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Background: <i>Enterococcus</i> is an important cause of infection in the hospital as well as in the community. Methods: A prospective study was done in Medical College, Kolkata for a period of 2 years (from January 2018 to December 2019). After obtaining clearance from the Institutional Ethics Committee, <i>Enterococcus</i> isolates from cases of vaginitis were included in the study. Identification of <i>Enterococcus</i> species was done by Gram stain and conventional biochemical tests along with automated identification by VITEK 2 Compact. These isolates were tested for antimicrobial susceptibility to different antibiotics by Kirby Bauer disc diffusion method and minimum inhibitory concentration (MIC) by VITEK 2 Compact. Interpretation of susceptibility was done according to the Clinical and Laboratory Standards Institute (CLSI) 2017 guidelines. Biofilm detection for <i>Enterococcus</i> species was done. Results: During the period of 2 years, 39 isolates of <i>Enterococcus</i> spp. were obtained from vaginitis cases. Among these, 27 were <i>Enterococcus faecalis</i> and 12 <i>Enterococcus faecium</i>. All isolates were highly susceptible to vancomycin, teicoplanin, and linezolid. Biofilm was detected in eight isolates of which five were strong biofilm producer and three moderate biofilm producers. Conclusion: Biofilm production is an important virulence factor in <i>Enterococcus</i> isolates from vaginitis.