Inducible Expression of spo0A as a Universal Tool for Studying Sporulation in Clostridium difficile.
Inducible Expression of spo0A as a Universal Tool for Studying Sporulation in Clostridium difficile.
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DOI：
10.3389/fmicb.2017.01793
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发表时间：
2017
影响因子：
5.2
通讯作者：
Cutting SM
中科院分区：
文献类型：
--
作者：
Dembek M;Willing SE;Hong HA;Hosseini S;Salgado PS;Cutting SM
艰难梭菌仍然是一种主要的医院病原体，给医疗保健系统带来相当大的压力。形成内生孢子的能力，高度抵抗环境的侮辱，是其持久性和传播的关键。然而，重要的差异之间存在孢子形成途径的C。艰难梭菌和模式革兰氏阳性生物枯草芽孢杆菌。在研究C.艰难梭菌的特征是孢子形成水平相对较差和孢子形成过程中的高度异质性。为了克服这些限制，我们将Ptet调节元件置于孢子形成的主调节因子spo 0A的上游，产生了一种新的菌株，该菌株可以通过添加脱水四环素（ATc）人工诱导孢子形成。我们证明，该菌株是asporogenous在没有ATc，ATc可以用来驱动更快，更有效的孢子形成。Spo 0A的诱导是可滴定的，这可用于spo 0A调节子的体外和体内研究，如使用C.艰难梭菌感染（CDI）。深入了解B中孢子形成途径之间的差异。subtilis和C. difficile获得的诱导菌株的研究进行了讨论，进一步突出了这个工具的普遍利益。Ptet-spo 0A菌株提供了一个有用的背景，在其中产生涉及孢子形成的基因突变，因此提供了一个令人兴奋的新工具，以解开芽孢杆菌孢子形成的关键方面。很难
Clostridium difficile remains a leading nosocomial pathogen, putting considerable strain on the healthcare system. The ability to form endospores, highly resistant to environmental insults, is key to its persistence and transmission. However, important differences exist between the sporulation pathways of C. difficile and the model Gram-positive organism Bacillus subtilis. Amongst the challenges in studying sporulation in C. difficile is the relatively poor levels of sporulation and high heterogeneity in the sporulation process. To overcome these limitations we placed Ptet regulatory elements upstream of the master regulator of sporulation, spo0A, generating a new strain that can be artificially induced to sporulate by addition of anhydrotetracycline (ATc). We demonstrate that this strain is asporogenous in the absence of ATc, and that ATc can be used to drive faster and more efficient sporulation. Induction of Spo0A is titratable and this can be used in the study of the spo0A regulon both in vitro and in vivo, as demonstrated using a mouse model of C. difficile infection (CDI). Insights into differences between the sporulation pathways in B. subtilis and C. difficile gained by study of the inducible strain are discussed, further highlighting the universal interest of this tool. The Ptet-spo0A strain provides a useful background in which to generate mutations in genes involved in sporulation, therefore providing an exciting new tool to unravel key aspects of sporulation in C. difficile.