Isolating patients infected or colonized with vancomycin-resistant enterococci (VRE) in a private room or cohort room to prevent hospital transmission is controversial. To evaluate the effect of a relaxed isolation policy for VRE-infected or colonized patients on healthcare-associated (HA) VRE bacteraemia in an acute care hospital with a predominantly shared-room setting. The incidence of HA VRE bacteraemia was compared during a private isolation era (October 2014–September 2017), a cohort isolation era (October 2017–June 2020), and a no isolation era (July 2020–June 2022). Using Poisson regression modelling, an interrupted time-series analysis was conducted to analyse level changes and trends in incidences of HA VRE bacteraemia for each era. The proportion of VRE-infected or -colonized patients staying in shared rooms increased from 18.3% in the private isolation era to 82.6% in the no isolation era (P < 0.001). There was no significant difference in the incidences of HA VRE bacteraemia between the private isolation era and the cohort isolation era (relative risk: 1.01; 95% confidence interval: 0.52–1.98; P = 0.977) or between the cohort isolation era and the no isolation era (0.99; 0.77–1.26; P = 0.903). In addition, there was no significant slope increase in the incidence of HA VRE bacteraemia between any of the eras. In a hospital with predominantly shared rooms, the relaxation of isolation policy did not result in increased HA VRE bacteraemia, when other infection control measures were maintained.