Tandem amplification of the vanM gene cluster drives vancomycin resistance in vancomycin-variable enterococci.

Background Vancomycin-variable enterococci (VVE) are a potential risk factor for vancomycin resistance gene dissemination and clinical treatment failure. vanM has emerged as a new prevalent resistance determinant among clinical enterococci in China. A total of 54 vancomycin-susceptible enterococci (VSE) isolates carrying incomplete vanM gene clusters were isolated in our previous study. Objectives To determine the potential of vanM-carrying VSE to develop vancomycin resistance and investigate the mechanism of alteration of the resistance phenotype. Methods Fifty-four vanM-positive VSE strains were induced in vitro by culturing in increasing concentrations of vancomycin. Genetic changes between three parent VVE strains and their resistant variants were analysed using Illumina and long-read sequencing technologies, quantitative PCR and Southern blot hybridization. Changes in expression level were determined by quantitative RT-PCR. Results Twenty-five of the 54 VSE strains carrying vanM became resistant upon vancomycin exposure. A significant increase in vanM copy number was observed ranging from 5.28 to 127.64 copies per cell in induced resistant VVE strains. The vanM transposon was identified as tandem repeats with IS1216E between them, and occurred in either the plasmid or the chromosome of resistant VVE cells. In addition, an increase in vanM expression was observed after resistance conversion in VVE. Conclusions This study identified tandem amplification of the vanM gene cluster as a new mechanism for vancomycin resistance in VVE strains, offering a competitive advantage for VVE under antibiotic pressure.