Emergence of two novel tmexCD-toprJ subtypes mediating tigecycline resistance in the megaplasmids from Pseudomonas putida.

The widespread antimicrobial resistance (AMR) problem poses a serious health threat, leaving few drug choices, including tigecycline, to treat multidrug resistance pathogens. However, a plasmid-borne tigecycline resistance gene cluster, tmexCD1-toprJ1, emerged and conferred tigecycline resistance. In this study, we identified two novel subtypes, tmexCD2.3-toprJ2.3 and tmexCD2.4-toprJ1b, obtained from three chicken-origin Pseudomonas putida isolates. Two types of megaplasmids were found as the vital vehicle of these tmexCD-toprJ variants. Phylogenetic and genomic analysis indicated the two variants were mainly distributed in Pseudomonas and acted as an evolved intermediated state precursor of tmexCD2-toprJ2. Further gene cloning assay revealed both the expression of tmexCD2.3-toprJ2.3 and tmexCD2.4-toprJ1b could confer multiple antimicrobial resistance, mediating 8- to 16-fold increase of tigecycline MIC. Importantly, two key nucleotide differences in promoter region influence the promoter activity between P_tmexC2.3 and P_tmexC2.4, while the downregulation effect of TNfxB on the transcriptional expression level of tmexCD2.3-toprJ2.3 and tmexCD2.4-toprJ1b were observed. The emergency of two novel tmexCD-toprJ variants necessitates preventive measures to curb their spread and highlights concerns about more emerging tmexCD-toprJ variants.