Antibiotic resistant bacteria are widespread in songbirds across rural and urban environments.

The widespread use of antibiotics in human and veterinary medicine to treat pathogenic bacteria has resulted in the rapid emergence of antibiotic-resistant bacteria (ARB). Wild animals may enable the spread of pathogenic and non-pathogenic ARB when they are exposed to reservoirs (e.g., contaminated soil, water, or crops) and carry ARB in and on their bodies to other environments. We tested for the presence of ARB in four songbird species in southwest Michigan across a gradient of land use. Our specific objectives were to: 1) quantify the prevalence of ARB found in the gut microbiome of birds; 2) identify the specific bacteria exhibiting resistance; 3) assess whether ARB prevalence and identity varied among bird species; and 4) assess whether anthropogenic land use influenced the prevalence and identity of ARB found on birds. We sampled birds across a land use gradient consisting of urban, agricultural, and natural land covers using a randomized, spatially-balanced sampling design and cultured bacteria from fecal samples in the presence of three different antibiotics (amoxicillin, tetracycline, and ciprofloxacin). Overall prevalence of ARB was high, with 88% of total birds carrying ARB resistant to one of three antibiotics that we tested. Resistance to amoxicillin was more common (83% of sampled birds) than resistance to tetracycline (15%) or ciprofloxacin (1%). Identified ARB were diverse, and included 135 isolates representing 5 bacterial phyla and 22 genera. There was no effect of land use on ARB prevalence, with 90% of sampled birds captured in rural sites and 85% of sampled birds in urban sites carrying ARB. We provide the first analysis of ARB prevalence across multiple bird species and land uses utilizing a spatially-balanced, randomized study design. Our results demonstrate that nearly all sampled birds carried at least some ARB, and that they may serve as important dispersal agents of ARB across large spatial scales.