Changes in the microbial populations, their activities, and the ruminal fermentation were monitored for 50 d following the reintroduction of ciliate protozoa into four defaunated sheep. A protozoal population was reestablished successfully in each recipient, using a washed inoculum containing approximately 103cells, although there were between-animal differences in the rates of recolonization and genus establishment.Entodiniumspp. predominated in the initial stages of the refaunation period and had an apparent maximal generation time of 9–10 h. Bacterial and fungal numbers did not decline following the reintroduction of protozoa and a small transient increase in the numbers of amylolytic and xylanolytic bacteria and fungal zoospores occurred in the early stages of refaunation when the protozoal population was < 105/g ruminal contents, but these subsequently declined as the protozoa established. Although the fibrolytic bacterial population was lowest in period 3 (> 105protozoa/g), thein saccoruminal digestion ofLolium perennehay and polysaccharolytic enzyme activities in the solids-associated populations were either maintained or increased when protozoa were present confirming the important contribution of protozoa to fibre breakdown in the rumen. Significant changes in ruminal microbial activities occurred after protozoal reinoculation but before the rumen had refaunated completely. Arylamidase activities in the liquor-phase population and ruminal ammonia concentrations increased significantly within 48 h of transfaunation; the magnitude of the effects became more pronounced as the protozoal population developed. However, volatile fatty acid formation and ruminal pH were not affected after the reintroduction of protozoa.Key words: rumen, sheep, ciliate protozoa.