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Journal of Clinical Microbiology, May 2001, p. 1941-1946, Vol. 39, No. 5
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.5.1941-1946.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

New Tests for Syphilis: Rational Design of a PCR Method for Detection of Treponema pallidum in Clinical Specimens Using Unique Regions of the DNA Polymerase I Gene

Hsi Liu,* Berta Rodes, C.-Y. Chen, and Bret Steiner

Centers for Disease Control and Prevention, Atlanta, Georgia 30333

Received 14 December 2000/Returned for modification 18 January 2001/Accepted 21 February 2001

A sensitive and specific PCR method to detect Treponema pallidum in clinical specimens was developed. PCR primers were designed based on two unique features of the DNA polymerase I gene (polA). The first distinctive characteristic is that the region codes for a high cysteine content and has low homology with similar regions of DNA polymerase I gene from known microorganisms. The second unique feature is the presence of four insertions in the gene. PCR tests using primers designed on the basis these regions reacted with various pathogenic T. pallidum subspecies but did not react with nonpathogenic treponemal species or other spirochetes. An additional 59 species of bacteria and viruses, including those that cause genital ulcers, tested negative. This PCR method is extremely robust and sensitive. The detection limit is about 10 to 25 organisms when analyzed on gel. However, the analytic sensitivity can be increased by at least 1 log, to a detection limit of a single organism, when the ABI 310 Prism Genetic Analyzer is used to detect fluorescence-labeled amplicons. We further used this test in a clinical setting and compared the results with results from a previously reported multiplex-PCR test (for T. pallidum, Haemophilus ducreyi, and herpes simplex virus). We tested 112 genital ulcer specimens by the polA PCR, obtaining a sensitivity of 95.8% and a specificity of 95.7%. These results suggest that the polA PCR is applicable as a routine clinical diagnostic test for syphilis.

* Corresponding author. Mailing address: Centers for Disease Control and Prevention, 1600 Clifton Rd., Mail Stop D13, Atlanta, GA 30333. Phone: (404) 639-3348. Fax: (404) 639-3976. E-mail: hcl6{at}cdc.gov.

Journal of Clinical Microbiology, May 2001, p. 1941-1946, Vol. 39, No. 5
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.5.1941-1946.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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