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Journal of Clinical Microbiology, August 2007, p. 2649-2653, Vol. 45, No. 8
0095-1137/07/$08.00+0     doi:10.1128/JCM.00451-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Development of Real-Time PCR Assays for Genotyping of Chlamydia trachomatis{triangledown}

Hamid Jalal,1* Hannah Stephen,1 Sarah Alexander,2 Christopher Carne,3 and Christopher Sonnex3

Clinical Microbiology & Public Health Laboratory, Box 236, Addenbrooke's Hospital, Hills Road, Cambridge CB2 2QW, United Kingdom,1 Sexually Transmitted Bacteria Reference Laboratory, Centre for Infection, 61 Colindale Avenue, London NW9 5HT, United Kingdom,2 Department of Genitourinary Medicine, Clinic 1A, Box 38, Addenbrooke's Hospital, Hills Road, Cambridge CB2 2QQ, United Kingdom3

Received 27 February 2007/ Returned for modification 27 March 2007/ Accepted 30 May 2007

We have developed and validated a nested real-time PCR (NRT-PCR) for the genotyping of Chlamydia trachomatis and used it specifically for the typing of either eight genovars from D to K or three genovars of lymphogranuloma venereum (LGV). The 11 probes used in the NRT-PCR correctly identified the DNA from D to K and LGV reference strains and did not cross-react with the DNA from 26 strains representing the bacterial pathogens and commensals of the oropharynx, genital tract, and rectum. The NRT-PCR had a 95% probability of detection at four genome copies (confidence interval, three to six copies) of C. trachomatis per reaction. One hundred cervical and urethral swab specimens containing C. trachomatis DNA from 63 women and 37 men were used to validate the method. The results from the NRT-PCR and the DNA sequencing of amplicons generated from the omp1 gene showed 100% correlation for these samples. The assay also identified the LGV-II genotype in 24 of 48 rectal swab specimens containing C. trachomatis DNA that were obtained from men having sex with men. The Sexually Transmitted Bacteria Reference Laboratory, London, independently confirmed these results using group-specific LGV real-time PCR and restriction fragment length polymorphism analysis. Compared with the NRT-PCR, non-NRT-PCR was found to be less sensitive: it typed C. trachomatis DNA in only 80% of the genital samples and 90% of the rectal swab samples. This is the first successful demonstration of the use of real-time PCR for the genotype-specific typing of C. trachomatis strains that cause sexually transmitted diseases.

* Corresponding author. Mailing address: Clinical Microbiology & Public Health Laboratory, Box 236, Addenbrooke's Hospital, Hills Road, Cambridge CB2 2QW, United Kingdom. Phone: 44-1223-257036. Fax: 44-1223-242775. E-mail: hamid.jalal{at}addenbrookes.nhs.uk

{triangledown} Published ahead of print on 13 June 2007.

Journal of Clinical Microbiology, August 2007, p. 2649-2653, Vol. 45, No. 8
0095-1137/07/$08.00+0     doi:10.1128/JCM.00451-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Morre, S. A, Ouburg, S., van Agtmael, M. A, de Vries, H. J C (2008). Lymphogranuloma venereum diagnostics: from culture to real-time quadriplex polymerase chain reaction. Sex. Transm. Infect. 84: 252-253 [Full Text]  
  • Huang, C.-T., Wong, W.-W., Li, L.-H., Chiang, C.-C., Chen, B.-D., Li, S.-Y. (2008). Genotyping of Chlamydia trachomatis by Microsphere Suspension Array. J. Clin. Microbiol. 46: 1126-1128 [Abstract] [Full Text]  


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