Zoological gardens as a neglected One Health interface for tigecycline-resistant Escherichia coli

The tigecycline resistance gene tet(X4) is increasingly reported from food-producing animals and humans, but its circulation at wildlife-environment interfaces, including zoos, remains poorly resolved. Here, we combined systematic sampling of zoo animals and zoo-associated environments with comparative genomics against a global collection of 1102 tet(X)-positive Escherichia coli genomes to characterize the occurrence, genomic context and potential dissemination of tet(X4) in zoological settings. We recovered 60 tet(X4)-positive isolates (58 E. coli and two Enterobacter hormaechei) from 188 samples, predominantly from animal feces (91.7%), with frequent detection in kangaroos and wolves. All isolates were resistant to tigecycline and commonly multidrug-resistant; 26/60 (43.3%) transferred tet(X4) by conjugation. Whole-genome sequencing showed that tet(X4) was primarily plasmid-borne within an ISCR2/IS1-associated module and was mainly carried by IncX1 backbones, including cointegrate plasmids, consistent with plasmid-mediated horizontal transfer and plasmid remodeling. Core-genome SNP phylogenies identified several near-identical clonal clusters (0-2 SNPs), indicating potential local dissemination within the zoo. Importantly, global phylogenomic analysis revealed that several zoo-associated isolates were phylogenomically close to strains from food-producing animals and farm-associated environments in China (<70 core-genome SNPs), suggesting possible ecological connectivity between zoo-associated and agro-environmental reservoirs. Together, these findings position zoos as an under-recognized mixing interface where tet(X4)-positive lineages and mobile elements may be introduced and maintained, underscoring the need for integrated One Health surveillance spanning animals, husbandry inputs and enclosure-associated environments.