Epidemiology and Pathogenesis

Key features of the epidemiology of diseases caused by the pathogenic treponemes are summarized in Table 46-2. In general, these organisms enter the host by either penetrating intact mucous membranes (as is the case for T. pallidum subsp. pallidum—hereafter referred to as T. pallidum) or entering through breaks in the skin. T. pallidum is transmitted by sexual contact and vertically from mother to the unborn fetus. After penetration, T. pallidum subsequently invades the bloodstream and spreads to other body sites. Although the mechanisms by which damage is done to the host are unclear, T. pallidum has a remarkable tropism (attraction) to arterioles; infection ultimately leads to endarteritis (inflammation of the lining of arteries) and subsequent progressive tissue destruction.

Spectrum of Disease

Treponema pallidum causes venereal (transmitted through sexual contact) syphilis. The clinical presentation of venereal syphilis is varied and complex, often mimicking many other diseases. This disease is divided into stages: incubating, primary, secondary, early latent, latent, and tertiary. Primary syphilis is characterized by the appearance of a chancre (a painless ulcer) usually at the site of inoculation, most commonly the genitalia. Within 3 to 6 weeks, the chancre heals. Dissemination of the organism occurs during this primary stage; once the organism has reached a sufficient number (usually within 2 to 24 weeks), clinical manifestations of secondary syphilis become apparent. During this phase the patient is ill and seeks medical attention. Systemic symptoms such as fever, weight loss, malaise, and loss of appetite are present in about half of the patients. The skin is the organ most commonly affected in secondary syphilis, with patients having a widespread rash with generalized lymphadenopathy. Aseptic meningitis may also occur. After the secondary phase, the disease becomes subclinical but not necessarily dormant (inactive); during this latent period, diagnosis can be made using serologic methods. Relapses are common during early (≤ 1 year) latent syphilis. Late latent syphilis (≥ 1 year) is usually asymptomatic and noninfectious. Tertiary syphilis is the tissue-destructive phase that appears 10 to 25 years after the initial infection in up to 35% of untreated patients. Complications of syphilis at this stage include central nervous disease (neurosyphilis), cardiovascular abnormalities, eye disease, and granuloma-like lesions, called gummas, found in the skin, bones, or visceral organs. Congenital syphilis is transmitted from mother to the unborn fetus during any stage of infection, but is most often associated with early syphilis. The unborn fetus may develop an asymptomatic infection or symptomatic infection with damage to the bone and teeth, deafness, neurosyphilis, or neonatal death.

The additional pathogenic treponemes are major health concerns in developing countries. Although morphologically and antigenically similar, these agents differ epidemiologically and with respect to their clinical presentation from T. pallidum. The diseases caused by these treponemes are summarized in Table 46-2.

Samples collected from ulcers and lesions should not be contaminated with blood, microorganisms, or tissue debris. The site should be cleansed with sterile gauze moistened with saline. The sample should be placed on a clean glass slide and cover slipped. Polymerase chain reaction (PCR) samples should be collected on a sterile Dacron or cotton swab and placed in a cryotube containing nucleic acid transport medium or universal transport medium. Tissue or needle aspirates of lymph nodes should be placed in 10% buffered formalin at room temperature. To test for congenital syphilis, a small section of the umbilical cord is collected and fixed in formalin or refrigerated until processed. Serum is the specimen of choice for serology; however, whole blood or plasma may be used in some assays.

Direct Detection

Treponemes can be detected in material taken from skin lesions by dark-field examination or fluorescent antibody staining and microscopic examination. Material for microscopic examination is collected from suspicious lesions. The area around the lesion must first be cleansed with a sterile gauze pad moistened in saline. The surface of the ulcer is then abraded until some blood is expressed. After blotting the lesion until there is no further bleeding, the area is squeezed until serous fluid is expressed. The surface of a clean glass slide is touched to the exudate, allowed to air dry, and transported in a dust-free container for fluorescent antibody staining. A T. pallidum fluorescein-labeled antibody is commercially available for staining (Viro Stat, Portland, Maine). For dark-field examination, the expressed fluid is aspirated using a sterile pipette, dropped onto a clean glass slide, and cover slipped. The slide containing material for dark-field examination must be transported to the laboratory immediately. Because positive lesions may be teeming with viable spirochetes that are highly infectious, all supplies and patient specimens must be handled with extreme caution and carefully discarded as required for contaminated materials. Gloves should always be worn.

Material for dark-field examination is examined immediately under 400× high-dry magnification for the presence of motile spirochetes. Treponemes are long (8 to 10 µm, slightly larger than a red blood cell) and consist of 8 to 14 tightly coiled, even spirals (Figure 46-2). Once seen, characteristic forms should be verified by examination under oil immersion magnification (1000×). Although the darkfield examination depends greatly on technical expertise and the numbers of organisms in the lesion, it can be highly specific when performed on genital lesions.

Lesion exudates or tissue samples may be used for direct fluorescent antibody detection for T. pallidum (DFA-TP). DFA-TP visualizes specimens on slides with fluorescein isothiocyanate (FITC) labeled antibodies. Polyclonal and monoclonal antibodies may be used; however, the Food and Drug Administration (FDA) in the United States has not approved this test.

