Salmonella enterica and Escherichia coli are bacteria of concern to veterinary public health and poultry health. Our research aimed to determine the factors associated with S. enterica and E. coli in commercial broiler chicken barns during the rest period between flocks to identify the best methods of sanitation for bacterial load reduction. This involved collecting samples from September 2015 to July 2016 from the floors of 36 barns before sanitation (baseline) and at 2 time intervals after sanitation, followed by microbiological and molecular analysis. A priori variables of interest included sanitation procedure (dry cleaning, wet cleaning, disinfection), sampling point (baseline, 2 d after sanitation, 6 d after sanitation), and flooring type (concrete, wood). The odds of detecting S. enterica were higher on wooden floors that were wet-cleaned than on concrete floors that were dry-cleaned, lower in the winter and spring than in the fall, and lower when samples were collected 2 d and 6 d after sanitation than at baseline. For E. coli , the concentration was higher on wooden floors than on concrete floors and in the summer than in the fall, and it was lower in postsanitation samples from disinfected barns than in presanitation samples from dry-cleaned barns and in the winter than in the fall. Among E. coli isolates, factors associated with the presence of qacEΔ1 , a gene associated with resistance to quaternary ammonium compounds, included sanitation procedure, flooring type, cycle length, and the number of times per yr the barn is disinfected. Our findings highlight the importance of cleaning after litter removal, although the sanitation procedure chosen might differ depending on which pathogen is present and causing disease issues; dry cleaning appears to be preferable for S. enterica control, especially in barns with wooden floors, whereas disinfection appears to be preferable for E. coli reduction.