ROCHELLE D.

DARLUCIO, RMT
OUR LADY OF FATIMA UNIVERSITY
COLLEGE OF MEDICAL LABORATORY SCIENCE
 -- bacterial infection caused by spirochetes

SYPHILIS
◦ most commonly acquired spirochete disease
in the US
Treponema pallidum subs. pallidum
◦ Causative agent: __________________

◦ Mode of Transmission:
◦ Sexual transmission -- most common start as a painless sore (lesion on genital/rectum/mouth) depending on
the site of contact w/ bacteria
◦ Transmission to the fetus is possible in mothers with -- bacteria can remain inactive for a decade & maging active ulit
clinically latent disease -- can be cure (antibiotic: penicillin)
-- if untreated: brain damage, heart, or organ damage
◦ Parenteral exposure -- can be life threatening
Detect
Antibodies against treponemal antigens and nontreponemal cardiolipin antigens (Wasser-mann antigens) develop
and elicit a cell-mediated and humoral Immune response, which results in the formation of immune complexes.
Incubation period: T. pallidum enters the body, reaches the bloodstream, and
is disseminated to all organs. This early asymptomatic phase lasts 10 days to
(incubation)
10 weeks.
 STAGES OF SYPHILIS
Primary (Early) Stage
"chancre" -- appear on spot where bacteria enter body
◦Initial lesion: ______________
◦appear 2 to 8 weeks after infection and last for 1 to 5 weeks.
◦Serum tests for syphilis are positive in 90% of patients after 3 weeks. (IgM)
 STAGES OF SYPHILIS
Secondary Stage
◦6 to 8 weeks after chancres first appear
◦generalized lymphadenopathy, ,malaise, fever, pharyngitis, and rash on skin and mucous
membranes, and Involvement of the CNS.
◦lesions are highly contagious, but heal spontaneously within 2 to 6
weeks.
◦Serologic tests are positive in secondary syphilis.
◦Antibodies are mostly IgG.
STAGES OF SYPHILIS
Latent Stage
◦ contagious and is generally considered to begin after the second year of
infection.
◦characterized by lack of clinical symptoms
◦Patients are non infectious except for pregnant women who can pass the disease to the fetus

Serologic tests are still positive.

 STAGES OF SYPHILIS
Tertiary Stage
◦Occurs between 10-30 years
after secondary stage
◦3 MANIFESTATIONS:
◦Gummatous syphilis
◦Cardiovascular disease
◦Neurosyphilis
Gummas – localized areas of granulomatous
inflammation that are most often found in bones, skin
or subcutaneous tissues
Cardiovascular complication – usually involve the
ascending aorta, symptoms are due to destruction of
elastic tissue, especially in the ascending and transverse
segment of the aortic arch.
Neurosyphilis- complication most often associated with
the tertiary stage, but it can actually occur anytime after
the primary stage and can span all stages of the disease.
 Congenital syphilis
Infection of the fetus causes late abortion, stillbirth, neonatal death, neonatal
disease, or latent infection.

Outcome: depends on the stage of the mother's disease

-diffuse maculopapular desquamatous rash (particularly around the mouth and on

the palms and soles)
-hemolytic anemia, jaundice, hepatosplenomegaly, abnormal cartilage and bone
involvement, and mental retardation.
 LABORATORY DIAGNOSIS
1. DIRECT DETECTION
◦ Dark field microscopy or
Fluorescent Antibody
testing
◦ Primary and secondary
syphilis can be diagnosed

◦ Requirement for direct

detection: ACTIVE LESION
 LABORATORY DIAGNOSIS
2. SEROLOGICAL TESTS
2.1 Non-treponemal Serologic Test Components of Non-
◦determine the presence of an
treponemal methods:
antibody that forms against Cardiolipin
cardiolipin (Reagin)
◦Principle: Flocculation
Cholesterol
◦Positive within 1-4 weeks after the Lecithin
appearance of primary chancre.
 Non-Treponemal Tests
Composition of VDRL Ag Reagent:
2.1.1 Venereal Disease Research
◦0.03% cardiolipin-__________
Laboratory (VDRL)
◦0.9% cholesterol-___________
◦ both qualitative and quantitative slide ◦0.21% lecithin-_____________
flocculation test for serum

