American Alligator Fecal Coliform and Pathogen Identification, Genotypic Fingerprinting, and REP-PCR Bacterial Source Tracking in South Carolina

Research for this study was performed to assess numerous coastal nonpoint source pollution components from poikilothermic (i.e. cold-blooded) wildlife sources, including fecal coliform identification, genotypic fingerprinting, and bacterial source tracking. The overall goals were to identify, fingerprint, and detect American alligator (Alligator mississippiensis) fecal coliform bacteria in surface waters, as alligators may be an unrecognized source of fecal pollution and/or potential pathogens in coastal South Carolina surface waters. The research was divided into three sections, where each component improves our knowledge of the contribution of fecal bacteria from poikilothermic animals. The first research component determined the accuracy of two Enterobacteriaceae identification techniques, the API 20E biochemical test system and 16S rRNA gene sequencing methods, in the identification of alligator cloacal isolates. The second component evaluated the ability of Repetitive Extragenic Palindromic PCR (REP-PCR) to produce identifiable, unique, and stable genomic fingerprints from alligator fecal coliform and potential pathogenic bacteria. The final component determined if alligator fecal coliform and potential pathogenic bacteria contributed to water quality degradation in coastal South Carolina surface waters using REP-PCR as a source tracking tool. Results from this research show that when 16S rRNA gene sequencing methods are utilized, the predominant enteric bacteria identified from alligator fecal samples include: Aeromonas spp., Citrobacter freundii, Edwardsiella tarda, Enterobacter spp., Escherichia coli, Plesiomonas shigelloides, and Salmonella enterica. Alligator fecal coliform bacteria can also be genotypically characterized with REP-PCR using the BOX A1R primer. Finally, alligators are a source of fecal coliform bacteria in South Carolina surface waters, as alligator REP-PCR fingerprints were similar to bacteria fingerprints isolated from surface waters. By attempting to identify, characterize, and match alligator bacterial fingerprints to those found in surface water samples, this research increases our understanding of bacterial sources and source tracking tools, indicating that alligators are a pollution source of enteric bacteria and potential pathogens in coastal surface waters in the southeastern United States. This research will hopefully demonstrate that there is a continued need to study and develop effective bacterial source tracking tools and indicators to help identify and manage sources that contribute to fecal pollution in coastal surface waters