title: Comparative genomic analysis of toxin-negative strains of Clostridium difficile from humans and animals with symptoms of gastrointestinal disease.
abstract: BACKGROUND
  Clostridium difficile infections (CDI) are a significant health problem to humans and food animals. Clostridial toxins ToxA and ToxB encoded by genes tcdA and tcdB are located on a pathogenicity locus known as the PaLoc and are the major virulence factors of C. difficile. While toxin-negative strains of C. difficile are often isolated from faeces of animals and patients suffering from CDI, they are not considered to play a role in disease. Toxin-negative strains of C. difficile have been used successfully to treat recurring CDI but their propensity to acquire the PaLoc via lateral gene transfer and express clinically relevant levels of toxins has reinforced the need to characterise them genetically. In addition, further studies that examine the pathogenic potential of toxin-negative strains of C. difficile and the frequency by which toxin-negative strains may acquire the PaLoc are needed.
  
  
  RESULTS
  We undertook a comparative genomic analysis of five Australian toxin-negative isolates of C. difficile that lack tcdA, tcdB and both binary toxin genes cdtA and cdtB that were recovered from humans and farm animals with symptoms of gastrointestinal disease. Our analyses show that the five C. difficile isolates cluster closely with virulent toxigenic strains of C. difficile belonging to the same sequence type (ST) and have virulence gene profiles akin to those in toxigenic strains. Furthermore, phage acquisition appears to have played a key role in the evolution of C. difficile.
  
  
  CONCLUSIONS
  Our results are consistent with the C. difficile global population structure comprising six clades each containing both toxin-positive and toxin-negative strains. Our data also suggests that toxin-negative strains of C. difficile encode a repertoire of putative virulence factors that are similar to those found in toxigenic strains of C. difficile, raising the possibility that acquisition of PaLoc by toxin-negative strains poses a threat to human health. Studies in appropriate animal models are needed to examine the pathogenic potential of toxin-negative strains of C. difficile and to determine the frequency by which toxin-negative strains may acquire the PaLoc.
authors.affiliation: The ithree institute, University of Technology Sydney, Sydney, 2007, Australia. Piklu.Bhattacharya@uts.edu.au.|NSW Department of Primary Industries, Elizabeth Macarthur Agricultural Institute, PMB 8, Camden, NSW, 2570, Australia. Piklu.Bhattacharya@uts.edu.au.
authors.forename: Piklu
authors.initials: P
authors.lastname: Roy Chowdhury
authors.affiliation: The ithree institute, University of Technology Sydney, Sydney, 2007, Australia.
authors.forename: Matthew
authors.initials: M
authors.lastname: DeMaere
authors.affiliation: NSW Department of Primary Industries, Elizabeth Macarthur Agricultural Institute, PMB 8, Camden, NSW, 2570, Australia.
authors.forename: Toni
authors.initials: T
authors.lastname: Chapman
authors.affiliation: The ithree institute, University of Technology Sydney, Sydney, 2007, Australia.
authors.forename: Paul
authors.initials: P
authors.lastname: Worden
authors.affiliation: The ithree institute, University of Technology Sydney, Sydney, 2007, Australia.|Institute of Food Research, Norwich Research Park, Colney, Norwich, NR4 7UA, UK.
authors.forename: Ian G
authors.initials: IG
authors.lastname: Charles
authors.affiliation: The ithree institute, University of Technology Sydney, Sydney, 2007, Australia.
authors.forename: Aaron E
authors.initials: AE
authors.lastname: Darling
authors.affiliation: The ithree institute, University of Technology Sydney, Sydney, 2007, Australia. Steven.Djordjevic@uts.edu.au.
authors.forename: Steven P
authors.initials: SP
authors.lastname: Djordjevic
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics; D001427:Bacterial Toxins / Q000378:metabolism; D016360:Clostridioides difficile / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D003015:Clostridium Infections / Q000382:microbiology* / Q000662:veterinary*; D005767:Gastrointestinal Diseases / Q000382:microbiology* / Q000662:veterinary*; D006734:Horse Diseases / Q000382:microbiology*; D006736:Horses; D006801:Humans; D008969:Molecular Sequence Data; D010802:Phylogeny; D016415:Sequence Alignment; D013552:Swine; D013553:Swine Diseases / Q000382:microbiology*
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins
keywords: CDI; Clostridium difficile; Comparative genomics; Toxin-negative isolates; Zoonosis