Prophage-encoded antibiotic resistance genes (ARGs) are enriched in human-impacted environments. A global analysis of 38,605 bacterial genomes, 1,432 metagenomes, and 1,186 metatranscriptomes across 12 habitats revealed that human activities significantly increase the abundance, diversity, and activity of prophage-encoded ARGs in environments with higher antibiotic exposure. Prophages, which can integrate into bacterial genomes, often carry ARGs that can be mobilized to enhance resistance in heterologous hosts. This study shows that human activities have altered bacteria-phage interactions, enriching ARGs in prophages and increasing their mobility across habitats. Prophages serve as a reservoir for ARGs, which can be transferred between bacterial species and environments. The presence of ARGs in prophages allows bacteria to persist and adapt to antibiotic exposure, contributing to the development of antibiotic-resistant strains. The study also found that prophage-encoded ARGs are more abundant and active in human-impacted environments, with higher transmission potential and wider geographical distribution. These findings highlight the role of prophages in the global spread of ARGs and the need for further research to understand the impact of human activities on antibiotic resistance.Prophage-encoded antibiotic resistance genes (ARGs) are enriched in human-impacted environments. A global analysis of 38,605 bacterial genomes, 1,432 metagenomes, and 1,186 metatranscriptomes across 12 habitats revealed that human activities significantly increase the abundance, diversity, and activity of prophage-encoded ARGs in environments with higher antibiotic exposure. Prophages, which can integrate into bacterial genomes, often carry ARGs that can be mobilized to enhance resistance in heterologous hosts. This study shows that human activities have altered bacteria-phage interactions, enriching ARGs in prophages and increasing their mobility across habitats. Prophages serve as a reservoir for ARGs, which can be transferred between bacterial species and environments. The presence of ARGs in prophages allows bacteria to persist and adapt to antibiotic exposure, contributing to the development of antibiotic-resistant strains. The study also found that prophage-encoded ARGs are more abundant and active in human-impacted environments, with higher transmission potential and wider geographical distribution. These findings highlight the role of prophages in the global spread of ARGs and the need for further research to understand the impact of human activities on antibiotic resistance.