◦ Serum are heated at ________________

to inactivate complement
 Non-Treponemal Tests
*****
Venereal Disease Research
SERUM VDRL:
Laboratory (VDRL) ◦ Size of ring: 14mm diameter
◦Sera and patient samples are spread out to fill the entire ring. ◦ Ag delivery needle: 18 gauge needle
◦One drop (1/60 mL) of the VDRL antigen is then added to
each ring.
◦The slide is rotated for 4 minutes on a rotator at 180 rpm. CSF VDRL: neurosyphilis
◦It is read microscopically to determine the presence of ◦ Size of ring: 16mm diameter
flocculation.
◦ Ag delivery needle: 21 or 22 gauge
needle
◦ Rotate for 8 mins at 180 rpm
 Non-Treponemal Tests
2.1.2 RAPID PLASMA REAGIN (RPR)
modified VDRL test involving macroscopic
agglutination
RPR Antigen Reagent: similar to the VDRL
antigen with the addition of the following:
◦Charcoal-__________
◦EDTA-_____________
◦Thimerosal-________
◦Choline chloride-____
RAPID PLASMA REAGIN (RPR)
Specimen: 0.05 mL of serum

- Patient serum is placed in a test circle (18 mm) on a

plastic-coated disposable card.
-Antigen is dispensed from a small plastic dispensing
bottle with a calibrated 20-gauge needle.
-The card is mechanically rotated for 8 mins at 100rpm.
-Cards are read under a high-intensity light source.
 LABORATORY DIAGNOSIS
2.2 Treponemal Serologic Test
◦detects antibody directed 2 Sources of Ag:
against the T.pallidum organism 1. Non pathogenic Reiter strain
or against secific treponemal 2. Pathogenic Reiter strain
antigens

◦100% reactive in secondary and

latent syphilis
 Treponemal Tests
2.2.1 Fluorescent Treponemal
Antibody Absoption Test (FTA-ABS)
Principle: Indirect Immunofluorescence
-one of the earliest confirmatory test
- Highly sensitive and specific, but it is time
consuming to perform
◦ Rgt antigen: Nichol’s strain dried and fixed on slide
◦ Absorbent: Reiter treponemes
FTA-ABS
Specimen - 1:5 heat inactivated serum reacted with sorbent
containing non-pathogenic treponemes (Reiter strain)
Slides: contains Nichol’s strain of T. pallidum
Incubated at 37 C
ͦ for 30 minutes
Positive result: green fluorescence
REPORTED on a scale of 0-4+
0 : no fluorescence (non-reactive)
1+ : minimally reactive, test must be repeated with a
second specimen drawn in 1-2 weeks
2-4+ : REACTIVE

Washed with deionized water then rinse with phosphate-

buffer saline for 5 minutes
Add antibody conjugate (antihuman immunoglobulin
conjugated with fluorescein I isothiocyanate label)
Incubated at 37 C
ͦ for 30 minutes
Wash
Examine under fluorescent microscope
 Treponemal Tests
2.2.2 Hemagglutination
Treponemal Test for Syphilis
Rgt antigen: glutaraldehyde
stabilized turkey RBC coated with
treponemal antigen
 Treponemal Test
2.2.3 Treponema pallidum
Immobilization Test
◦ Standard test
◦ Principle: the antibody produced vs T.
pallidum + complement can immobilized
the live treponemes
◦ Rgt antigen: live actively motile T. pallidum
organisms

◦(+) >50% immobilized

treponemes
2.2.4 Treponema pallidum Particle Agglutination
(TP-PA) Test

use colored gelatin particles coated with treponemal

antigens
More sensitive in detecting primary syphilis
Traditional testing algorithm for syphilis