title: A Clostridium difficile Cell Wall Glycopolymer Locus Influences Bacterial Shape, Polysaccharide Production and Virulence.
abstract: Clostridium difficile is a diarrheagenic pathogen associated with significant mortality and morbidity. While its glucosylating toxins are primary virulence determinants, there is increasing appreciation of important roles for non-toxin factors in C. difficile pathogenesis. Cell wall glycopolymers (CWGs) influence the virulence of various pathogens. Five C. difficile CWGs, including PSII, have been structurally characterized, but their biosynthesis and significance in C. difficile infection is unknown. We explored the contribution of a conserved CWG locus to C. difficile cell-surface integrity and virulence. Attempts at disrupting multiple genes in the locus, including one encoding a predicted CWG exporter mviN, were unsuccessful, suggesting essentiality of the respective gene products. However, antisense RNA-mediated mviN downregulation resulted in slight morphology defects, retarded growth, and decreased surface PSII deposition. Two other genes, lcpA and lcpB, with putative roles in CWG anchoring, could be disrupted by insertional inactivation. lcpA- and lcpB- mutants had distinct phenotypes, implying non-redundant roles for the respective proteins. The lcpB- mutant was defective in surface PSII deposition and shedding, and exhibited a remodeled cell surface characterized by elongated and helical morphology, aberrantly-localized cell septae, and an altered surface-anchored protein profile. Both lcpA- and lcpB- strains also displayed heightened virulence in a hamster model of C. difficile disease. We propose that gene products of the C. difficile CWG locus are essential, that they direct the production/assembly of key antigenic surface polysaccharides, and thereby have complex roles in virulence.
authors.affiliation: School of Animal and Comparative Biomedical Sciences, University of Arizona, Tucson, Arizona, United States of America.
authors.forename: Michele
authors.initials: M
authors.lastname: Chu
authors.affiliation: School of Animal and Comparative Biomedical Sciences, University of Arizona, Tucson, Arizona, United States of America.
authors.forename: Michael J G
authors.initials: MJ
authors.lastname: Mallozzi
authors.affiliation: School of Animal and Comparative Biomedical Sciences, University of Arizona, Tucson, Arizona, United States of America.
authors.forename: Bryan P
authors.identifier: http://orcid.org/0000-0002-0139-2232
authors.initials: BP
authors.lastname: Roxas
authors.affiliation: Department of Chemistry, University of Guelph, Guelph, Ontario, Canada.
authors.forename: Lisa
authors.initials: L
authors.lastname: Bertolo
authors.affiliation: Department of Chemistry, University of Guelph, Guelph, Ontario, Canada.
authors.forename: Mario A
authors.initials: MA
authors.lastname: Monteiro
authors.affiliation: School of Animal and Comparative Biomedical Sciences, University of Arizona, Tucson, Arizona, United States of America.
authors.forename: Al
authors.initials: A
authors.lastname: Agellon
authors.affiliation: School of Animal and Comparative Biomedical Sciences, University of Arizona, Tucson, Arizona, United States of America.|Department of Immunobiology, Bio5 Institute for Collaborative Research, University of Arizona, Tucson, Arizona, United States of America.
authors.forename: V K
authors.initials: VK
authors.lastname: Viswanathan
authors.affiliation: School of Animal and Comparative Biomedical Sciences, University of Arizona, Tucson, Arizona, United States of America.|Department of Immunobiology, Bio5 Institute for Collaborative Research, University of Arizona, Tucson, Arizona, United States of America.|Southern Arizona VA Healthcare System, Tucson, Arizona, United States of America.
authors.forename: Gayatri
authors.initials: G
authors.lastname: Vedantam
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism*; D002473:Cell Wall / Q000737:chemistry / Q000648:ultrastructure*; D016360:Clostridioides difficile / Q000472:pathogenicity* / Q000648:ultrastructure*; D003015:Clostridium Infections / Q000821:virology*; D006224:Cricetinae; D004195:Disease Models, Animal; D005455:Fluorescent Antibody Technique; D015151:Immunoblotting; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism; D008647:Mesocricetus; D008854:Microscopy, Electron; D016254:Mutagenesis, Insertional; D016297:Mutagenesis, Site-Directed; D016133:Polymerase Chain Reaction; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism; D014774:Virulence; D037521:Virulence Factors / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D008562:Membrane Glycoproteins; D011134:Polysaccharides; D037521:Virulence Factors