Enterococcus faecium can cause nosocomial infections associated with intravenous catheter, invasive infections in the abdomen, and bacteraemia [1]. A Danish cohort study investigated enterococcal bacteraemia and found the incidence of E. faecium bacteraemia to be 6.6/100,000 person-years, and the 30-day mortality in this population to be 34.6% [2]. In another Danish study, the authors founds no difference in the 30-day mortality between patients with vancomycin-resistant E. faecium (VREfm) bacteraemia, and patients with vancomycin-susceptible E. faecium bacteraemia (37.6% vs.37.0%) [1]. Te in-hospital mortality rate of VREfm infections is 18%, risin to above 69% at 90 ays [3,4,5].

Vancomycin-resistance in E. faecium from human isolates is mainly caused by the acquisition of vanA or vanB. Studies have shown that the van operons can be transmitted between E. faecium strains by horizontal gene transfer [6, 7]. In this study, we focus on the vanA gene, which is located on Transposon 1546 (Tn1546) and most often carried on plasmids, and the vanB gene, which is located on Transposon 1549 (Tn1549), inserted in the E. faecium chromosome, or rarely on plasmids [8, 9].

From 2013, Denmark observed an increase in vancomycin-resistant E. faecium (VREfm) due to the emergence of several vanA VREfm clones. From 2019, vanB VREfm started to increase and outcompeted vanA VREfm [10, 11].

The emergence of vanB VREfm has challenged rapid diagnostic of VREfm from rectal screening samples by PCR since vanB is present in multiple gut commensals other than E. faecium. As previously described, a multiplex PCR, targeting both vanB, and the specific Tn1549 insertion sites in the chromosome in the two most dominating vanB clones (a L-arabinose isomerase gene (araA2), and a silent information regulator gene (sir2) in our area, was developed at the DCM Hvidovre Hospital to overcome this diagnostic challenge [8, 12].

Accurate and rapid diagnostic of VREfm performed directly on rectal swabs (within 12 h) is crucial to maintain infectious hygiene precautions in a hospital setting, and to prevent hospital outbreaks [13]. In our regions, we carry out contact isolation precautions for all patients that are carriers of or infected with VREfm. In one of the regions’ four DCMs (Hvidovre Hospital) we use the results from the multiplex PCR to determine which VREfm clone patients are colonised or infected with. This allows for timely analysis of VREfm dissemination in hospital wards. In addition, when we need to perform cohort isolation in case of lack of single rooms in the wards, we use the results from the multiplex PCR. In those cases, we perform cohort isolation of patients that have the same van gene and Tn1549 insertion site on the multiplex PCR. We use VREfm whole genome sequencing (WGS) for continuous surveillance of VRE on a regional level. This is important to identify the VREfm clone-distribution in our hospitals and emergence of new clones that require diagnostic changes.

The aims of this study were to describe the genomic epidemiology of VREfm in Eastern Denmark, from 2020 to 2022 and to identify emergence of new clones, especially vanB clones with non-araA2/non-sir2 Tn1549 insertion sites. Furthermore, we aimed to identify and characterise the Tn1549 insertion sites among vanB VREfm isolates in our collection to evaluate how many different Tn1549 insertion sites that are present in our vanB VREfm collection and to determine the proportion of vanB VREfm that will be identified by the laboratory developed multiplex-PCR.