Articles on Medical Diseases and Conditions

Tag: Rickettsial Infections

  • Other Rickettsial Diseases

    Bartonellosis (Oroya fever or Carrion’s disease) is caused by the rickettsial organism Bartonella bacilliformis and occurs only in Andean mountain regions of South America. The vector is the sandfly Phlebotomus (species). The organism is related to Rochalimaea quintana, the rickettsial etiology of trench fever and therefore is also related to Rochalimaea henselae, one of the etiologies of cat scratch fever.

    Q fever is a rickettsial disease whose etiology is Coxiella burnetii. Goats and cows are the major vectors; infection occurs through exposure to contaminated milk or animal placentas, or may occur by inhalation of infected material. The organism is very resistant to drying, so that dust inhalation can spread the disease in dry areas. The incubation period is 2-4 weeks. Clinical disease is similar to moderately severe influenza. Infection usually (not always) does not produce a rash, which is unusual among the rickettsiae. Liver function tests are abnormal in 80% of patients, but only to a small degree. Diagnosis is usually by immunofluorescent serologic tests, generally performed in large reference laboratories or public health laboratories. The Weil-Felix test panel is nonreactive.

    Ehrlichiosis is a rickettsial disease caused by Ehrlichia chaffeensis. The disease is spread by a tick vector and is clinically similar to Rocky Mountain Spotted Fever, except that only about 20% of patients develop a rash. Over half of Ehrlichiosis patients develop some degree of leukopenia (74%-85%) or thrombocytopenia (72%-84%), and mild aminotransferase enzyme level elevations (78%-100%) are common. Usually the disease is not severe. About one third of patients have symptoms that raise the question of CNS involvement. CSF shows elevated WBC count in 67%-71% of cases, with lymphocytes predominating in one third of cases. Total protein is elevated in 33%-62% of cases. There is also a canine Ehrlichiosis caused by a related but not identical tick-borne Rickettsia. In fact, for several years the usual diagnostic test for human Ehrlichiosis was an immunofluorescent test based on cross-reaction with canine Ehrlichiosis organisms. Now indirect immunofluorescent assays (IFA) are available that are specific for E. chaffeensis and that are being done in some large reference laboratories or public health laboratories. Acute and convalescent serum specimens may be required to document active or recent infection (vs. old infection).

  • Rickettsial Diseases

    The rickettsiae to some extent resemble small bacteria but are not stained with Gram stain and cannot be cultured on artificial media. These organisms are spread only by insect vectors that have fed on blood from a patient with the disease, not from personal contact with a patient. Blood cultures can sometimes isolate and identify rickettsiae, especially in Rocky Mountain spotted fever. The blood specimens should be frozen and sent to the laboratory packed in dry ice. Since artificial culture media are not available, chick embryo or live animal inoculation must be done. Very few laboratories will perform rickettsial culture. Therefore, serologic tests or other procedures by far overshadow culture as diagnostic aids. The most commonly used procedure is the Weil-Felix reaction. This test takes advantage of the fact that certain rickettsial diseases produce antibodies that also react (or cross-react) with antigen contained in certain strains of Proteus bacteria. These Proteus groups are called OX-19 and OX-K. Titers of 1:80 are suspicious, and titers of 1:160 are definitely significant. Antibodies appear 7-10 days after onset of illness. Rickettsial diseases that may be diagnosed by means of the Weil-Felix reaction are the following:

    Rickettsial Diseases

    Unfortunately, there are serious limitations to the Weil-Felix test. First, there are a fairly large number of borderline false positive results as well as occasional outright false positives. Since the Weil-Felix reaction depends on Proteus antigen, urinary tract infection by Proteus should be ruled out if the Weil-Felix test result is positive. Second, about two thirds of patients with Rocky Mountain spotted fever or Brill’s disease (recrudescent typhus) have false negative reactions. In these two diseases, other serologic tests (e.g., the microimmunofluorescence procedure) are preferred to the Weil-Felix test.

    Rocky Mountain spotted fever

    Rocky Mountain spotted fever occurs in spring and summer, predominantly in the eastern two thirds of the United States (the area east of the Rocky Mountains). The disease is transmitted through the bite of an infected tick, with an incubation period of 3-12 days. Fever, severe frontal headache, severe myalgias, and a macular or maculopapular skin rash that develops on the third to fifth day and eventually develops a petechial component are the most characteristic clinical findings, with the rash typically involving the palms and soles. Nausea, vomiting, and diarrhea are reported in about 60% of patients and abdominal pain is reported in about 35%. The WBC count is usually normal, but thrombocytopenia is said to be common. In severe cases disseminated intravascular coagulation may develop. Serologic tests include the Weil-Felix test, CF test, microimmunofluorescence (MIF) test, latex agglutination (LA) test, and tissue biopsy with immunofluorescent staining. The Weil-Felix test has very poor sensitivity and is not specific, and the antibody does not develop until some time between 7-21 days after infective contact (compared with incubation period of 3-12 days). Complement fixation is more specific but detects 50% or fewer cases and is very laborious to perform, the antibody does not appear until 14-21 days after infection, and antibody elevations persist for years. Antibodies detected by the MIF and LA tests begin to appear 7-10 days after onset of illness. These tests are reported to detect considerably more than one half of Rocky Mountain spotted fever cases but are available only in reference laboratories or public health laboratories. The MIF test is currently considered the gold standard. LA is reported to detect about 80%-85% of patients detected by MIF. Skin biopsy of a lesion with immunofluorescent antirickettsial antibody studies on the tissue can also be done but the degree of sensitivity is controversial (50%-70%) and the test is not widely available.

