Date of Completion

1-14-2019

Embargo Period

1-14-2020

Keywords

Clostridium difficile, sodium selenite, baicalin, lactic acid bacteria, mouse model

Major Advisor

Dr. Kumar Venkitanarayanan

Associate Advisor

Dr. Mary Anne Roshni Amalaradjou

Associate Advisor

Dr. Dennis J. D'Amico

Associate Advisor

Dr. Mazhar I. Khan

Associate Advisor

Dr. Bhushan M. Jayarao

Field of Study

Animal Science

Degree

Doctor of Philosophy

Open Access

Open Access

Abstract

Clostridium difficile is a significant enteric pathogen causing a toxin-mediated infection and diarrhea in humans. There has been an increased incidence of C. difficile infection (CDI) in the United States with the emergence of hypervirulent strains and community associated outbreaks. CDI is commonly observed among hospital in-patients undergoing protracted antibiotic therapy, which results in gastrointestinal dysbiosis, creating a conducive environment for spore germination, pathogen colonization in the gut, and subsequent toxin production. An ideal anti-C.difficile therapeutic agent should inhibit critical virulence factors of the pathogen such as toxin production, sporulation and spore germination without inducing gut dysbiosis. Such agents when provided as an adjunct to C. difficile antibiotic therapy could help to improve the clinical outcome of CDI and prevent the relapse of the infection. In this Ph.D. research, the efficacy of three alternative treatment approaches as antivirulence agents was tested for potential future development as therapies against CDI. This included sodium selenite (metalloid), baicalin (flavone glycoside), and selected lactic acid bacteria. All treatment modalities were tested for inhibiting toxin production, sporulation and spore outgrowth in two hypervirulent C. difficile isolates. Moreover, gene expression and cell culture studies were performed to elucidate the anti-toxigenic mechanism of sodium selenite and baicalin. In addition, the effect of sodium selenite in increasing pathogen sensitivity to ciprofloxacin and vancomycin, two common antibiotics used in treating C. difficile, was also tested. Furthermore, the effect of baicalin on CDI was investigated in a mouse model, with special reference to its effect on disease severity and the mouse gut microbiome. The results revealed that sub-minimum inhibitory concentration (sub-MIC) and sub-inhibitory concentrations (SIC) of sodium selenite and baicalin, respectively, reduced C. difficile toxin production and cytotoxicity in vitro. In addition, sodium selenite and baicalin inhibited spore outgrowth. Oral supplementation of baicalin improved the clinical outcome in challenged mice, and positively altered the gut microbiome composition. Collectively, these results indicate that the three approaches identified in this study significantly reduced C. difficile virulence, however, follow up validation in animal models for long-term safety and dose standardization, and clinical trials in human subjects are necessary.

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