Publication Date

2008

Document Type

Thesis

Committee Members

Michael P. Markey (Committee Member), John V. Paietta (Committee Member), Oleg Paliy (Committee Chair)

Degree Name

Master of Science (MS)

Abstract

Intestinal microflora refers to all the different species of bacteria that reside in the human gut and is an important organ of the human body because almost all the digestive reactions of the host occur in the intestine. The bacteria of the intestine play a key role in this process by supplementing the intestine with various enzymes and proteins that are required for the digestive process. At the same time, these bacteria were shown to be implicated in a variety of gastrointestinal disorders like Irritable Bowel Syndrome, Inflammatory Bowel Disorder and Gastrointestinal Cancer, but with the current knowledge about the microflora it is difficult to determine which exact species is responsible for a particular disease caused. The knowledge about the composition of the typical intestinal microflora is very limited, the cause at large being the lack of properculture techniques to isolate and study the microfloral species in artificial media. Majority of the species of the microflora are obligate anaerobes and selective culturing techniques provide very limited knowledge about the composition of such complex microflora. Phylogenetic microarrays are one such approach to study various members of the microflora because they contain probes for numerous species of bacteria on a single glass slide and are also known to provide robust and high throughput analysis.

ENTREZ nucleotide database was used to compile a list of 16S ribosomal DNA (rDNA) sequences of bacterial species isolated from the human intestine and they were grouped into various phylo-species. Representative sequences for each phylo-species were extracted and the probes on the microarray were designed based on these representative sequences. 16 different bacterial species were used for validation experiments, which represented bacteria from various groups. The results showed that the microarray correctly identified 15 of a total 16 bacterial species. The detection sensitivity of the microarray was at least 1pg. As a test, fecal samples from adults and children were analyzed by the microarray. Clostridia were the dominant group of the microflora followed by Bacteroidetes in both adults and children. The analysis of the fecal samples showed clear differences between the microflora composition of adults and children.

Page Count

77

Department or Program

Department of Biochemistry and Molecular Biology

Year Degree Awarded

2008

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