This study was conducted to determine if a relationship exists between temperament, stress response, and the shedding of Escerhichia coli O157:H7. Cattle (n = 150) were evaluated for disposition and stress response before shipping to the feeding operation, upon arrival at the feedlot, at approximately 70d on feed, and prior to transport to the harvesting facility. Chute and pen scores, as well as serum cortisol concentrations, were measured in order to assess individual temperament and stress response. A temperament index was created to classify cattle as Excitable, Intermediate, or Calm. The presence of E. coli O157:H7 was determined by rectal swabs on the live cattle and swabs of colons collected postmortem at the processing facility. As expected, variables for pre-shipment temperament index, exit velocity, pen score, arrival and midpoint exit velocity, and mid-point cortisol concentrations differed (P < 0.05) greatly between temperament groups. However, pre-shipment chute scores and cortisol concentration, as well as arrival and final cortisol concentrations differed (P < 0.05) only for Excitable cattle compared to both Calm and Intermediate groups. The percentage of cattle shedding the pathogen at arrival was approximately equal between temperament groups. When sampled before shipment to the processing facility, a higher proportion (P = 0.03) of cattle displaying Calm temperaments shed E. coli O157:H7 than the other groups. Results from postmortem colon samples exhibited a similar trend. When the results from all four sampling periods were pooled, the Calm cattle had a greater numerical percentage test positive for E. coli O157:H7. However, the pooled frequency distribution is largely dictated by the results of the final sampling time. Based on these results, it appears that Excitable cattle are not more likely to shed E. coli O157:H7. In fact, it seems that Calm cattle may be equally or more susceptible to shed at later points in the feeding period. However, it is important to note that a relatively small number of the samples tested positive for E. coli O157:H7, thus, potentially causing dramatic changes in the distributions.