title: Surface-layer (S-layer) of human and animal Clostridium difficile strains and their behaviour in adherence to epithelial cells and intestinal colonization.
abstract: Clostridium difficile is a frequent cause of severe, recurrent post-antibiotic diarrhoea and pseudomembranous colitis. The surface layer (S-layer) is the predominant outer surface component of C. difficile which is involved in pathogen-host interactions critical to pathogenesis. In this study, we characterized the S-layer protein A (SlpA) of animal and human strains belonging to different PCR-ribotypes (PR) and compared the in vitro adherence and in vivo colonization properties of strains showing different SlpA variants. Since each SlpA variant has been recently associated with an S-layer cassette, we were able to deduce the cassette for each of our strains. In this study, an identity of 99-100 % was found among the SlpA of isolates belonging to PR 012, 014/020, 045 and 078. One exception was the SlpA of a poultry isolate, PR 014/020, which showed 99 % identity with that of strain 0160, another PR 014/020 which contains an S-layer cassette 6. Interestingly, this cassette has also been found in a PR 018 strain, an emerging virulent type currently predominant in Italy. Five other SlpA variants (v014/020a-e) were identified in strains PR 014/020. In vitro adherence assays and in vivo colonization experiments were performed on five PR 014/020 strains: human 1064 (v014/020e), human 4684/08 (v014/020b), human IT1106 (v078a), poultry P30 (v014/020d) and poultry PB90 (v014/020b) strains. Adhesion assays indicate that C. difficile strains vary in their capacity to adhere to cells in culture and that adhesion seems to be independent of the SlpA variant. Colonization properties were assessed in vivo using a dixenic mouse model of colonization. The kinetics of faecal shedding and caecal colonization were similar when human 4684/08 (v014/020b) strain was compared with human 1064 (v014/020e) and poultry PB90 (v014/020b) strain. In contrast, poultry P30 (v014/020d) strain outcompeted both human 4684/08 (v014/020b) and IT1106 (v078a) strains and its adherence to caeca at day 7 was significantly higher. The peculiar characteristics of C. difficile P30 seem to advantage it in colonizing the intestinal mice niche, increasing its ability to compete and adapt. The results obtained underline the need of an increased attention to the genetic evolution of C. difficile to prevent and limit the consequences of the emergence of increasingly virulent strains.
authors.affiliation: Istituto Superiore di Sanità, Roma, Italy.
authors.forename: Patrizia
authors.initials: P
authors.lastname: Spigaglia
authors.affiliation: EA 4043, USC INRA Faculté de Pharmacie, Université Paris-Sud 11, France.
authors.forename: Amira
authors.initials: A
authors.lastname: Barketi-Klai
authors.affiliation: EA 4043, USC INRA Faculté de Pharmacie, Université Paris-Sud 11, France.
authors.forename: Anne
authors.initials: A
authors.lastname: Collignon
authors.affiliation: Istituto Superiore di Sanità, Roma, Italy.
authors.forename: Paola
authors.initials: P
authors.lastname: Mastrantonio
authors.affiliation: Istituto Superiore di Sanità, Roma, Italy.
authors.forename: Fabrizio
authors.initials: F
authors.lastname: Barbanti
authors.affiliation: Centre of Excellence for Integrated Approaches in Chemistry and Biology of Proteins, Ljubljana, Slovenia.|University of Maribor, Faculty of Medicine, Maribor, Slovenia.|Institute of Public Health Maribor, Maribor, Slovenia.
authors.forename: Maja
authors.initials: M
authors.lastname: Rupnik
authors.affiliation: Institute of Public Health Maribor, Maribor, Slovenia.
authors.forename: Sandra
authors.initials: S
authors.lastname: Janezic
authors.affiliation: EA 4043, USC INRA Faculté de Pharmacie, Université Paris-Sud 11, France.
authors.forename: Imad
authors.initials: I
authors.lastname: Kansau
mesh_terms: D000222:Adaptation, Physiological; D000595:Amino Acid Sequence; D000818:Animals; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000378:metabolism*; D016360:Clostridioides difficile / Q000145:classification / Q000235:genetics / Q000502:physiology*; D004761:Enterocolitis, Pseudomembranous / Q000382:microbiology; D004847:Epithelial Cells / Q000382:microbiology*; D014644:Genetic Variation; D006801:Humans; D007422:Intestines / Q000382:microbiology*; D051379:Mice; D008809:Mice, Inbred C3H; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D010802:Phylogeny; D011200:Poultry / Q000382:microbiology
chemical_list: D001426:Bacterial Proteins; C070683:surface layer protein A, Bacteria