Advanced Techniques in Diagnostic Microbiology

22. Molecular Techniques for STDs Diagnosis 361
also been commonly used (Romanowski et al., 1987). Both dark-field microscopy
and DFA-TP, however, do not distinguish T. pallidum from the other pathogenic
species of Treponema (Larsen et al., 1995). On the other hand, nontreponemal serologic
tests are based on detection of antibodies to a cardiolipid–cholesterol–lecithin
antigen. They include the Venereal Diseases Research Laboratory (VDRL) and the
Rapid Plasma Reagin (RPR) card test. Both are modified from the original Wasserman
reaction (Larsen et al., 1989; Young, 1992). Limitations of these tests include
the lack of sensitivity in early and latent stages of syphilis and the well-known
possibility of biological false-positive reactions. Treponemal serologic tests are
based on detection of treponemal specific antibodies to cellular component of T.
pallidum. They include the serum fluorescent treponemal antibody absorption test
(FTA-ABS), the microhemagglutination test (MHA-TP), and the Treponema pallidum
particle agglutination (TPPA) test (Rodolph 1976). These tests have higher
sensitivities and specificities than nontreponemal serologic tests and are used as
confirmatory tests for syphilis. Although false-positive results are rare in FTA-ABS
compared with nontreponemal serologic tests, they have been well documented
to occur in association with autoimmune diseases, viral infections, and pregnancy
(Sparling, 1971). Enzyme immunoassay (EIA) is a semiautomated method used to
detect treponemal antibodies (Young et al., 1998). A format that can directly detect
T. pallidum antigens in early syphilitic lesions has also been developed (Young
et al., 1998). Antigens are extracted from swabs containing lesion exudates within
72 h and the antigen–antibody complexes are measured in an automated spectrophotometer
device. Advantages of the EIA test are more objectivity in interpretation;
less labor intensive; and potentially automatable.
Molecular Detection Methods
Polymerase chain reaction (PCR) is the newest laboratory technique for direct
detection of T. pallidum. Only a few laboratories use PCR for routine case detection.
The CDC has developed a PCR test that is available for specimens from patients
with suspected neurosyphilis or with genital ulcer diseases (CDC, 2002). PCR has
also been found to be useful for detection of congenital syphilis, and the sensitivity
of the test is comparable to RIT for amniotic fluid specimens (Hollier, 2001). In
addition, a syphilitic aortitis case (which often has atypical clinical presentation,
and available serological tests are nonspecific) has recently been described that
was diagnosed by the use of PCR (O’Regan et al., 2002). Dot blot hybridization
is another technique also being used for detection of T. pallidum (Jethwa et al.,
1995).
PCR
Several PCR-based tests of T. pallidum have been developed on the basis of membrane
lipoproteins (Orle et al., 1996), TmpA and 4D genes (Hay et al., 1990), 16S
RNA (Centurion-Lara et al., 1997), DNA polymerase I (polA) gene (Liu et al.,
2001). Levels of detection of these assays ranged between 10 −3 equivalents of