Single base extension reaction and its applications in environmental microbiological studies

Abstract

A molecular method termed hierarchical oligonucleotide primer extension (HOPE) is developed to complement the current suite of quantitative molecular tools, and to provide quantitative measures of the relative abundances of bacterial targets among total amplified 16S rRNA genes. To demonstrate, this dissertation first validates the use of HOPE to quantify 14 Bacteroides spp. in the feces and fecal-contaminated water samples. The method is subsequently applied to examine two areas related to the fecal environment, namely, fecal contamination in water bodies and health-related fecal microbiota. This dissertation shows that HOPE is able to quantify the abundances of host-specific Bacteroidales, and is capable of identifying fecal contamination sources in blind samples and environmental waters. Furthermore, this dissertation shows that HOPE can provide a time- and cost-effective approach to quantify numerous health-related Bifidobacterium spp. that are present in a statistically representative sample size.