamental data regarding C. difficile plasmids and their potential function(s). New virulence or fitness factors are expected to be found within these mobile genetic elements, which could help us better understand the microbial diversity of C. difficile.