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Journal of Clinical Microbiology logoLink to Journal of Clinical Microbiology
. 1993 Jul;31(7):1753–1758. doi: 10.1128/jcm.31.7.1753-1758.1993

Comparison of plasmid- and chromosome-based polymerase chain reaction assays for detecting Chlamydia trachomatis nucleic acids.

J B Mahony 1, K E Luinstra 1, J W Sellors 1, M A Chernesky 1
PMCID: PMC265626  PMID: 7688753

Abstract

Several laboratories have demonstrated that the polymerase chain reaction (PCR) is more sensitive than culture or enzyme immunoassay (EIA) for detecting Chlamydia trachomatis in genitourinary tract specimens when various DNA targets are used for amplification, including the cryptic plasmid, major outer membrane protein (MOMP), or rRNA genes. We compared the performances of five different PCR assays, including assays with two plasmid, two MOMP, and one rRNA targets, by amplifying serial dilutions of C. trachomatis DNA and testing genitourinary tract specimens. By using published procedures, two different plasmid primers had sensitivities of 0.1 fg for C. trachomatis plasmid DNA and 10 fg for total cellular DNA. The sensitivities of the assays with the two MOMP primers were 0.1 and 10 pg, and the sensitivity for the assay with the rRNA primers was 1 pg for cellular DNA. Both plasmid-based assays detected 38 of 38 confirmed Chlamydiazyme-positive specimens, whereas the assays with the MOMP and rRNA primers detected 36 of 38 and 29 of 38 confirmed Chlamydiazyme-positive specimens, respectively. Six of 18 Chlamydiazyme-negative specimens collected from individuals whose specimens were positive by culture or immunofluorescence were positive by both plasmid-based PCRs; 4 of these were positive by PCR with the MOMP primers and 3 were positive by PCR with the rRNA primers. The results obtained with both purified DNA and genitourinary tract specimens indicated that the plasmid-based PCRs are more sensitive than bacterial chromosome-based PCRs for detecting C. trachomatis.

