title: Surface-layer protein A (SlpA) is a major contributor to host-cell adherence of Clostridium difficile.
abstract: Clostridium difficile is a leading cause of antibiotic-associated diarrhea, and a significant etiologic agent of healthcare-associated infections. The mechanisms of attachment and host colonization of C. difficile are not well defined. We hypothesize that non-toxin bacterial factors, especially those facilitating the interaction of C. difficile with the host gut, contribute to the initiation of C. difficile infection. In this work, we optimized a completely anaerobic, quantitative, epithelial-cell adherence assay for vegetative C. difficile cells, determined adherence proficiency under multiple conditions, and investigated C. difficile surface protein variation via immunological and DNA sequencing approaches focused on Surface-Layer Protein A (SlpA). In total, thirty-six epidemic-associated and non-epidemic associated C. difficile clinical isolates were tested in this study, and displayed intra- and inter-clade differences in attachment that were unrelated to toxin production. SlpA was a major contributor to bacterial adherence, and individual subunits of the protein (varying in sequence between strains) mediated host-cell attachment to different extents. Pre-treatment of host cells with crude or purified SlpA subunits, or incubation of vegetative bacteria with anti-SlpA antisera significantly reduced C. difficile attachment. SlpA-mediated adherence-interference correlated with the attachment efficiency of the strain from which the protein was derived, with maximal blockage observed when SlpA was derived from highly adherent strains. In addition, SlpA-containing preparations from a non-toxigenic strain effectively blocked adherence of a phylogenetically distant, epidemic-associated strain, and vice-versa. Taken together, these results suggest that SlpA plays a major role in C. difficile infection, and that it may represent an attractive target for interventions aimed at abrogating gut colonization by this pathogen.
authors.affiliation: Department of Microbiology and Immunology, Loyola University Medical Center, Maywood, Illinois, United States of America.
authors.forename: Michelle M
authors.initials: MM
authors.lastname: Merrigan
authors.forename: Anilrudh
authors.initials: A
authors.lastname: Venugopal
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Roxas
authors.forename: Farhan
authors.initials: F
authors.lastname: Anwar
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Mallozzi
authors.forename: Bryan A P
authors.initials: BA
authors.lastname: Roxas
authors.forename: Dale N
authors.initials: DN
authors.lastname: Gerding
authors.forename: V K
authors.initials: VK
authors.lastname: Viswanathan
authors.forename: Gayatri
authors.initials: G
authors.lastname: Vedantam
mesh_terms: D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001427:Bacterial Toxins / Q000096:biosynthesis; D016360:Clostridioides difficile / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D004847:Epithelial Cells / Q000382:microbiology; D005838:Genotype; D006801:Humans; D013045:Species Specificity
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; C070683:surface layer protein A, Bacteria