Identification of Enterococcus species and phenotypically similar Lactococcus and Vagococcus species by reverse checkerboard hybridization to chaperonin 60 gene sequences

Data from four recent studies (S. H. Goh et al., J. Clin. Microbiol. 36:216 4-2166, 1998; S. H. Goh et al., J. Clin. Microbiol. 34:818-823, 1996; S. H. Goh et al., J. Clin. Microbiol. 35:3116-3121, 1997; A. Y. C. Kwok et al., Int. J. Syst. Bacteriol. 49:1181-1192, 1999) suggest that an approximately 600-bp region of the chaperonin 60 (Cpn60) gene, amplified by PCR with a si ngle pair of degenerate primers, has utility as a potentially universal tar get for bacterial identification IID). This Cpn60 gene ID method correctly identified isolates representative of numerous staphylococcal species and S treptococcus iniae, a human and animal pathogen. We report herein that this method enabled us to distinguish clearly between 17 Enterococcus species ( Enterococcus asini, Enterococcus rattus, Enterococcus dispar, Enterococcus gallinarum, Enterococcus hirae, Enterococcus durans, Enterococcus cecorum, Enterococcus faecalis, Enterococcus mundtii, Enterococcus casseliflavus, En terococcus faecium, Enterococcus malodoratus, Enterococcus raffinosus, Ente rococcus avium, Enterococcus pseudoavium, Enterococcus new sp. strain Fackl am, and Enterococcus saccharolyticus), and Vagococcus fluvialis, Lactococcu s lactis, and Lactococcus garvieae. From 123 blind-tested samples, only two discrepancies were observed between the Facklam and Collins phenotyping me thod (R. R. Facklam and M. D. Collins, J. Clin. Microbiol. 27:731-734, 1989 ) and the Cpn60 ID method. In each case, the discrepancies were resolved in favor of the Cpn60 ID method. The species distributions of the 123 blind-t ested isolates were Enterococcus new sp. strain Facklam (ATCC 700913), 3; E . asini, 1; E. rattus, 4; E. dispar, 2; E. gallinarum, 20; E. hirae, 9; E. durans, 9; E. faecalis, 12; E. mundtii, 3; E. casseliflavus, 8; E. faecium, 25; E. malodoratus, 3; E. raffinosus, 8; E. avium, 4; E, pseudoavium, 1; a n unknown Enterococcus clinical isolate, sp. strain R871; Vagococcus fluvia lis, 4; Lactococcus garvieae, 3; Lactococcus lactis, 3; Leuconostoc sp., 1; and Pediococcus sp., I. The Cpn60 gene ID method, coupled with reverse che ckerboard hybridization, is an effective method for the identification of E nterococcus and related organisms.