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Selected References

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  1. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bobo L., Coutlee F., Yolken R. H., Quinn T., Viscidi R. P. Diagnosis of Chlamydia trachomatis cervical infection by detection of amplified DNA with an enzyme immunoassay. J Clin Microbiol. 1990 Sep;28(9):1968–1973. doi: 10.1128/jcm.28.9.1968-1973.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bobo L., Munoz B., Viscidi R., Quinn T., Mkocha H., West S. Diagnosis of Chlamydia trachomatis eye infection in Tanzania by polymerase chain reaction/enzyme immunoassay. Lancet. 1991 Oct 5;338(8771):847–850. doi: 10.1016/0140-6736(91)91502-l. [DOI] [PubMed] [Google Scholar]
  4. Brunham R. C., Kuo C. C., Cles L., Holmes K. K. Correlation of host immune response with quantitative recovery of Chlamydia trachomatis from the human endocervix. Infect Immun. 1983 Mar;39(3):1491–1494. doi: 10.1128/iai.39.3.1491-1494.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chernesky M. A., Mahony J. B., Castriciano S., Mores M., Stewart I. O., Landis S. J., Seidelman W., Sargeant E. J., Leman C. Detection of Chlamydia trachomatis antigens by enzyme immunoassay and immunofluorescence in genital specimens from symptomatic and asymptomatic men and women. J Infect Dis. 1986 Jul;154(1):141–148. doi: 10.1093/infdis/154.1.141. [DOI] [PubMed] [Google Scholar]
  6. Chernesky M., Castriciano S., Sellors J., Stewart I., Cunningham I., Landis S., Seidelman W., Grant L., Devlin C., Mahony J. Detection of Chlamydia trachomatis antigens in urine as an alternative to swabs and cultures. J Infect Dis. 1990 Jan;161(1):124–126. doi: 10.1093/infdis/161.1.124. [DOI] [PubMed] [Google Scholar]
  7. Claas H. C., Melchers W. J., de Bruijn I. H., de Graaf M., van Dijk W. C., Lindeman J., Quint W. G. Detection of Chlamydia trachomatis in clinical specimens by the polymerase chain reaction. Eur J Clin Microbiol Infect Dis. 1990 Dec;9(12):864–868. doi: 10.1007/BF01967500. [DOI] [PubMed] [Google Scholar]
  8. Claas H. C., Wagenvoort J. H., Niesters H. G., Tio T. T., Van Rijsoort-Vos J. H., Quint W. G. Diagnostic value of the polymerase chain reaction for Chlamydia detection as determined in a follow-up study. J Clin Microbiol. 1991 Jan;29(1):42–45. doi: 10.1128/jcm.29.1.42-45.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Erlich H. A., Gelfand D., Sninsky J. J. Recent advances in the polymerase chain reaction. Science. 1991 Jun 21;252(5013):1643–1651. doi: 10.1126/science.2047872. [DOI] [PubMed] [Google Scholar]
  10. Griffais R., Thibon M. Detection of Chlamydia trachomatis by the polymerase chain reaction. Res Microbiol. 1989 Feb;140(2):139–141. doi: 10.1016/0923-2508(89)90047-8. [DOI] [PubMed] [Google Scholar]
  11. Holland S. M., Hudson A. P., Bobo L., Whittum-Hudson J. A., Viscidi R. P., Quinn T. C., Taylor H. R. Demonstration of chlamydial RNA and DNA during a culture-negative state. Infect Immun. 1992 May;60(5):2040–2047. doi: 10.1128/iai.60.5.2040-2047.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Jang D., Sellors J. W., Mahony J. B., Pickard L., Chernesky M. A. Effects of broadening the gold standard on the performance of a chemiluminometric immunoassay to detect Chlamydia trachomatis antigens in centrifuged first void urine and urethral swab samples from men. Sex Transm Dis. 1992 Nov-Dec;19(6):315–319. [PubMed] [Google Scholar]
  13. Kwok S., Higuchi R. Avoiding false positives with PCR. Nature. 1989 May 18;339(6221):237–238. doi: 10.1038/339237a0. [DOI] [PubMed] [Google Scholar]
  14. Loeffelholz M. J., Lewinski C. A., Silver S. R., Purohit A. P., Herman S. A., Buonagurio D. A., Dragon E. A. Detection of Chlamydia trachomatis in endocervical specimens by polymerase chain reaction. J Clin Microbiol. 1992 Nov;30(11):2847–2851. doi: 10.1128/jcm.30.11.2847-2851.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Mahony J. B., Chernesky M. A. Effect of swab type and storage temperature on the isolation of Chlamydia trachomatis from clinical specimens. J Clin Microbiol. 1985 Nov;22(5):865–867. doi: 10.1128/jcm.22.5.865-867.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Mahony J. B., Luinstra K. E., Jang D., Sellors J., Chernesky M. A. Chlamydia trachomatis confirmatory testing of PCR-positive genitourinary specimens using a second set of plasmid primers. Mol Cell Probes. 1992 Oct;6(5):381–388. doi: 10.1016/0890-8508(92)90031-r. [DOI] [PubMed] [Google Scholar]
