Date of Award

12-2014

Document Type

Thesis

Degree Name

Master of Science (MS)

Legacy Department

Biological Sciences

Advisor

Dr. Min Cao

Committee Member

Dr. Yuqing Dong

Committee Member

Dr. Thomas A. Hughes

Abstract

Vibrio cholerae, a Gram-negative bacterium found in natural aquatic environments, is the causative agent of cholera, a severe diarrheal disease most commonly spread through drinking water. An essential component to this pathogen's success and persistence in the environment is its ability to attach to both biotic and abiotic surfaces via biofilm formation. Biofilms not only aid in surface attachment, but also provide a barrier that protects and enhances survival. Water-soluble extracts from the North American cranberry (Vaccinium macrocarpon) were found to dramatically inhibit V. cholerae biofilm formation at a low concentration of 2mg/ml. This inhibition was not due to acidification of the growth medium or any bactericidal or bacteriostatic effects. The biofilm inhibition also appeared to be specific to V. cholerae. Furthermore, a decrease in biofilm was also observed when water-soluble cranberry extracts (WSCE) were added after four hours of pre-incubation of the culture statically. The vps operons, which encode the major component of V. cholerae biofilm, exopolysaccharide, were significantly down-regulated in the presence of WSCE by qRT-PCR assays. The vps operons are controlled by a set of transcriptional activators and repressors within a tangled signaling network and are regulated in a unique cell density dependent manner known as quorum sensing. To test whether the master quorum-sensing regulator HapR is involved in this cranberry-mediated anti-biofilm effect, the same biofilm inhibition assay was conducted in a hapR deletion mutant. A similar level of inhibition was detected, suggesting that HapR does not appear to be required for the inhibition by WSCE. To further confirm this finding, a wild type V. cholerae strain containing a hapR-lacZ fusion plasmid was used to determine in vivo expression driven by the hapR promoter and measured by β-galactosidase activities. Additionally, it was determined that the intracellular concentration of cyclic diguanylate (c-di-GMP), an important second messenger used in signal transduction, is significantly reduced in the presence of WSCE. In summary, the results suggest a potential application of WSCE as an alternative and safe means in V. cholerae biofilm control.

Included in

Microbiology Commons

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