We have compared nine Enterococcus faecalis strains with E. faecalis V583 by comparative genomic hybridization using microarrays (CGH). The strains used in this study (the "test" strains) originated from various environments. CGH is a powerful and promising tool for obtaining novel information on genome diversity in bacteria. By CGH, one obtains clues about which genes are present or divergent in the strains, compared to a reference strain (here, V583). The information obtained by CGH is important from both ecological and systematic points of view. CGH of E. faecalis showed considerable diversity in gene content: Compared to V583, the percentage of divergent genes in the test strains varied from 15% to 23%, and 154 genes were divergent in all strains. The main variation was found in regions corresponding to exogenously acquired or mobile DNA in V583. Antibiotic resistance genes, virulence factors, and integrated plasmid genes dominated among the divergent genes. The strains examined showed various contents of genes corresponding to the pTEF1, pTEF2, and pTEF3 genes in V583. The extensive transport and metabolic capabilities of V583 appeared similar in the test strains; CGH indicated that the ability to transport and metabolize various carbohydrates was similar in the test strains (verified by API 50 CH assays). The contents of genes related to stress tolerance appeared similar in V583 and the nine test strains, supporting the view of E. faecalis as an organism able to resist harsh conditions.