Understanding sediment Escherichia coli levels (i.e., pathogen indicators) and their contribution to the water column during resuspension is critical for predicting in-stream E. coli levels and the potential risk to human health. The U.S. Environmental Protection Agency's current water quality testing strategies, however, rely on water borne E. coli concentrations to assess stream E. coli levels and identify impaired waters. In this work, we conducted a scenario analysis using a range of flows, sediment/water bacteria fractions, and particle sizes to which E. coli attach to assess the impact of E. coli in streambed sediments on water column E. coli levels. We used simple sediment transport theory to calculate the potential total E. coli concentrations in a stream with and without the resuspension process. Results clearly indicate that inclusion of resuspending sediment attached E. coli is necessary for watershed assessments and data on sediment attached E. coli concentrations is much needed. When neglecting the streambed sediment E. coli concentrations, the model predicted average E. coli loads of 107 Colony Forming Units (CFU)/s; however, when streambed sediment E. coli concentrations were included in the model, the predictions ranged from 1010 to 1014 CFU/s. To evaluate the predictions, E. coli data in the streambed sediment and the water column were monitored in Squaw Creek, Iowa. Comparisons between measured and predicted E. coli loads yielded an R2-value of 0.85.