The polymerase chain reaction (PCR) was used to amplify small-subunit ribosomal DNA from the anaerobic ciliated protozoon , and from its uncultured endosymbiotic bacteria. This was accomplished directly from total DNA extracted from protozoa without prior isolation or enrichment for symbiont cells. The double-stranded amplification products were precipitated and directly sequenced using the linear PCR reaction. Fluorescent oligonucleotide probes were designed and used in whole-cell hybridizations to provide direct visual evidence that the sequences originated from the host ciliate and from the endosymbiont. Phylogenetic analysis of the sequence consistently placed it as a deep-branching lineage near the root of the ciliate tree. However, the present data were insufficient to resolve the detailed relationship between and and thus to determine if the heterotrichs are mono- or paraphyletic. Phylogenetic analysis of the symbiont partial sequence clearly demonstrated that it is an archaeobacterium and that it is closely related to, but distinct from, .


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