To date, no genes have been characterized from mitochondrial DNA of hypotrichous ciliates. Here we present the complete sequence of the gene (cyb) encoding cytochrome b in the hypotrichous ciliate Pseudourostyla cristata. Analysis of nucleotide and deduced amino acids sequences predicts a protein of 393 amino acids encoded by cyb. CYB amino acid similarity and phylogenetic tree analyses strongly support a close evolutionary relationship between Paramecium, Tetrahymena and Pseudourostyla. The heat stress-dependent expression pattern of CYB was verified by semi-quantitative real-time reverse transcriptase polymerase chain reaction (RT-PCR) analysis.