title: Analysis of TcdB Proteins within the Hypervirulent Clade 2 Reveals an Impact of RhoA Glucosylation on Clostridium difficile Proinflammatory Activities.
abstract: Clostridium difficile strains within the hypervirulent clade 2 are responsible for nosocomial outbreaks worldwide. The increased pathogenic potential of these strains has been attributed to several factors but is still poorly understood. During a C. difficile outbreak, a strain from this clade was found to induce a variant cytopathic effect (CPE), different from the canonical arborizing CPE. This strain (NAP1V) belongs to the NAP1 genotype but to a ribotype different from the epidemic NAP1/RT027 strain. NAP1V and NAP1 share some properties, including the overproduction of toxins, the binary toxin, and mutations in tcdC. NAP1V is not resistant to fluoroquinolones, however. A comparative analysis of TcdB proteins from NAP1/RT027 and NAP1V strains indicated that both target Rac, Cdc42, Rap, and R-Ras but only the former glucosylates RhoA. Thus, TcdB from hypervirulent clade 2 strains possesses an extended substrate profile, and RhoA is crucial for the type of CPE induced. Sequence comparison and structural modeling revealed that TcdBNAP1 and TcdBNAP1V share the receptor-binding and autoprocessing activities but vary in the glucosyltransferase domain, consistent with the different substrate profile. Whereas the two toxins displayed identical cytotoxic potencies, TcdBNAP1 induced a stronger proinflammatory response than TcdBNAP1V as determined in ex vivo experiments and animal models. Since immune activation at the level of intestinal mucosa is a hallmark of C. difficile-induced infections, we propose that the panel of substrates targeted by TcdB is a determining factor in the pathogenesis of this pathogen and in the differential virulence potential seen among C. difficile strains.
authors.affiliation: Facultad de Microbiología and Centro de Investigación en Enfermedades Tropicales, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: Carlos
authors.initials: C
authors.lastname: Quesada-Gómez
authors.affiliation: Facultad de Microbiología and Centro de Investigación en Enfermedades Tropicales, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: Diana
authors.initials: D
authors.lastname: López-Ureña
authors.affiliation: Department of Pathology, Microbiology, and Immunology, Vanderbilt University School of Medicine, Nashville, Tennessee, USA.
authors.forename: Nicole
authors.initials: N
authors.lastname: Chumbler
authors.affiliation: Department of Pathology, Microbiology, and Immunology, Vanderbilt University School of Medicine, Nashville, Tennessee, USA.
authors.forename: Heather K
authors.initials: HK
authors.lastname: Kroh
authors.affiliation: Facultad de Microbiología and Centro de Investigación en Enfermedades Tropicales, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: Carolina
authors.initials: C
authors.lastname: Castro-Peña
authors.affiliation: Facultad de Microbiología and Centro de Investigación en Enfermedades Tropicales, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: César
authors.initials: C
authors.lastname: Rodríguez
authors.affiliation: Facultad de Farmacia, Universidad de Costa Rica, San José, Costa Rica Laboratorio de Ensayos Biológicos, Escuela de Medicina, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: Josué
authors.initials: J
authors.lastname: Orozco-Aguilar
authors.affiliation: Laboratorio de Ensayos Biológicos, Escuela de Medicina, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: Sara
authors.initials: S
authors.lastname: González-Camacho
authors.affiliation: Instituto Clodomiro Picado, Facultad de Microbiología, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Rucavado
authors.affiliation: Programa de Investigación en Enfermedades Tropicales, Escuela de Medicina Veterinaria, Universidad Nacional, Heredia, Costa Rica.
authors.forename: Caterina
authors.initials: C
authors.lastname: Guzmán-Verri
authors.affiliation: Host-Microbiota Interactions Laboratory, Wellcome Trust Sanger Institute, Hinxton, United Kingdom.
authors.forename: Trevor D
authors.initials: TD
authors.lastname: Lawley
authors.affiliation: Department of Pathology, Microbiology, and Immunology, Vanderbilt University School of Medicine, Nashville, Tennessee, USA The Veterans Affairs Tennessee Valley Healthcare System, Nashville, Tennessee, USA.
authors.forename: D Borden
authors.initials: DB
authors.lastname: Lacy
authors.affiliation: Facultad de Microbiología and Centro de Investigación en Enfermedades Tropicales, Universidad de Costa Rica, San José, Costa Rica esteban.chaves@ucr.ac.cr.
authors.forename: Esteban
authors.initials: E
authors.lastname: Chaves-Olarte
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001427:Bacterial Toxins / Q000235:genetics / Q000378:metabolism*; D016360:Clostridioides difficile / Q000145:classification / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity*; D004761:Enterocolitis, Pseudomembranous / Q000201:enzymology* / Q000235:genetics / Q000382:microbiology*; D005838:Genotype; D006031:Glycosylation; D054884:Host-Pathogen Interactions; D006801:Humans; D008297:Male; D051379:Mice; D014774:Virulence; D020742:rhoA GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; C031815:toxB protein, Clostridium difficile; D020742:rhoA GTP-Binding Protein