Serodiagnosis

Serologic tests for treponematosis measure the presence of two types of antibodies: treponemal and nontreponemal. Treponemal antibodies are produced against antigens of the organisms themselves, whereas nontreponemal antibodies, often referred to as reagin antibodies, are produced in infected patients against components of mammalian cells. Reaginic antibodies, although almost always produced in patients with syphilis, are also produced in patients with other infectious diseases such as leprosy, tuberculosis, chancroid, leptospirosis, malaria, rickettsial disease, trypanosomiasis, lymphogranuloma venereum (LGV), measles, chickenpox, hepatitis, and infectious mononucleosis; noninfectious conditions such as drug addiction; autoimmune disorders, including rheumatoid disease and systemic lupus erythematosus; and in conjunction with increasing age, pregnancy, and recent immunization.

The two most widely used nontreponemal serologic tests are the Venereal Disease Research Laboratory (VDRL) and rapid plasma reagin (RPR) tests. Each of these tests is a flocculation (or agglutination) test, in which soluble antigen particles are coalesced to form larger particles that are visible as clumps when they are aggregated in the presence of antibody. The VDRL is used as a quantitative test and may be performed on serum or CSF in suspected cases of neurosyphilis. See Procedures 46-1 and 46-2 on the Evolve site for details and limitations for the VDRL and RPR.

Procedure 46-1   Rapid Plasma Reagin (RPR) Test

Purpose

The rapid plasma reagin (RPR) test is a presumptive serologic screening test for syphilis.

Principle

The test is based on the presence of reagin, a nonspecific antilipid antibody found in the serum of patients infected with T. pallidum. Syphilis infection results in the breakdown of human tissue releasing fatty substances that combine with T. pallidum protein to form an antigen, resulting in the formation of nonspecific and specific T. pallidum antibodies. The RPR antigen consists of cardiolipin, lecithin, and cholesterol bound to charcoal particles. The nonspecific antibody reacts with the test antigen resulting in flocculation. The flocculation is visible on a white test card by incorporating charcoal particles in the antigen preparation.

Specimen

Patient serum or plasma is tested at room temperature, 20° to 25° C.

Method

1. Place a 50 µL sample of patient serum or plasma, positive and negative controls into three separate circles on the test card.

2. Mix the RPR-carbon reagent gently prior to applying to each circle.

3. Add exactly one drop (20 µL) of reagent to each sample.

4. Mix the specimen and reagent with a stirrer carefully spreading each sample over the entire surface of the circle. Use a clean stirrer for each sample.

5. Place the test card on a mechanical rotator or shaker at 80 to 100 rpm for 8 minutes.

6. Read the results prior to drying.

Expected Results

Reactive: samples that demonstrate small to large clumps; a weak positive may show fine granulation or partial clumping.

Nonreactive: samples that demonstrate no clumping or very slight roughness.

Limitations

1. Slide flocculation tests are affected by room temperature. All test reagents and specimens should be warmed to room temperature prior to testing.

2. Failure to read results within 8 minutes may lead to drying and the appearance of false-positive results.

3. Patient samples that are hemolyzed, lipemic, or contain a high concentration of bilirubin may interfere with test results.

Quality Control

Positive controls should demonstrate readily visible flocculation

Negative controls will appear uniformly turbid.

Procedure 46-2   Venereal Disease Research Laboratory (VDRL) Test

Purpose

The Venereal Disease Research Laboratory Test (VDRL) is a slide flocculation test used for screening patients for infection with T. pallidum.

Principle

Patients infected with T. pallidum produce nonspecific antibodies capable of reacting with the cardiolipin test antigen. Cardiolipin is a lipid antigen extracted from beef heart that contains cardiolipin, lecithin, and cholesterol. The VDRL test is typically positive within 1 to 2 weeks following the appearance of the primary lesion. The test becomes reactive in late phase primary syphilis and highly reactive in secondary syphilis. The results will slowly decrease and become less reactive in late or tertiary syphilis. The VDRL test is also useful in the diagnosis for congenital syphilis. Maternal antibodies are capable of crossing the placenta; a positive VDRL immediately following birth may be solely a result of the presence of maternal antibodies. A quantitative titer is therefore required at birth, followed by a second titer approximately 1 month following birth. No increase in titer will assist in ruling out the possibility of congenital syphilis.

Specimen

Patient serum or CSF samples. The serum must be inactivated prior to testing. The sample is heated at 56° C for 30 minutes. This inactivates serum proteins such as complement within the patient specimen. CSF samples should not be heated prior to testing.

Method

1. Place 50 µl of patient sample, a positive reactive, and a negative nonreactive control into separate circles on the test card.

2. Place 20 µl of reagent next to the sample and stir each, making sure to completely cover the circle on the test card. Use a separate stirrer for each sample to avoid contamination.

3. Place the test card on a rotator or shaker at 180 rpm for 4 minutes.

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