Summary: Respiratory tract infections in man may be caused by several members of the genus and also by two -like strains, (Z-agent) and (Bn). To facilitate diagnostic procedures a PCR assay able to detect all known sequences in one reaction was developed. For this purpose, primers were selected to amplify a fragment of the 16S rRNA gene. Characterization of the amplified fragments was done by hybridization with specific probes and by sequencing. PCR assays were carried out using DNA isolated from nose/throat specimens or from peripheral blood mononuclear cells of patients with respiratory tract infections, and from vessel wall specimens of abdominal aneurysms. Six of the 42 nose/throat swab specimens analysed yielded strong bands and one yielded a faint band. Three of these bands were identified as and one as by sequencing. Analysis of the three other bands yielded two different new sequences. DNA isolated from peripheral blood mononuclear cells of one patient yielded a third new sequence. DNA isolated from peripheral blood mononuclear cells of four healthy controls was negative. One of the abdominal aneurysm specimens also yielded a strong band. Sequencing revealed a fourth new sequence. All negative controls included during specimen processing and PCR analysis remained negative. The typical secondary structure of microbial 16S genes was present in all four new sequences indicating the validity of the sequence data. All four new sequences were distinct from other bacteria and clustered together with known sequences. Phylogenetic analysis suggested a new lineage, separating the four new sequences, and from the genus with the four known chlamydial species. In conclusion, this study provides evidence for the existence of several new members of the order Since the source of the -like strains has not been identified and serological and/or molecular cross-reactivities may be expected, results of identification of infecting recognized organisms should be interpreted cautiously.


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