Eetection of treponema pallidum by polymerase chain reaction.
- Author:
Kee Yang CHUNG
;
Jung Bock LEE
- Publication Type:In Vitro ; Original Article
- MeSH:
Amniotic Fluid;
Cerebrospinal Fluid;
Dermatology;
Diagnosis;
DNA;
Female;
Humans;
Leukocyte Count;
Neurosyphilis;
Paraffin;
Polymerase Chain Reaction*;
Pregnant Women;
Sensitivity and Specificity;
Serologic Tests;
Syphilis;
Syphilis, Congenital;
Syphilis, Latent;
Treponema pallidum*;
Treponema*
- From:Korean Journal of Dermatology
1993;31(4):481-494
- CountryRepublic of Korea
- Language:Korean
-
Abstract:
BACKGROUND: Definite criteria for the diagnosis and treatment evaluation for different clinical stags of syphilis are not yet present dute to the inability to dultivate Treponema pallidum in vitro. However, as the staining methods and the serological tests currently used have their limitations, a more definite method for its confirmation is needed. Polymerase chain reaction (PCR), due to its high sensitivity and specificity, is currently being applied to the detection of T. pallidum. OBJECTIVE: We have used PCR for the detection of T. pallidum DNA in various clinical specimens in orber to evaluate its potenital as a diagnostic tool. METHOD: Clinical specimens were collected from patients with different stages of syphilis who visited ithe Deparment of Dermatology of Yonsei medical Center for 1 year beginning from May, 1991. Sera from 63 patients, cerebrospinal fluids from 24 patients, amniotic fluids from 3 patients, and 21 tissues from 19 patients were subjected to PCR and the results were analyzed to evaluate its usefulness as a diagnostic and treatment evaluation tool. A portion of the T. pallidum-specific chromosomal DNA, tpp47, which encodes the 47 kDa surface protein, was used as the template DNA to amplify the 658 bp DNA fragment, and the following results were obtained. PCR using primers 47-1 and 47-2 were applied to amplify 658 bp DNA fragments from the T. pallidum-specific tpp47 gene encoding 47 kDa surface protein. RESULT: 1. To evaluate the sensitivity of the PCR, T. pallida and their chromosomal DNA were diluted. The diluents contataining a single organism and 1 fg of the chromosomal DNA showed positive reactions by the amplification. 2. Specificity of the PCR was determined by using T. pallidum, 4 species beloging to genus Treponema, and 9 species of nonpathogenic or pathogenic organisms. A positive reaction was obtained only when T. pallidum chromosomal DNA was used. 3. PCR was positive in 5 of 9 (55%) sera in primary syphilis, 22 of 26(84%) in secondary syphilis, 3 to 15(20%) in early latent syphilis, 1 of 19 (11%0 in late latent syphilis, 2 of 2 (100%) in neurosyphlis, and 0 of 2 (0%) in congenital syphilis. The differences in the positive rates were statistically significant (P<0.01) in all stages except neurosyphilis and congenital syphilis, as their numbers were too smalll to deduce any significant meaning. Despite their high VDRL titers, the positive rate in early latent syphilis was relatively low when compared to the rate in secondary syphilis. 4. Follw-up PCR of sera in some patients showed positive results 9 months after treatment. However, some with negative PCR before treatment showed positive results after treatment. 5. PCR was positive in 1 of 1 (50%) cerebrospinal fluid in primary syphilis, 3 of 14 (21%) in secondary syphilis, 2 of 7 (29%) in early latent syphilis, and 1 of 1(100%) in neurosyphlis. The differences in the positive rates showed no statistical significance in relation to the clinical stages. Cerebrospinal fluid VDRL test, white blood cell count, and protein content showed no correlation to the PCR results in early syphilis patients. 6. Amniotic fluid showed a positive PCR result only in a pregnant woman whose serum showed a high VDRL titer and a positive PCR. 7. PCR positive rates were 90% in frozed tissues and 50% in paraffin embedded tissues. CONCLUSION: From the results, it is suggested that PCR is not suitable for treatment evaluation but is useful for the detection of T. pallidum in sera of secondary syphilis patients and syphilitic lesions, and for the confirmation of the diagnosis the diagnosis in these cases.
