The nucleotide sequences of 16S ribosomal DNA (rDNA) were determined for 39 strains of Chlamydia psittaci (34 from birds and 5 from mammals) and for 4 Chlamydia pecorum strains. The sequences were compared phylogenetically with the gene sequences of nine Chlamydia strains (covering four species of the genus) retrieved from nucleotide databases. In the neighbor-joining tree, C. psittaci strains were more closely related to each other than to the other Chlamydia species, although a feline pneumonitis strain was distinct (98.3 to 98.6% similarity to other strains) and appeared to form the deepest subline within the species of C. psittaci (bootstrap value, 99%). The other strains of C. psittaci exhibiting similarity values of more than 99% were branched into several subgroups. Two pigeon strains and one turkey strain formed a distinct clade recovered in 97% of the bootstrapped trees. The other pigeon strains seemed to be distinct from the strains from psittacine birds, with 88% of bootstrap value. In the cluster of psittacine strains, three parakeet strains and an ovine abortion strain exhibited a specific association (level of sequence similarity, 99.9% or more; bootstrap value, 95%). These suggest that at least four groups of strains exist within the species C. psittaci. The 16S rDNA sequence is a valuable phylogenetic marker for the taxonomy of chlamydiae, and its analysis is a reliable tool for identification of the organisms.Chlamydiae constitute a group of obligate intracellular organisms that are differentiated from other bacteria by their unique developmental cycles (43). The genus Chlamydia currently contains four species, Chlamydia trachomatis, Chlamydia pneumoniae, Chlamydia pecorum, and Chlamydia psittaci (16,17,22). C. trachomatis has been divided into three biotypes and 15 serovars (44). C. pneumoniae, which has been separated from C. psittaci based on DNA-DNA reassociation and the morphology of the elementary body (EB), is clearly discriminated by antigenic structures that react with monoclonal antibodies (MAbs) (32,38). The genealogical characteristics of another newly classified species, C. pecorum, are also distinct (7,16,27). Strains of C. psittaci, which has a broader host range than other species, have been grouped by MAbs (5,6,19,28,39) and by genetic diversity (5,15,18,20,27,41). We have reported that C. psittaci strains from pigeons and budgerigars can be grouped into six immunotypes by using MAbs to a pigeon strain and a budgerigar strain (40). Furthermore, Fukushi et al. (19) and Kikuta et al. (28) have demonstrated that extensive antigenic diversity exists not only between avian and mammalian C. psittaci strains but also within each of the groups. A comprehensive survey has been published by Anderson (6), who discriminated four avian serovars and three or more mammalian types by using serovar-specific MAbs.The importance of genetic relationships among chlamydial species and strains has been established, and typing of the organisms has been performed by many investigators. DNA-DNA homology...