Date of Award

May 2017

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Department

Freshwater Sciences

First Advisor

Sandra L. McLellan

Committee Members

Harvey Bootsma, Elizabeth Alm, Mark McBride, Matthew Smith

Abstract

Fecal indicator bacteria (FIB) such as Escherichia coli (E. coli) and enterococci are used to assess microbiological water quality in recreational waters. The use of FIB follows the assumption that their presence correlates with that of fecal-associated pathogens in recreational waters. The beach ecosystem is complex however and multiple factors can influence the concentration of E. coli and enterococci in the beach environment. Microbial communities within beach sand play a key role in nutrient cycling and are important to the nearshore ecosystem function. E. coli and enterococci, two common indicators of fecal pollution, have been shown to persist in the beach sand, but little is known about how microbial community assemblages are related to these fecal indicator bacteria (FIB) reservoirs. The first objective of this project was to harness the power of next-generation sequencing to profile the indigenous communities within beach sand and examine key environmental drivers of community structure. FIB were found at similar levels in sand at beaches adjacent to urban, forested, and agricultural land and in both the berm and backshore. However, there were striking differences in the berm and backshore microbial communities, even within the same beach, reflecting the very different environmental conditions in these beach zones in which FIB can survive. Results indicate that microbial community structure in beach sand is most associated with the concentrations of total organic carbon (TOC) and total phosphorus (TP). Fine scale nucleotide differences in the V4V5 region of the 16S rRNA gene of abundant taxa were identified and sequence patterns suggest a biogeographic influence. This work demonstrates that microbial communities are reflective of environmental conditions at freshwater beaches and are able to provide useful information regarding long-term anthropogenic stress. The second objective of this project was to use host-specific alternative fecal indicator assays to identify the major pollution sources that are responsible for contributing high levels of E. coli in both beach sand and water. At the six beaches studied, berm sand contained the highest levels of E. coli versus to water (P < 0.01), using a weight-to-volume comparison. The gull-specific assay (Gull2) was detected more than any other host-specific alternative fecal indicator assay with 80% detection in water samples during water quality advisories. Human-specific Bacteroides (HB) and Lachnospiraceae (Lachno2) were detected in only 2.4% of water samples, however a large number of sand samples had an uncoupled occurrence of the two human-specific alternative indicators. Results from in situ microcosm experiments indicate that the HB and Lachno2 markers decay at different rates, helping explain their differential occurrence in environmental samples. In situ microcosm experiments also revealed that signals from the alternative indicators decay approximately 20% faster than culturable E. coli. Overall, a significant amount of the E. coli burden in sand cannot be accounted for with the use of host-specific alternative indicators suggesting that E. coli concentrations in sand are uncoupled from fecal sources and that E. coli may be able to persist in beach sand post-deposition. The final objective of this project is to assess the survival of different E. coli isolates and to identify possible genomic characteristics that may support a persistence phenotype. In situ survival experiments revealed that the die-off of a non-environmental E. coli type strain was significantly faster than an environmental E. coli strain isolated from beach sand. Comparative genomics suggested that biofilm formation and programmed cell death might be important mechanisms supporting the increased survival of the environmental E. coli strain. Overall, the findings presented in this dissertation provide new insights into the environmental ecology of enteric bacteria, highlighting the importance of nutrients, land-use, the indigenous microbial community, and genomic elements as determinants of the fate of FIB in the beach environment.

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