Infections caused by multidrug resistant bacteria represent a therapeutic challenge both inclinical settings and in livestock production, but the prevalence of antibiotic resistance genesamong the species of bacteria that colonize the gastrointestinal tract of ruminants is not wellcharacterized. Here, we investigate the resistome of 435 ruminal microbial genomes in silicoand confirm representative phenotypes in vitro. We find a high abundance of genes encodingtetracycline resistance and evidence that the tet(W) gene is under positive selective pressure. Our findings reveal that tet(W) is located in a novel integrative and conjugative elementin several ruminal bacterial genomes. Analyses of rumen microbial metatranscriptomesconfirm the expression of the most abundant antibiotic resistance genes. Our data provideinsight into antibiotic resistange gene profiles of the main species of ruminal bacteria andreveal the potential role of mobile genetic elements in shaping the resistome of the rumenmicrobiome, with implications for human and animal health.