The emergence of antibiotic-resistant bacteria has created a significant threat to global health. Some strains of Escherichia coli (E. coli), a common bacteria with both commensal and pathogenic strains, can be resistant to common antibiotics. This study investigates the prevalence of E. coli resistance to ampicillin, erythromycin, and ciprofloxacin in Waller Creek, an urban waterway in Austin, Texas with chronically high levels of fecal bacteria. Approximately 134 samples were collected from three different sites along Waller Creek and spiked with varying antibiotic concentrations. E. coli resistance to ampicillin and erythromycin was found to be up to 26.92% and 48.37% respectively, while resistance to ciprofloxacin was much lower ranging between 0%-2%. The highest levels of resistance were associated with dense urbanization, high levels of homelessness, and after heavy precipitation. These findings indicate the need for regular monitoring of antibiotic concentrations and antibiotic resistant fecal bacteria within urban waterways. This data is needed to identify potential human health risks and understand key factors that contribute to the quantity of antibiotic-resistant pathogens.