Abstract: This experiment was conducted to investigate the effect of radix bupleuri (Bupleurum chinensis) herbal supplementation (RBH) on the diversity of cellulolytic bacteria including Fibrobacter fuccinogene, Ruminococcus albus and Ruminococcus flavefaciens in the rumen of lactating dairy cows. Forty Holstein cows were randomly assigned to 4 diet groups (n=10) according to milk yield (37.5±1.8) kg/d and lactation length (75±15) days. Four treatment diets consisted of 0, 0.25, 0.50 or 1.00 g/kg of RHB dry matter, respectively. The experiment lasted for 10 weeks; rumen fluid samples were collected using a stomach-tube during week 6 of the trial. The diversity of the bacterial community in the rumen was assessed using denaturing gradient gel electrophoresis (DGGE) and real-time quantitative polymerase chain reaction (RT-PCR) analysis. The DGGE analysis indicated that there were no differences among the treatment groups. Compared with the control group, the Shannon-Weiner index was significantly reduced (P<0.05) in the 1.0 g/kg RBH group, while the dominance index was significantly increased (P<0.05) in the 0.50 and 1.00 g/kg treatments. RT-PCR analysis also showed that RBH had no effect on cellulolytic bacteria. It was concluded that RBH supplementation had little effect on diversity of the bacterial community in the rumen of lactating dairy cows, possibly due to gradual adaptation to the RBH supplementation by rumen bacteria.