Direct fecal deposits from cattle provide a significant source of E. coli to streams and therefore pose a threat to human health in agricultural watersheds. Experiments were conducted in a flume (9.1 m long, 0.6 m wide, and 0.6 m deep) with flow of 0.0106 m3 s-1 , an average velocity of 11.4 cm s-1 ,and water depth of 15.24 cm to measure the resuspension and deposition of E. coli from an undisturbed standard cowpat. Water samples were collected 1.22 m and 3.66 m downstream of the deposited cowpat, and at each downstream cross-section nine samples were collected to characterize the bacterial movement. E. coli in water samples were separated into the attached and unattached phases by filtration to assess the mechanism of transport. The cumulative load contribution from a single deposited cowpat after one hour was 2.49×10 9 cfu 3.66 m downstream. The composite E. coli concentrations at all sampling points and times exceeded the federal standards for primary contact in the United States of 126 cfu/100 ml. Between 77.2 and 99.5% of all E. coli downstream of the direct deposit were associated with particulates. The resuspension rate was 5.91×107 and 9.52×104 cfu m-2 s-1 0.5 min and 60 minutes after deposition, respectively, 1.22 m downstream of the deposit and 2.19×106 and 3.14×103 cfu m-2 s-1 0.5 min and 60 min after deposition, respectively, 3.66 m downstream of the deposit. Results from this study are useful to improve modeling techniques to predict in-stream E. coli concentrations from direct fecal deposits and emphasize the need to implement management practices to reduce livestock access to streams.