Identification of Chlamydia pneumoniae by DNA amplification of the 16S rRNA gene

Abstract

Chlamydia pneumoniae is an important cause of respiratory disease in humans, but diagnosis of C. pneumoniae is hindered by difficulties in the in vitro growth of the organism. In order to improve detection and identification, we recently developed a polymerase chain reaction (PCR) assay which uses oligonucleotide primers specific for C. pneumoniae. The nucleic acid sequence was determined for the 16S rRNA of C. pneumoniae, and regions in which C. pneumoniae differed from both Chlamydia psittaci and Chlamydia trachomatis were identified. Oligonucleotide primers corresponding to these unique regions were then synthesized and used in a PCR for the detection of C. pneumoniae. The C. pneumoniae-specific primers permitted the identification of six isolates of C. pneunwniae, but no reaction was observed with the 15 serovars of C. trachomatis or two strains of C. psittaci. PCR should prove to be valuable in confirming the identification of C. pneumoniae and in the diagnosis of C. pneumoniae infections.

Development of a simplified polymerase chain reactionenzyme immunoassay for the detection of Chlamydia pneumoniae

“…A comparison of alignments of these and other chlamydia] 16s rRNA gene sequences extracted from the GenBankT" database (accession numbers M59178 and M13769) revealed uniformity between strains within a species but significant variation between strains of different species (100% between the two C. pneumoniae strains, 100% between the two C. psittaci strains, 95.9% between C. pneumoniae and C. psittaci strains and 93.6% similarity between C. pneumoniae and C. trachomatis strains). As in previous studies (Gaydos et al 1992) it was noted that several locations within the sequences exhibited considerable diversity. In particular the divergence observed between regions (using E. coli 16s rRNA gene sequence numbering) 158-290 and 432-452 was significant enough to allow production of nested PCR primers specific to C. pneumoniae.…”

“…pneumoniae have been described. These procedures were based on the detection of the 16s rRNA (Gaydos et al 1992(Gaydos et al , 1993(Gaydos et al , 1994, the major outer membrane protein (MOMP) (Holland et al 1990) or other chromosomal sequences (Campbell et al 1992). The aim of this study was to develop an improved PCR-EIA for C. pneumoniae detection, based on the method of Gaydos et al (1993) which combines the specificity and sensitivity of PCR with the ease and convenience of colorimetric detection in ELISA plates.…”

What Is the Role of Newer Molecular Tests in the Management of CAP?

“…72,73 The CDC workshop identified a few assays that were considered to be "validated" enough to be used for research studies; others have been developed and used. 66,[74][75][76][77][78][79][80][81][82] The advantages of these assays are their sensitivity, decreased possibility of contamination, and ability to quantify DNA. A PCR assay has recently been used to identify an outbreak of CAP among navy SEALs.…”

Serology of Chlamydia pneumoniae in patients with chronic cough

“…PCR was performed using a Premix kit (Bioneer, Seoul, Korea), and the final mixture contained 10 pmol of each C. P. DNA primer (CPN 90: 5′-GGT CTC AAC CCC ATC CGT GTC GG-3′ and CPN 91: 5′-TGC GGA AAG CTG TAT TTC TAC AGT T-3′). 11 For each reaction, the total volume was 20 µL. The amplification was performed in an iCycler DNA thermal cycler (Bio-Rad, Hercules, CA, USA) with an initial 75 s at 95°C, followed by 60 cycles of 94°C for 45 s, annealing for 45 s and 72°C for 1 min.…”