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IMPACT OF BIOACTIVE MOLECULES SECRETED BY LACTOBACILLUS ACIDOPHILUS LA-5 ON CLOSTRIDIUM DIFFICILE VIRULENCE FACTORS

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Title: IMPACT OF BIOACTIVE MOLECULES SECRETED BY LACTOBACILLUS ACIDOPHILUS LA-5 ON CLOSTRIDIUM DIFFICILE VIRULENCE FACTORS
Author: NAJARIAN, AFSANEH
Department: Department of Food Science
Program: Food Science
Advisor: Griffiths, ManselShayan, Sharif
Abstract: Clostridium difficile is a leading pathogen of hospital-associated diseases including infectious diarrhea and pseudomembranous colitis. Increasing frequency and severity of diseases besides inconsistent results of antibiotic therapy, demand alternative therapeutic candidates using a different strategy to control pathogenicity of C. difficile. Unlike antibiotics, antivirulence compounds control bacterial infection without killing them or developing antibiotic-resistant. Reducing essential virulence factors in C. difficile including toxin production, adherence, and biofilm formation could substantially minimize its pathogenicity and lead to a faster recovery from the disease. This study aimed to investigate antivirulence effects of La-5 bioactive molecules produced by Lactobacillus acidophilus La-5 namely proteobiotics on the pathogenicity of C. difficile. Three clinically important strains of C. difficile (ribotypes 027, 078, and 001) were incubated with or without reconstituted La-5 cell-free supernatant (CFS). The C. difficile culture filtrates were collected for the assessment of quorum sensing, and expression of several virulence genes performing reverse transcription q-PCR. C. difficile attachment and cytotoxicity in human epithelial cells were monitored in vitro using human epithelial cells HT-29 and Caco-2 monolayers. Additionally, L. acidophilus La-5 was incubated in a medium containing whey protein, and inhibitory effects of its CFS were tested performing quantitative biofilm assay then visualized in scanning electron microscopy (SEM). Bioactive molecules caused a significant down-regulation in expression of some of the virulence genes mainly two essential toxin genes tcdA and tcdB and adhesion gene Cwp84 in all C. difficile strains. A substantial down-regulation in luxS expression was observed in ribotype 027 and 078, and 001 (P<0.05). La-5 bioactive molecules also reduced cytotoxicity and cytopathic effect of C. difficile culture filtrate on HT-29 and Caco-2 monolayers significantly (P < 0.0.5). Furthermore, pre-incubation of cell monolayers and La-5 CFS reduced attachment of the C. difficile bacterial cells on both cell monolayers. The biofilm formation and population of C. difficile cells encased in biofilms were significantly decreased within the treated cultures (P < 0.0001). SEM analysis also confirmed quantitative results showing a sufficient reduction in biofilm formation. Our study suggests that La-5 bioactive molecules provide a supportive adjunct therapy for C. difficile infection, prevent relapses, and improve disease outcome.
URI: http://hdl.handle.net/10214/12153
Date: 2017-11
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