Three growing Polish Merino rams were defaunated and then consecutively refaunated with Eudiplodinium maggii, Entodinium caudatum and Dasytricha ruminantium. Establishment of Eudiplodinium maggii in the rumen of defaunated animals resulted in the increase of β-endoglucanase and xylanase activities by about 42 and 46%, respectively (P < 0.01). Enzyme activities did not change following the development of Entodinium caudatum whereas significantly decreased (P < 0.01) when Dasytricha ruminanium was added to Entodinium caudatum and Eudiplodinium maggii. Rumen digesta weight tended to increase following the consecutive refaunations. ADF and NDF in rumen digesta varied in the range of 4.1–5.6 and 6.5–7.9%, respectively, and increased significantly after establishment of Entodinium caudatum and Dasytricha ruminantium. Disappearance rate of ADF and NDF was affected by the microfauna composition and varied from 0.21 to 0.30 and from 0.36 to 0.50 kg/12 h, respectively. Establishment of Entodinium caudatum and Dasytricha ruminantium in the rumen was accompanied by decrease in disappearance rate of ADF and NDF (P < 0.01). The negative correlation was found between both the ADF and NDF contents in rumen digesta and their disappearance rate from the rumen.