Date of Graduation

Summer 2015

Degree

Master of Science in Biology

Department

Biology

Committee Chair

Paul Schweiger

Abstract

Fecal waste enters waterways from run-off during rain events, and is especially high in agricultural areas due to manure application and livestock. Increased fecal waste may also be due to direct human influence. This fecal loading may lead to eutrophication and poses health hazards to humans. By determining the source of fecal pollution, practices can be implemented to reduce the amount entering waterways. Bacteroides sp. have previously shown high host specificity allows for species-specific identification of fecal sources. Using quantitative real-time PCR (qPCR), human-associated and bovine-associated Bacteroides DNA, as well as total Bacteroides DNA can be quantified and concentrations of fecal contamination in lakes and streams can be determined. Amplification control plasmids containing 16S rDNA from human and bovine specific Bacteroides were constructed using TOPO cloning. Samples taken from McDaniel Lake in Springfield, MO Dec. 2013 – Dec. 2014 were analyzed with end-point PCR and fecal concentrations determined with qPCR. Quantitative PCR showed increased sensitivity and selectivity, compared to end-point PCR. While Bacteroides sp. were present in all samples, less than 1% of total Bacteroides in McDaniel Lake was identified as human- or bovine-associated Bacteroides for the sample period. The use of qPCR is shown to be a valuable tool in identifying and quantifying fecal inputs and monitoring nonpoint source pollution as urban and agricultural areas encroach on natural systems.

Keywords

microbial source tracking, human-associated Bacteroides, bovine-associated Bacteroides, qPCR, hydrolysis probes

Subject Categories

Biology

Copyright

© Kalie Michelle Somerville

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