Gut microbiota is a reservoir of antibiotic resistance genes (ARGs). Yet, limited information is available regarding the presence (metagenomic DNA level) and expression profiles (metatranscriptomic RNA level) of ARGs in gut microbiota. Here, we used both metagenomic and metatranscriptomic approaches to comprehensively reveal the abundance, diversity, and expression of ARGs in human, chicken, and pig gut microbiomes in China. Based on deep sequencing data and ARG databases, a total of 330 ARGs associated with 21 antibiotic classes were identified in 18 human, chicken, and pig fecal samples. Metatranscriptomic analysis revealed that 49.4, 66.5, and 56.6% of ARGs identified in human, chicken, and pig gut microbiota, respectively, were expressed, indicating that a large proportion of ARGs were not transcriptionally active. Further analysis demonstrated that transcript abundance of tetracycline, aminoglycoside, and beta-lactam resistance genes was mainly contributed by acquired ARGs. We also found that various biocide, chemical, and metal resistance genes were actively transcribed in human and animal guts. The combination of metagenomic and metatranscriptomic analysis in this study allowed us to specifically link ARGs to their transcripts, providing a comprehensive view of the prevalence and expression of ARGs in gut microbiota. Taken together, these data deepen our understanding of the distribution, evolution, and dissemination of ARGs and metal resistance genes in human, chicken, and pig gut microbiota.