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Nutrient and microbial transport by runoff may vary at different locations within a beef cattle feedlot. If the areas making the largest contributions to nutrient and microbial transport can be identified, it may be possible to institute site‐specific management practices to reduce runoff nutrient and microbial transport. The objectives of this study were to: (1) measure selected feedlot soil properties and nutrient and microbial transport in runoff from various feedlot locations, (2) compare the effects of unconsolidated surface materials (USM) (loose manure pack) and consolidated subsurface materials (CSM) (compacted manure and underlying layers) on nutrient and microbial transport, and (3) determine if nutrient and microbial transport in runoff are correlated to selected feedlot soil characteristics. Simulated rainfall events were applied to 0.75 m wide by 2 m long plots. No significant differences (P < 0.05) in feedlot soil characteristics or nutrient transport in runoff were found between USM and CSM. However, concentrations of E. coli were significantly greater in the USM than the CSM. Pen location was found to significantly influence feedlot soil measurements of Bray‐1 P, calcium, chloride, copper, electrical conductivity (EC), loss on ignition, organic N, phosphorus, potassium, sodium, sulfur, total N (TN), water‐soluble P, and zinc. Runoff measurements of dissolved phosphorus (DP), EC, and NH4-N were significantly influenced by pen location and were correlated to selected feedlot soil characteristics. Thus, it may be possible to estimate DP, EC, and NH4-N in runoff from selected feedlot soil parameters.