Department of Population Medicine and Diagnostic Sciences
Contact Information: Email: firstname.lastname@example.org, Phone: 607-253-3675
Sponsor: Cornell University, College of Veterinary Medicine, Research Grants Program in Animal Health
Grant Number: N/A
Title: Characterization of a Hypervirulent Strain of C. difficile (ribotype 078)
Annual Direct Cost: $47,075
Project Period: 10/01/2012-09/30/2014
DESCRIPTION (provided by applicant): Clostridium difficile–associated disease (CDAD) is increasing in prevalence with increasing numbers of cases associated with infection by hypervirulent C. difficile strains (HCDS) requiring emergency care. Infections by HCDS are thought to cause a more severe disease that has higher relapse rates and increased mortality. Recently, C. difficile has also emerged as either a food animal pathogen or commensal organism. The most prevalent C. difficile ribotype found in neonatal piglets in the U.S. and worldwide is ribotype 078, which is indistinguishable from human strains of the same ribotype. Ribotype 078 produces two main virulence factors, toxin A and B, the concentration of which mainly determines disease severity. The two major molecular hallmarks of HCDS are: (i) increased host cell adhesion and colonization capacity, and (ii) enhanced toxin production or toxin activity. Although these two parameters are major virulence attributes of hypervirulent strains, the underlying mechanisms of CDAD and ancillary genes that might drive these virulence attributes are not yet fully understood. Forward genetic studies using random transposon mutagenesis is an ideal tool to unravel new virulence associated genes without making prior assumptions. We propose to identify new determinants that specifically contribute to the C. difficile hypervirulent phenotype by applying a mariner-based transposon system for random mutagenesis in a porcine C. difficile strain (ribotype 078) and virulence assessment in hamsters and a fetal rabbit xenograft animal model.
Transconjugants will be screened for phenotypes affecting (i) host cell adhesion and colonization capacity, and (ii) toxin production. The highest ranking mutants from this screen will be further selected using a SCID Mouse Fetal Rabbit Intestinal Xenograft (FRIX) model. The most promising mutants from these experiments will then be further characterized using whole genome expression microarrays and their effect on pathogenicity will be studied in a hamster model. We have conducted a wide range of molecular pathogenesis studies in C. difficile, and thus, we are in an excellent position to conduct all of the proposed studies. We expect that we will identify new genes that contribute to our understanding of CDAD pathogenesis that is essential for development of targeted treatments and more effective vaccines for prevention of C. difficile disease in animals.