BACKGROUND: The vanA gene is the most frequently encountered gene among isolates, causing vancomycin-resistant Enterococci (VRE) infections in humans, and it is part of the transposable element Tn1546. Knowledge of the diversity of Tn1546 is important to distinguish between the dissemination of a single VRE clone and the transmission of a particular Tn1546 type through a genetically divergent population of enterococci. Recently, we studied molecular diversity of Tn1546-related elements in enterococci isolated in one hospital to facilitate understanding of the molecular epidemiology of vancomycin resistance. METHODS: Nineteen VanA-type VRE clinical isolates, collected in one university hospital during 1997 and 1999, were investigated for characteristics such as antibiotic resistance, structure of vanA gene cluster and genomic DNA type by means of antibiotic susceptibility test, PCR amplified length polymorphism of vanA gene cluster and pulse-field gel electrophoresis (PFGE), respectively. RESULTS: Nine (A, B1 to B5, C, D and E) different vanA gene cluster types were identified. Three isolates were grouped into vanA gene cluster type A, similar to that of Tn1546 prototype, and twelve isolates were grouped into type B that has an insertion of IS1216V at vanX-Y intergenic region. Type B was further subdivided into B1 to B5 according to the size variation of vanX-Y intergenic region, which was resulted from the insertion of IS1216V and deletions associated with the insertion. Both vanY and vanZ were deleted in three isolates, suggesting that these genes are not essential for vancomycin resistance. Notably, three E. faecalis and three E. faecium strains isolated during the same period were shown to carry the same vanA gene cluster, showing intergenic transmission of vancomycin resistance. CONCLUSION: The presence of variable types of vanA gene cluster among VRE strains isolated in one hospital suggests that several evolutionary changes of vanA gene clusters have occurred during the horizontal spread of resistance gene in the hospital environment. This approach may be useful for monitoring the evolution of VanA resistance.