Acta Dermatovenerol Croat

2012;20(2):84-88 CLINICAL ARTICLE

Serologic Diagnosis of Syphilis: Comparison

of Different Diagnostic Methods
Yunus Saral1, Aziz Ramazan Dilek2, Nursel Dilek1, İlkay Bahçeci2,
Deniz Zehra Ulusan2
1
Department of Dermatology; 2Department of Microbiology, Rize Education and
Research Hospital, Rize, Turkey

Corresponding author: SUMMARY Diagnostic and therapeutic approaches in syphilis show wide
Aziz Ramazan Dilek, MD variation. The use of only one type of serologic test is insufficient for diag-
nosis. However, current international recommendations cannot be applied
Department of Microbiology due to various reasons (cost, availability, etc.). The aim of the study was
Rize Education and Research Hospital to review serologic data of syphilis patients to determine diagnostic per-
Rize formance of three different methods. In 117 patients suspected of having
syphilis, syphilis was diagnosed serologically and clinically. Three different
Turkey
methods were used for detection of antibodies: Rapid Plasma Reagin (RPR),
ar.dilek@hotmail.com Treponemal Chemiluminescence Microparticle Enzyme Immunoassay
(CMIA) and Treponema pallidum hemagglutination (TPHA). The sensitivity,
Received: June 10, 2011 specificity, positive predictive value, negative predictive value and accuracy
were calculated for the former two methods against TPHA. The sensitivity of
Accepted: May 15, 2012 RPR and CMIA against TPHA was 58% and 98%, respectively. The specificity
of RPR and CMIA against TPHA was 0% and 100%, respectively. Automated
enzyme immunoassay systems could contribute to reducing errors that de-
pend on the person, especially while monitoring titration changes.

KEY WORDS: syphilis, Treponema pallidum, enzyme immunoassays, TPHA,

RPR

INTRODUCTION
Syphilis is a sexually transmitted disease caused
by the spirochete Treponema (T.) pallidum, which af-
fects 12 million people each year and results in signif-
icant morbidity and mortality. Syphilis seropositivity
is common in some areas of the world and antenatal
syphilis infections are another aspect which should
not be forgotten (1-7). Despite the availability of rela-
tively sensitive tests and affordable treatment, the
disease remains a global health problem (8). Syphilis
can be treated easily and inexpensively with antibiot-
ics (9). The course of the infection may lead to various
clinical presentations, which are classified into early
(infectious) and late (non-infectious) stages. Early
syphilis can be further divided into primary and sec-
ondary syphilis and early latent syphilis. Late syphi-
lis includes late latent and various forms of tertiary
syphilis (10). While primary syphilis occurs as the re-
sult of direct contact of skin or mucous membranes
with those of an infected person (2), cutaneous and
mucosal lesions are outstanding manifestations of
secondary syphilis (11). Although the spirochetes or
their DNA can be consistently detected in lesions by
either microscopy (dark field, immunofluorescence) or
PCR, the most reliable method for laboratory diagnosis

