To comparatively analyze the molecular characteristics of linear plasmids in Enterococcus avium, clarify the global transmission trajectory of linear plasmids in Enterococcus, and explore the genetic stability of linear plasmids in different host strains through subculture of transconjugants, thereby providing a theoretical basis for the prevention and control of clinically drug-resistant bacteria.

The minimum inhibitory concentrations (MICs) of vancomycin against Enterococcus avium ZB195, three recipient strains (Enterococcus avium 07H170, Enterococcus faecalis FA2-2, and Enterococcus faecium BM4105RF), and their corresponding transconjugants were determined using the broth microdilution method. Whole-genome sequencing of strain ZB195 was performed to characterize its plasmid features. Linear plasmid sequences from 18 global Enterococcus faecium strains isolated between 2000 and 2019 were collected, and phylogenetic trees and Bayesian evolutionary trees were constructed using MAFFT, IQtree, and BEAST software. The global transmission trajectory was simulated with SpreaD3 software. The genetic stability of linear plasmids in different host strains was evaluated through a 15-day subculture experiment without antibiotic selection pressure.

For the first time, this study identified a linear plasmid in the genome of Enterococcus avium ZB195, which harbors the vancomycin resistance vanA gene and was capable of horizontal transfer to three recipient strains. Sequence alignment showed that the linear plasmid of ZB195 had the highest sequence identity with that of Enterococcus faecium AA708 from Japan, and phylogenetic analysis confirmed that they were most closely related. Simulation of global transmission trajectories revealed that the linear plasmid in Enterococcus was first detected in Japan in 2000, and then gradually spreaded to China, the United States, Norway, Denmark, India and other countries. Passage experiments indicated that the linear plasmid exhibited the highest stability in Enterococcus avium hosts.

The linear plasmids detected in Enterococcus strains in China may have originated from the horizontal transmission of Enterococcus faecium from Japan. Moreover, after transferring to Enterococcus avium, these plasmids can form better host adaptability. This suggests that it is necessary to strengthen cross-regional monitoring of linear plasmids in Enterococcus to curb the global spread of drug-resistant genes.