Investigating the mobility and host range of mobile genetic elements harbouring antimicrobial resistance genes in enterococci

In this study, the abundance and conjugation capacity of mobile genetic elements (MGEs) carrying resistance genes such as vanA , tet (M) and erm (B) were investigated to enhance our understanding of antimicrobial resistance (AMR) dissemination across the One Health continuum in high-priority, highly prevalent enterococcal pathogens. The abundance of MGEs was estimated using replicon typing and both reference-based and reference-free clustering approaches. Conjugation potential was assessed using agar plate mating between Enterococcus faecium donors and E. faecium , Enterococcus faecalis and Enterococcus hirae recipients, with conjugated MGE verified via long-read sequencing. Key findings include the identification of a vanA gene cluster from E. faecium VRE0008 associated with a Tn 1546 -like transposon embedded in a RepA_N-type putative plasmid (232,902 bp). This plasmid successfully conjugated with E. faecium , E. faecalis and E. hirae recipients from clinical, environmental and agricultural sources. The transfer predominantly involved the modular movement of a 46-kb region surrounding the vanA gene cluster, with E. hirae of agricultural origin (i.e. 0093A) being the exception, as it retained the entire plasmid. The tet (M) gene from E. faecium Ent0189 was located on a putative Rep_Trans-like plasmid, with features of Tn 916 integrative conjugative elements. The entire plasmid from Ent0189 was successfully transferred to intra-species recipients from clinical and environmental sources, but transfer to E. faecalis and E. hirae was less common. Attempts to transfer tet (M) associated with Tn 916 from bovine E. hirae to any of the E. hirae , E. faecium and E. faecalis isolates were unsuccessful. Additionally, the erm (B) gene from E. faecium NS0794 was carried by an MGE matching the RepA_N-type plasmid, but lacking the vanA gene cluster. Successful conjugative transfer of this plasmid was observed with E. faecium , E. faecalis and E. hirae of various origins, except one clinical E. faecalis isolate. These findings highlight the broad conjugation capabilities and modular mobility of MGEs carrying ARGs in enterococci, enhancing our understanding of dynamic MGE-mediated ARG dissemination and informing strategies to address the spread of AMR between species and habitats.