title: Characterization of the Adherence ofClostridium difficileSpores: The Integrity of the Outermost Layer Affects Adherence Properties of Spores of the Epidemic Strain R20291 to Components of the Intestinal Mucosa.
abstract: Clostridium difficileis the causative agent of the most frequently reported nosocomial diarrhea worldwide. The high incidence of recurrent infection is the main clinical challenge ofC. difficileinfections (CDI). Formation ofC. difficilespores of the epidemic strain R20291 has been shown to be essential for recurrent infection and transmission of the disease in a mouse model. However, the underlying mechanisms of how these spores persist in the colonic environment remains unclear. In this work, we characterized the adherence properties of epidemic R20291 spores to components of the intestinal mucosa, and we assessed the role of the exosporium integrity in the adherence properties by usingcdeCmutant spores with a defective exosporium layer. Our results showed that spores and vegetative cells of the epidemic R20291 strain adhered at high levels to monolayers of Caco-2 cells and mucin. Transmission electron micrographs of Caco-2 cells demonstrated that the hair-like projections on the surface of R20291 spores are in close proximity with the plasma membrane and microvilli of undifferentiated and differentiated monolayers of Caco-2 cells. Competitive-binding assay in differentiated Caco-2 cells suggests that spore-adherence is mediated by specific binding sites. By using spores of acdeCmutant we demonstrated that the integrity of the exosporium layer determines the affinity of adherence ofC. difficilespores to Caco-2 cells and mucin. Binding of fibronectin and vitronectin to the spore surface was concentration-dependent, and depending on the concentration, spore-adherence to Caco-2 cells was enhanced. In the presence of an aberrantly-assembled exosporium (cdeCspores), binding of fibronectin, but not vitronectin, was increased. Notably, independent of the exosporium integrity, only a fraction of the spores had fibronectin and vitronectin molecules binding to their surface. Collectively, these results demonstrate that the integrity of the exosporium layer of strain R20291 contributes to selective spore adherence to components of the intestinal mucosa.
authors.affiliation: Microbiota-Host Interactions and Clostridia Research Group, Departamento de Ciencias Biológicas, Facultad de Ciencias Biológicas, Universidad Andres BelloSantiago, Chile; Center for Bioinformatics and Integrative Biology, Facultad de Ciencias Biológicas, Universidad Andres BelloSantiago, Chile.
authors.forename: Paola
authors.initials: P
authors.lastname: Mora-Uribe
authors.affiliation: Microbiota-Host Interactions and Clostridia Research Group, Departamento de Ciencias Biológicas, Facultad de Ciencias Biológicas, Universidad Andres Bello Santiago, Chile.
authors.forename: Camila
authors.initials: C
authors.lastname: Miranda-Cárdenas
authors.affiliation: Microbiota-Host Interactions and Clostridia Research Group, Departamento de Ciencias Biológicas, Facultad de Ciencias Biológicas, Universidad Andres BelloSantiago, Chile; Center for Bioinformatics and Integrative Biology, Facultad de Ciencias Biológicas, Universidad Andres BelloSantiago, Chile.
authors.forename: Pablo
authors.initials: P
authors.lastname: Castro-Córdova
authors.affiliation: Microbiota-Host Interactions and Clostridia Research Group, Departamento de Ciencias Biológicas, Facultad de Ciencias Biológicas, Universidad Andres Bello Santiago, Chile.
authors.forename: Fernando
authors.initials: F
authors.lastname: Gil
authors.affiliation: Laboratorio de Genética y Patogénesis Bacteriana, Departamento de Ciencias Biológicas, Facultad de Ciencias Biológicas, Universidad Andres Bello Santiago, Chile.
authors.forename: Iván
authors.initials: I
authors.lastname: Calderón
authors.affiliation: Laboratorio de Genética y Patogénesis Bacteriana, Departamento de Ciencias Biológicas, Facultad de Ciencias Biológicas, Universidad Andres Bello Santiago, Chile.
authors.forename: Juan A
authors.initials: JA
authors.lastname: Fuentes
authors.affiliation: Facultad de Medicina, Center for Integrative Medicine and Innovative Sciences, Universidad Andres Bello Santiago, Chile.
authors.forename: Paula I
authors.initials: PI
authors.lastname: Rodas
authors.affiliation: Department of Biomedical Sciences, Oregon State UniversityCorvallis, OR, USA; Medical Laboratories Department, College of Science Al-Zulfi, Majmaah UniversityAl Majma'ah, Saudi Arabia.
authors.forename: Saeed
authors.initials: S
authors.lastname: Banawas
authors.affiliation: Department of Biomedical Sciences, Oregon State University Corvallis, OR, USA.
authors.forename: Mahfuzur R
authors.initials: MR
authors.lastname: Sarker
authors.affiliation: Microbiota-Host Interactions and Clostridia Research Group, Departamento de Ciencias Biológicas, Facultad de Ciencias Biológicas, Universidad Andres BelloSantiago, Chile; Center for Bioinformatics and Integrative Biology, Facultad de Ciencias Biológicas, Universidad Andres BelloSantiago, Chile.
authors.forename: Daniel
authors.initials: D
authors.lastname: Paredes-Sabja
mesh_terms: D000818:Animals; D001422:Bacterial Adhesion / Q000502:physiology*; D001426:Bacterial Proteins / Q000235:genetics; D018938:Caco-2 Cells / Q000382:microbiology; D002473:Cell Wall; D016360:Clostridioides difficile / Q000472:pathogenicity / Q000502:physiology*; D004195:Disease Models, Animal; D004761:Enterocolitis, Pseudomembranous / Q000382:microbiology*; D005353:Fibronectins / Q000378:metabolism; D006801:Humans; D007413:Intestinal Mucosa / Q000382:microbiology; D051379:Mice; D046529:Microscopy, Electron, Transmission; D008871:Microvilli / Q000382:microbiology; D009077:Mucins; D013171:Spores, Bacterial / Q000502:physiology*; D019096:Vitronectin / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D005353:Fibronectins; D009077:Mucins; D019096:Vitronectin
keywords: BclA; C. difficile spores; CdeC; exosporium; spore adherence