Author
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Everett, Karin |
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Andersen, Arthur |
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Submitted to: American Society for Microbiology
Publication Type: Abstract Only Publication Acceptance Date: 3/1/1996 Publication Date: N/A Citation: N/A Interpretive Summary: Technical Abstract: There is evidence that chlamydia is a widespread infectious agent in livestock; however, diagnosis of infection and subsequent identification of strains suffer from the wide diversity of isolates and in vitro growth requirements. The objective of this study was to develop a simple PCR- based diagnostic test that detects the presence of chlamydia and identifies sthe species of the infecting strain. To create a set of data on which a universal, chlamydia-specific diagnostic test could be based, DNA was prepared from over thirty isolates of chlamydia and a 3 kb region of the ribosomal operon was PCR amplified, cloned, and partially sequenced. The sequenced portion of the 16S rRNA gene was highly conserved (>95%) and enabled us to design a genus-specific oligonucleotide primer. The sequenced portion of the 23S gene was also conserved (85-95% over 900 bases), permitting the selection of a second PCR primer recognizing all the echlamydial isolates. Amplification of a short DNA segment using these primers was specific for chlamydia, and nested primers were identified for both species and subspecies typing. In addition, analysis of the 23S data was used to construct a rooted genealogical tree for chlamydia, and the intergenic spacer was used to construct an unrooted tree. Both these trees were consistent with existing genealogies based on the amino acid sequence of the major outer membrane protein of chlamydia, and with existing biological and DNA hybridization data. The results indicate that PCR amplification of the 16S/23S domain and the ribosomal intergenic spacer can be used for both diagnosis and classification of chlamydial strains. |
