We sequenced the 16 metagenomes using the whole-genome shotgun approach and generated 157 Gbp of metagenomic sequences. Through de novo assembly and annotation, we obtained a 13.1 Gbp reference contig assembly and identified 22 million microbial genes for cattle rectum microbiota. We discovered a significant reduction of microbial diversity after toxic seed treatment (P < 0.01), suggesting dysbiosis of the microbiome. Six bacterial families and 31 species are significantly increased in the fecal microbiota (P-adjusted < 0.05), including members of the top abundant rumen core taxa. This global elevation of rumen microbes in the rectum microbiota suggests a potential impairment of rumen microbiota under fescue toxicosis. Among these, Ruminococcaceae bacterium P7, an important species accounting for ~2% of rumen microbiota, was the most impacted with a 16-fold increase from 0.17% to 2.8% in feces (P < 0.01). Functional enrichment analysis revealed that the overrepresented pathways shifted from energy metabolism to antimicrobial resistance and DNA replication after the consumption of toxic fescue seeds.