The glucosyltransferase activity of C. difficile toxin b is required for disease pathogenesis

Bilverstone, Terry W.; Garland, Megan; Cave, Rory J.; Kelly, Michelle L.; Tholen, Martina; Bouley, Donna M.; Kaye, Philip; Minton, Nigel P.; Bogyo, Matthew; Kuehne, Sarah A.; Melnyk, Roman A.

Abstract

© 2020 Bilverstone et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Enzymatic inactivation of Rho-family GTPases by the glucosyltransferase domain of Clostridioides difficile Toxin B (TcdB) gives rise to various pathogenic effects in cells that are classically thought to be responsible for the disease symptoms associated with C. difficile infection (CDI). Recent in vitro studies have shown that TcdB can, under certain circumstances, induce cellular toxicities that are independent of glucosyltransferase (GT) activity, calling into question the precise role of GT activity. Here, to establish the importance of GT activity in CDI disease pathogenesis, we generated the first described mutant strain of C. difficile producing glucosyltransferase-defective (GT-defective) toxin. Using allelic exchange (AE) technology, we first deleted tcdA in C. difficile 630Δerm and subsequently introduced a deactivating D270N substitution in the GT domain of TcdB. To examine the role of GT activity in vivo, we tested each strain in two different animal models of CDI pathogenesis. In the non-lethal murine model of infection, the GT-defective mutant induced minimal pathology in host tissues as compared to the profound caecal inflammation seen in the wild-type and 630ΔermΔtcdA (ΔtcdA) strains. In the more sensitive hamster model of CDI, whereas hamsters in the wild-type or ΔtcdA groups succumbed to fulminant infection within 4 days, all hamsters infected with the GT-defective mutant survived the 10-day infection period without primary symptoms of CDI or evidence of caecal inflammation. These data demonstrate that GT activity is indispensable for disease pathogenesis and reaffirm its central role in disease and its importance as a therapeutic target for small-molecule inhibition.

• Journal Article Type: Article
• Acceptance Date: Aug 3, 2020
• Online Publication Date: Sep 22, 2020
• Publication Date: Sep 1, 2020
• Deposit Date: Nov 3, 2020
• Publicly Available Date: Nov 3, 2020
• Journal: PLoS Pathogens
• Print ISSN: 1553-7366
• Electronic ISSN: 1553-7374
• Publisher: Public Library of Science
• Peer Reviewed: Peer Reviewed
• Volume: 16
• Issue: 9
• Article Number: e1008852
• DOI: https://doi.org/10.1371/journal.ppat.1008852
• Keywords: Immunology; Genetics; Molecular Biology; Microbiology; Parasitology; Virology
• Public URL: https://nottingham-repository.worktribe.com/output/4928197
• Publisher URL: https://journals.plos.org/plospathogens/article?id=10.1371/journal.ppat.1008852