84 ACTA DERMATOVENEROLOGICA CROATICA

Saral et al. Acta Dermatovenerol Croat
Serologic diagnosis of syphilis 2012;20(2):84-88
of syphilis, regardless of the stage of infection, is still
serology (9). However, the present international rec-
ommendations cannot be applied in some areas of
the world either because the recommended diagnos-
tic assays are not available or diagnostic strategies are
too expensive (7).
This study was designed to review serologic data
of syphilis patients in order to determine diagnostic
performance of three methods.
MATERIAL AND METHODS
Data on all cases of primary, secondary, and early
latent syphilis from 2008 through 2011 were extract-
ed from the Rize Education and Research Hospital
surveillance database. A total of 4226 serum speci-
mens were submitted to microbiology laboratory of
the Rize Education and Research Hospital for screen-
ing. The study was approved by the ethics committee
of the Rize University Education and Research Hospi-
tal. A total of 117 patients were diagnosed with syphi-
lis. The study included 117 patients (81 male and 36
female), aged 22-60 (mean 41.4±9.9) years, in whom
the diagnosis of syphilis was made by clinical, epide-
miological and laboratory methods (Fig. 1). The diag-
nosis of syphilis was made according to the Centers
for Disease Control (CDC) criteria; the diagnosis was
confirmed by examining the material from a chancre
using a special dark-field microscope or a blood test
(12). Three different methods were used for detection
of antibodies: Rapid Plasma Reagin (RPR), Treponema
pallidum hemagglutination (TPHA), and treponemal
Figure 1. Age of 117 patients diagnosed with syphilis
ACTA DERMATOVENEROLOGICA CROATICA
Chemiluminescence Microparticle enzyme Immu-
noassay (CMIA). The ARCHITECT Syphilis TP assay is
a chemiluminescent microparticle immunoassay for
qualitative detection of antibody to T. pallidum in hu-
man serum and plasma on the Architect System.
All samples were tested with the following syphilis
serology tests, according to the manufacturers’ in-
structions: (1) RPR test (Immutrep-RPR, Omega Diag-
nostics, Scotland, United Kingdom); TPHA test (Im-
mutrep-TPHA, Omega Diagnostics, Scotland, United
Kingdom); and CMIA test (Architect Syphilis TP, Ab-
bott Japan Co., Japan).
On the basis of consensus results derived from
these serologic tests (RPR, TPHA and CMIA), sera were
classified into the following categories: (1) syphi-
lis-negative, negative for both screening (RPR) and
confirmatory tests (TPHA, CMIA), and biological false
positives, which gave positive screening test results
that could not be confirmed by any of the confirma-
tory tests; and (2) probable syphilis positives, which
can be divided into probable past syphilis infection
(negative by screening but positive by confirmatory
tests); and probable active syphilis infection (posi-
tive by both screening and confirmatory tests). The
VDRL, TPHA and CMIA status of 117 samples studied
is shown in Table 1.
The sensitivity, specificity, positive predictive val-
ue (PPV), negative predictive value (NPV) and accura-
cy of the two methods were calculated against TPHA
(reference group) for the diagnosis of syphilis.
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Saral et al. Acta Dermatovenerol Croat
Serologic diagnosis of syphilis 2012;20(2):84-88

Table 1. VDRL, TPHA and CMIA status of 117 samples

VDRL CMIA
TPHA
Positive Negative Positive Negative
Positive 66 (58.9%) 46 (41.1%) 110 (98.2%) 2 (1.8%)
Negative 5 (100%) 0 (0.0%) 0 (0.0%) 5 (100%)

RESULTS specificity of RPR was very weak, the specificity of

As shown in Table 2, 4109 samples were negative
on both screening and confirmatory tests. Hence,
4109 samples were regarded as syphilis-negative,
whereas 66 samples were classified as probable active
syphilis (screening tests were positive and confirma-
tory test was positive); 46 samples were regarded as
originating from subjects who had probably had past
syphilis infections (screening tests were negative but
confirmatory test was positive); 5 samples were posi-
tive on RPR screening test but positive results could
not be confirmed by the confirmatory test. Hence,
positive RPR results were regarded as biological false
positives.
The most common clinical findings in symptom-
atic patients were chancre (11.1%) and condylomata
lata (7.6%). Clinical findings of study patients are
shown in Table 3.
Calculated sensitivity of RPR and CMIA against
TPHA was 58% and 98%, respectively. While the
Table 2. Serum samples according to their reactivity on
screening tests and confirmatory test
Total cases TPHA CMIA RPR
66 + + + of serologic test is insufficient for diagnosis because
46 + + - each type of test has some limitations (14). The sensi-
2 + - -
5 - - +
4109 - - - nemal serologic tests, the lack of sensitivity in early
CMIA was excellent, as shown in Table 4.
DISCUSSION
The incidence of syphilis has been rising in west-
ern countries, the prevalence of syphilis has increased
in human immunodeficiency virus (HIV) infected
persons in recent years, and congenital syphilis rates
closely parallel the rates of primary and secondary
syphilis in women of childbearing age (1,3). Diagnos-
tic and therapeutic approaches in syphilis show wide
variation (13). The traditional algorithm used in the di-
agnosis of the disease includes screening with a non-
treponemal test (microprecipitation reaction (MPR),
Venereal Disease Research Laboratory (VDRL), rapid
plasma reagin (RPR), with positive results confirmed
by a treponemal test (FTA-ABS test, ELISA, TPHA)
(7,12). All non-treponemal serologic tests measure an-
tibodies to cardiolipin. Generally, these tests take the
form of flocculation reactions. Heterophil IgG and IgM
antibodies to antigens associated with tissue damage
and to lipids in the cell wall of T. pallidum are detected
in such tests (7). Non-treponemal test antibody titers
may correlate with disease activity, and results should
be reported quantitatively. The use of only one type
tivity of non-treponemal tests depends on the stage
of syphilis (15). There are limitations of the non-trepo-

