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Inducible expression of spo0A as a universal tool for studying sporulation in Clostridium difficile

Lookup NU author(s): Dr Marcin Dembek, Dr Paula Salgado

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This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).


Abstract

© 2017 Dembek, Willing, Hong, Hosseini, Salgado and Cutting. 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.


Publication metadata

Author(s): Dembek M, Willing SE, Hong HA, Hosseini S, Salgado PS, Cutting SM

Publication type: Article

Publication status: Published

Journal: Frontiers in Microbiology

Year: 2017

Volume: 8

Online publication date: 21/09/2017

Acceptance date: 05/09/2017

Date deposited: 29/09/2017

ISSN (electronic): 1664-302X

Publisher: Frontiers Media S.A.

URL: https://doi.org/10.3389/fmicb.2017.01793

DOI: 10.3389/fmicb.2017.01793


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