Emergence of a conjugative hybrid plasmid co-harboring mcr-1.1, mcr-3.39, and tet(X4) in Escherichia coli from wastewater in Seoul: A potential threat to last-line antibiotics

The emergence of plasmid-mediated antimicrobial resistance to last-line antibiotics such as colistin and tigecycline poses an escalating public health threat. In this study, we identified an Escherichia coli strain (WWCOL-256) isolated from municipal wastewater in Seoul, South Korea, co-harboring the mcr-1.1, mcr-3.39, and tet(X4) genes. Genomic analysis revealed that these resistance genes were located on a 237.9-kb IncFIA(HI1)–HI1A–HI1B(R27) hybrid plasmid (pCOL256–1), with the tet(X4) gene embedded within an ISVsa3-mediated transposon [∆ISVsa3–estT–tet(X4)–ISVsa3] and mcr-1.1 associated with the ISApl1–mcr-1.1–pap2 structure. The mcr-3.39 gene (∆TnAs2–mcr-3.39–dgkA–ISKpn40) was likely acquired from a co-resident IncFII plasmid (pCOL256–2). Functional assays demonstrated that the plasmid pCOL256–1 can be transferred via conjugation and remains stable without antibiotic selection, underscoring its potential to disseminate colistin and tigecycline resistance. Although global surveillance has detected the frequent co-occurrence of the mcr and tet(X) genes, their co-localization on the same plasmid remains rare. To the best of our knowledge, this is the first report of a single plasmid co-harboring the mcr-1, mcr-3.39, and tet(X4) genes, highlighting the urgent need for enhanced environmental genomic surveillance to prevent the spread of multidrug resistance determinants.