Table 3. Clinical findings of syphilis in 117 patients

Finding Primary stage (n) Secondary stage (n) % (N=117)
Chancre 13 -- 11.1
Lymphadenopathy 5 -- 4.2
Condyloma latum -- 9 7.6
Mucous patch -- 5 4.2
Rash -- 7 5.9

Table 4. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) and accuracy of the two methods
Test Sensitivity (%) Specificity (%) PPV (%) NPV (%) Accuracy (%)
RPR 58% 0% 92% 0% 56%
CMIA 98% 100% 100% 71% 98%

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Saral et al. Acta Dermatovenerol Croat
Serologic diagnosis of syphilis 2012;20(2):84-88
dark-field positive primary cases and in late syphilis,
and a relatively high incidence of false-positive reac-
tions (16,17). Another disadvantage of non-trepone-
mal tests is prozone phenomenon, which occurs in
up to 2% of patients and causes false-negative results
(15). Although the VDRL and RPR are equally valid as-
says, quantitative results from these two tests cannot
be compared directly because RPR titers frequently
are slightly higher than VDRL titers (14). Although
confirmatory tests are done with treponemal tests,
they in general cannot differentiate between syphi-
lis and other treponomatoses such as yaws and pinta
(18). The resurgence of syphilis in recent years and
widespread acceptance of enzyme immunoassays
(EIA) in clinical diagnostic laboratories have led to the
research and development of a number of EIA tests
for syphilis diagnosis (9). As automated treponemal
tests were presented to service and widespread use
in the screening, problems occurred in the interpreta-
tion of results (19). Architect Syphilis TP is a two-step
immunoassay for qualitative detection of IgG and/or
IgM to T. pallidum in human serum or plasma using
the chemiluminescent microparticle immunoassay
technology. Sample microparticles are coated with
recombinant T. pallidum antigens (TpN15, TpN17, and
TpN47). In our study, the sensitivity of CMIA against
TPHA was 98%. In another study, ARCHITECT Syphilis
TP performed with a sensitivity of 99.2%, higher than
TPHA (97.1%) in 244 patients with syphilis (20).
CONCLUSIONS
Based on our results, we think that automated
enzyme immunoassay systems could contribute to
reduce errors that depend on the person, especially
while monitoring titration changes. Considering the
suitability for automation, the high sensitivity and
specificity of ARCHITECT Syphilis TP make it a good
choice as a screening test in microbiology laborato-
ries. The limits of screening tests for the diagnosis
of syphilis should not be forgotten, i.e. confirmatory
tests must be done. Serology has the prime impor-
tance in the laboratory diagnosis of syphilis, but must
be viewed in the context of clinical presentation.
Acknowledgment
The authors would like to thank microbiology
technicians of the Rize Education and Research Hos-
pital for their contribution to the study.
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Ladies using Taky are sure for their beauty; year 1934.
(From the collection of Mr. Zlatko Puntijar)

88 ACTA DERMATOVENEROLOGICA CROATICA