Proposal Title : A Molecular Approach to Determine the Origin of Fecal Bacteria in Catoma Creek of the Alabama River Basin

Many of Alabama's rivers and streams do not meet the Clean Water Act's "swimmable" goal due to fecal contamination. Catoma Creek in Montgomery County has been included on Alabama's Section 303(d) List due to elevated numbers of fecal coliform bacteria and organic enrichment. Both urban and agricultural land uses exist in the watershed. The objectives of this research were to construct a library of rep-PCR DNA fingerprints from Escherichia coli strains isolated from human and animal feces, enumerate and isolate E. coli in the water samples collected monthly from the watershed, and identify the sources of fecal contamination in the Catoma Creek watershed using the constructed library. We isolated and characterized a total of 484 E. coil isolates from cattle, chicken, horse, waterfowl, deer, wild turkey, dog, and human fecal samples. Rep-PCR DNA fingerprints of the E. coli isolates were obtained and construction of the rep-PCR DNA fingerprint library of known sources will soon be completed. Monthly water samples were collected from May 2003 to April 2004 at 8 locations in the Catoma Creek watershed. Data showed that E. coli densities ranged from 18 to 12,650 CFU/100 ml, with 70% of the samples exceeding the EPA criterion for swimming waters. There was a high correlation between£. coli density and stream flow. Chemical analyses ofthe water samples showed that the concentration ranges for ~-N, N03-N, phosphorus, and potassium were 0.19 to 0.78 mg/1, 0.12 to 0.38 mg/1, 0.13 to 0.65 mg/1, and 0.16 to 4.74 mg/1, respectively. Relatively high concentrations of phosphorus suggest that eutrophication may occur in the watershed. During the same period, 500 E coli isolates were also isolated from the water samples and rep-PCR DNA fingerprints of some water E. coli isolates were obtained. Once the sources of fecal contamination in the Catoma Creek watershed are determined, the corrective actions can be developed to target the contamination source(s), which will lead to improved water quality. Statement of the Problem and Research Objectives Fecal pollution impairs the quality of streams and rivers for recreational use and adversely affects fish and aquatic life. Fecal contamination can originate from both human and non-human sources including surface runoff from land application of animal wastes or farm animal feedlots, inadequate septic or sewer systems, improper waste disposal, and wildlife impact. Determining the sources of fecal contamination is necessary to develop effective pollution control strategies. Presence of fecal indicator bacteria is the second most common cause of surface water impairment in Alabama. Thirty percent of the 181 impaired water bodies in the state are attributed to fecal indicator bacteria according to Alabama's 2002 Section 303 (d) List (ADEM, 2003). Catoma Creek in Montgomery County is a tributary of the Alabama River. Both agricultural and urban land uses exist in the watershed. A 23-mile segment of Catoma Creek has been included on the State 303( d) list due to impairment of fecal pollution (ADEM, 2002). Urban runoff and pasture grazing are suspected to be the main sources of fecal contamination. The local citizen volunteer group, Montgomery Water Watch, has been monitoring water quality parameters (i.e., pH, alkalinity, dissolved oxygen, turbidity, and temperature) along the creek. The objectives of this research were to construct a library ofrep-PCR DNA fingerprints from Escherichia coli strains isolated from a wide range of collection of human and animal feces, enumerate and isolate E. coli in the water samples collected monthly from the watershed, and identify the sources of fecal contamination in the Catoma Creek watershed using the constructed library.