Dairy farm soil presents distinct microbiota and varied prevalence of antibiotic resistance across housing areas.

Dairy cattle of different ages experience different living conditions and varied frequency of antibiotic administration that likely influence the distribution of microbiome and resistome in ways that reflect different risks of microbial transmission. To assess the degree of variance in these distributions, fecal and soil samples were collected from six distinct housing areas on commercial dairy farms (n = 7) in Washington State. 16S rRNA gene sequencing indicated that the microbiota differed between different on-farm locations in feces and soil, and in both cases, the microbiota of dairy calves was often distinct from others (P < 0.05). Thirty-two specific antibiotic resistance genes (ARGs) were widely distributed on dairies, of which several clinically relevant ARGs (including cfr, cfrB, and optrA) were identified for the first time at U.S. dairies. Overall, ARGs were observed more frequently in feces and soil from dairy calves and heifers than from hospital, fresh, lactation and dry pens. Droplet-digital PCR demonstrated that the absolute abundance of floR varied greatly across housing areas and this gene was enriched the most in calves and heifers. Furthermore, in an extended analysis with 14 dairies, environmental soils in calf pens had the most antibiotic-resistant Escherichia coli followed by heifer and hospital pens. All soil E. coli isolates (n = 1,905) are resistant to at least 4 different antibiotics, and the PFGE analysis indicated that florfenicol-resistant E. coli is probably shared across geographically-separated farms. This study identified a discrete but predictable distribution of antibiotic resistance genes and organisms, which is important for designing mitigation for higher risk areas on dairy farms.