The ecology of the uncultured, but large and morphologically conspicuous, rumen bacterium Oscillospira spp. was studied. Oscillospira-specific 16S rRNA gene sequences were detected in North American domestic cattle, sheep from Australia and Japan, and Norwegian reindeer. Phylogenetic analysis of the sequences obtained allowed definition of three operational taxonomic units within the Oscillospira clade. Consistent with this genetic diversity, we observed atypical smaller morphotypes by using an Oscillospira-specific fluorescence in situ hybridization probe. Despite the visual disappearance of typical large Oscillospira morphotypes, the presence of Oscillospira spp. was still detected by Oscillospira-specific PCR in the rumen of cattle and sheep. These observations suggest the broad presence of Oscillospira species in various rumen ecosystems with the level, and most likely the morphological form, dependent on diet. An ecological analysis based on enumeration of the morphologically conspicuous, large-septate form confirms that the highest counts are associated with the feeding of fresh forage diets to cattle and sheep and in two different subspecies of reindeer investigated.Morphologically unique, large bacteria of the genus Oscillospira, which have not been grown in pure culture yet, can be routinely detected microscopically in the rumen contents of cattle and sheep (11,18). A number of Oscillospira spp. are responsive to the advent of green pastures and fluctuate seasonally (13, 17) (see also Table 1). The only species of Oscillospira described in Bergey's Manual of Systematic Bacteriology (8) is Oscillospira guilliermondii, and this entry refers to the original morphological description by Chatton and Perard in 1913. Warner (34) proposed that at least two strains or species be included in the Oscillospira group based on cell diameter and the tendency to form spores as the main characteristic differences. This implies that there are different morphological forms or species related to different diets or in different gut ecosystems.Detection and identification of microbial populations are the most basic prerequisites for microbial ecology studies. Over the last decade, several molecular techniques have been developed that when applied to the rumen microbiota have revealed enormous microbial diversity not recognized previously because of limitations and biases inherent in the cultivation approach (24,(30)(31)(32)37). The cultivation-independent approach provides technology for detection and monitoring of microorganisms such as Oscillospira, for which growth requirements are unknown and undetermined. The only requirement for the development of molecular detection techniques is the availability of a marker molecule. Fortunately, several 16S rRNA gene sequences have recently been retrieved from this large bacterium isolated by flow cytometric sorting (39), and this enabled the design of various molecular probes for detection and monitoring. We designed and validate here PCR and PCR-denaturing gradient gel electroph...