  17. Mahony J. B., Luinstra K. E., Sellors J. W., Jang D., Chernesky M. A. Confirmatory polymerase chain reaction testing for Chlamydia trachomatis in first-void urine from asymptomatic and symptomatic men. J Clin Microbiol. 1992 Sep;30(9):2241–2245. doi: 10.1128/jcm.30.9.2241-2245.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Mahony J. B., Schachter J., Chernesky M. A. Detection of antichlamydial immunoglobulin G and M antibodies by enzyme-linked immunosorbent assay. J Clin Microbiol. 1983 Aug;18(2):270–275. doi: 10.1128/jcm.18.2.270-275.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Mahony J., Castriciano S., Sellors J., Stewart I., Cunningham I., Landis S., Seidelman W., Grant L., Devlin C., Chernesky M. Diagnosis of Chlamydia trachomatis genital infections by cell culture and two enzyme immunoassays detecting different chlamydial antigens. J Clin Microbiol. 1989 Sep;27(9):1934–1938. doi: 10.1128/jcm.27.9.1934-1938.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Moncada J., Schachter J., Bolan G., Engelman J., Howard L., Mushahwar I., Ridgway G., Mumtaz G., Stamm W., Clark A. Confirmatory assay increases specificity of the chlamydiazyme test for Chlamydia trachomatis infection of the cervix. J Clin Microbiol. 1990 Aug;28(8):1770–1773. doi: 10.1128/jcm.28.8.1770-1773.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Näher H., Drzonek H., Wolf J., von Knebel Doeberitz M., Petzoldt D. Detection of C trachomatis in urogenital specimens by polymerase chain reaction. Genitourin Med. 1991 Jun;67(3):211–214. doi: 10.1136/sti.67.3.211. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Ossewaarde J. M., Rieffe M., Rozenberg-Arska M., Ossenkoppele P. M., Nawrocki R. P., van Loon A. M. Development and clinical evaluation of a polymerase chain reaction test for detection of Chlamydia trachomatis. J Clin Microbiol. 1992 Aug;30(8):2122–2128. doi: 10.1128/jcm.30.8.2122-2128.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Ostergaard L., Traulsen J., Birkelund S., Christiansen G. Evaluation of urogenital Chlamydia trachomatis infections by cell culture and the polymerase chain reaction using a closed system. Eur J Clin Microbiol Infect Dis. 1991 Dec;10(12):1057–1061. doi: 10.1007/BF01984929. [DOI] [PubMed] [Google Scholar]
  24. Palmer H. M., Gilroy C. B., Thomas B. J., Hay P. E., Gilchrist C., Taylor-Robinson D. Detection of Chlamydia trachomatis by the polymerase chain reaction in swabs and urine from men with non-gonococcal urethritis. J Clin Pathol. 1991 Apr;44(4):321–325. doi: 10.1136/jcp.44.4.321. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Ratti G., Moroni A., Cevenini R. Detection of Chlamydia trachomatis DNA in patients with non-gonococcal urethritis using the polymerase chain reaction. J Clin Pathol. 1991 Jul;44(7):564–568. doi: 10.1136/jcp.44.7.564. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Schachter J. Rapid diagnosis of sexually transmitted diseases--speed has a price. Diagn Microbiol Infect Dis. 1986 Mar;4(3):185–189. doi: 10.1016/0732-8893(86)90097-0. [DOI] [PubMed] [Google Scholar]
  27. Sellors J. W., Mahony J. B., Jang D., Pickard L., Goldsmith C. H., Gafni A., Chernesky M. A. Comparison of cervical, urethral, and urine specimens for the detection of Chlamydia trachomatis in women. J Infect Dis. 1991 Jul;164(1):205–208. doi: 10.1093/infdis/164.1.205. [DOI] [PubMed] [Google Scholar]
  28. Sellors J., Mahony J., Jang D., Pickard L., Castriciano S., Landis S., Stewart I., Seidelman W., Cunningham I., Chernesky M. Rapid, on-site diagnosis of chlamydial urethritis in men by detection of antigens in urethral swabs and urine. J Clin Microbiol. 1991 Feb;29(2):407–409. doi: 10.1128/jcm.29.2.407-409.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Stephens R. S., Mullenbach G., Sanchez-Pescador R., Agabian N. Sequence analysis of the major outer membrane protein gene from Chlamydia trachomatis serovar L2. J Bacteriol. 1986 Dec;168(3):1277–1282. doi: 10.1128/jb.168.3.1277-1282.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Tam J. E., Davis C. H., Thresher R. J., Wyrick P. B. Location of the origin of replication for the 7.5-kb Chlamydia trachomatis plasmid. Plasmid. 1992 May;27(3):231–236. doi: 10.1016/0147-619x(92)90025-6. [DOI] [PubMed] [Google Scholar]
  31. Taylor-Robinson D., Gilroy C. B., Thomas B. J., Keat A. C. Detection of Chlamydia trachomatis DNA in joints of reactive arthritis patients by polymerase chain reaction. Lancet. 1992 Jul 11;340(8811):81–82. doi: 10.1016/0140-6736(92)90399-n. [DOI] [PubMed] [Google Scholar]
  32. Watson M. W., Lambden P. R., Clarke I. N. Genetic diversity and identification of human infection by amplification of the chlamydial 60-kilodalton cysteine-rich outer membrane protein gene. J Clin Microbiol. 1991 Jun;29(6):1188–1193. doi: 10.1128/jcm.29.6.1188-1193.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. White T. J., Madej R., Persing D. H. The polymerase chain reaction: clinical applications. Adv Clin Chem. 1992;29:161–196. doi: 10.1016/s0065-2423(08)60224-3. [DOI] [PubMed] [Google Scholar]
  34. Williams T. W., Tyler S. D., Giercke S., Pollard D. R., McNicol P., Rozee K. R. Comparison of polymerase chain reaction and chlamydiazyme for the detection of Chlamydia trachomatis in clinical specimens. Eur J Clin Microbiol Infect Dis. 1992 Mar;11(3):233–236. doi: 10.1007/BF02098085. [DOI] [PubMed] [Google Scholar]
  35. Wu C. H., Lee M. F., Yin S. C., Yang D. M., Cheng S. F. Comparison of polymerase chain reaction, monoclonal antibody based enzyme immunoassay, and cell culture for detection of Chlamydia trachomatis in genital specimens. Sex Transm Dis. 1992 Jul-Aug;19(4):193–197. doi: 10.1097/00007435-199207000-00002. [DOI] [PubMed] [Google Scholar]

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