There is considerable debate over the contribution of virginiamycin use in animals to quinupristin/dalfopristin (Q/D) resistance in humans. In this study, the prevalence and mechanisms of streptogramin resistance in enterococci from animals and the environment were investigated. From 2000-2004, enterococci from samples were tested for antimicrobial susceptibility. Q/D-resistant isolates (minimum inhibitory concentration >or=4 microg/mL) were subjected to polymerase chain reaction (PCR) using primers for streptogramin resistance genes (ermB, msrC, vatD and vatE). From the analysis, 1029/6227 (17%) Q/D-resistant non-Enterococcus faecalis enterococci were identified. The majority of Q/D-resistant isolates were Enterococcus hirae (n=349; 34%), Enterococcus casseliflavus (n=271; 26%) and Enterococcus faecium (n=259; 25%). Using PCR, 55.5% (n=571) were positive for ermB, 3% (n=34) for msrC, 2% (n=20) for vatE and 0.3% (n=3) for vatD; 39% (n=401) were negative for all four genes. The vatD-positive samples comprised two E. faecium from chicken and one E. hirae from swine. The nucleotide sequence of vatD from the three isolates was 100% homologous to published vatD sequences. These data indicate that Q/D resistance among enterococci from animals remains low despite the long history of virginiamycin use. To date, this is the first report of vatD from enterococci in animals in the USA.