Tigecycline is one of the most critical drugs for treating Gram-negative bacterial infections; however, the emergence of the tigecycline resistance efflux pump TMexCD1-TOprJ1 poses a global health threat. The evolutionary relationships and epidemiological trends of tmexCD1-toprJ1-positive strains across various ecological niches remain largely unexplored. In this study, we employed whole-genome sequencing (WGS) of tmexCD1-toprJ1-positive bacteria from humans, food, animals, and the environment in China to assess the epidemiological and genomic features of these strains, analyzing both newly collected strains and data from the GenBank database. From 3 434 samples collected during 2019–2022, 145 tmexCD1-toprJ1-carrying strains (4.5%) were isolated. The majority of the tmexCD1-toprJ1-positive Enterobacterales exhibited resistance to nearly all antimicrobials, including colistin (42.13%). tmexCD1-toprJ1 was predominantly identified in Klebsiella pneumoniae (K. pneumoniae) from chicken feces in China but was also detected in multiple ecological niches and other countries. Phylogenetic analysis revealed the clonal transmission of tmexCD1-toprJ1-positive ST37 K. pneumoniae across diverse ecological niches as well as the international spread of the ST15 K. pneumoniae clone-producing TMexCD1-TOprJ1. tmexCD1-toprJ1 is mainly carried by Klebsiella spp. specific narrow host range plasmids, which may limit the spread of tmexCD1-toprJ1 across different bacterial species. Notably, due to the fitness cost posed by tmexCD1-toprJ1, the occurrence of tmexCD1-toprJ1-positive Enterobacterales in both food animals and humans in China has declined significantly following the withdrawal of antibiotics as growth promoters in food animals in China since 2020. However, tmexCD1-toprJ1 has been captured by broad-host-range plasmids and hypervirulent carbapenem-resistant K. pneumoniae ST11-KL64 strains in healthcare settings. The frequent use of tetracyclines in chicken farming likely contributes to the high detection rate of tmexCD1-toprJ1; therefore, to reduce the threat of tmexCD1-toprJ1-positive K. pneumoniae, continuous monitoring of tmexCD1-toprJ1 across different ecological niches and strict enforcement of antimicrobial policies in animal husbandry, particularly in the poultry industry, are urgently required.