    Cat scratch disease

    This condition has two possible etiologies. One is a small bacillus that stains faintly gram-negative, and the other is a member of the Rickettsial family. About 80% of patients affected are children (about 90% are less than 18 years old and about 70% are less than 10 years old). About 85% (range, 83%-90%) have a history of contact with cats, most of which (about 80%) are kittens or less than 1 year old. Cat inoculation via scratch (57%-83% of patients), bite, or lick is followed in 4-14 days (range, 3-50 days) by development of a primary inoculation site pustule or papule in 54%-93% of patients. The primary lesion persists 1-3 weeks but can persist up to 1-3 months. Regional lymphadenopathy develops in 1-3 weeks after appearance of the primary site lesion; the nodes are unilateral in about 90% (range, 85%-95%) of patients and are tender in about 80%. Cat scratch disease is possibly the most common cause of chronic lymphadenopathy in children. The node or nodes are located in the head and neck area in about 25% of cases; the axilla (occasionally the epitrochlear area) in about 45%, and the groin in about 20%. They generally regress in size in 2-4 months but can persist longer. Adenopathy is accompanied by fever and malaise in about 35% of patients (range, 30%-59%), but the fever is usually (90%) less than 102°F (38.9°C). The lymph nodes suppurate in 10%-30% of cases and may drain in 10%-20% of cases. Parinaud’s oculoglandular syndrome (unilateral conjunctivitis with regional lymphadenitis) is present in 4%-6% of patients. The lymph nodes often raise the clinical question of mycobacterial infection and sometimes of malignant lymphoma. Biopsy specimens display a characteristic lesion consisting of lymphocytes and also histiocytes with scanty cytoplasm surrounding a central area containing segmented neutrophils that usually becomes necrotic. The lesions may be single or can be stellate (branching) due to coalescence of the original lesions. The same histologic pattern can be found in tularemia and in lymphogranuloma venereum. It is extremely difficult to find the organisms by Gram stain, and more success is obtained with the Warthin-Starry silver stain. Interestingly, the more commonly used silver stain methods such as methenamine silver are not useful. Even with Warthin-Starry, the organisms may be difficult to find or may be absent. The best areas are the center of suppurative granulomas. A skin test has been described using antigen derived from lymph node pus, but the antigen is not readily available.

    In 1988, scientists at the Armed Forces Institute of Pathology (AFIP), using special media, cultured a gram-negative rod from several patients with cat scratch disease which was subsequently given the name Afipia felis. However, only a few years later it was found that considerably more patients with cat scratch disease had antibodies to a rickettsial organism named Rochalimaea henselae than evidence of Afipia felis infection. Later, R. henselae was isolated by culture from several patients with cat scratch disease and now appears to be the most frequent cause. Both organisms can be isolated using chocolate agar in 5% CO2 atmosphere; but growth may take up to 6 weeks. Both organisms also grow in Haemophilus test medium.

    The Rochalimaea genus includes two species, Rochalimaea quintana (etiology of trench fever) and R. henselae. R. henselae is now considered the etiologic agent of several conditions. One is bacillary angiomatosis, which is focal proliferation of small blood vessels, endothelial cells, and connective tissue cells (somewhat similar in appearance to Kaposi’s sarcoma), and which is found predominantly in the skin but sometimes in the liver or spleen, most often in patients with HIV-1 infection. R. quintana can also cause these lesions. Another condition is peliosis hepatis (multicystic angiomatoid structures with endothelium not seen by ordinary microscopy; usually in the liver). It also causes occasional cases of septicemia, most often in HIV-infected patients, and is one cause of cat scratch fever. The organism is a small gram-negative curved bacillus that requires special culture conditions for isolation. Diagnosis can be made by Warthin-Starry silver stains from tissue biopsies and culture on special media noted previously. There have been two published reports of a fluorescent antibody test for R. henselae and for A. felis. The sensitivity of these tests is not yet established. Nucleic acid probe methods have been used experimentally and will probably also become commercially available in the future, although most likely in many patients the disease will be treated empirically.

  • Vincent’s Angina

    Vincent’s angina (Vincent’s infection) is an infection of the mouth caused by an interesting synergistic group of organisms, including anaerobic streptococci, a fusiform gram-negative bacillus, and a spirochete. Gram-stained smears demonstrating all three organisms are usually sufficient for diagnosis.

  • Leptospirosis

    Leptospirosis is caused by several species of Leptospira organisms found most often in rats but sometimes present in some farm animals and in some cats and dogs (presumably from rat-transmitted infection). Transmission is most often through accidental contact with water contaminated by infected rat urine. Those most at risk are sewer workers and slaughterhouse employees, but farmers and campers sometimes come into contact with contaminated water. There is an incubation period of 4-20 days, then abrupt onset of fever, often accompanied by chills, headache, malaise, and conjunctivitis. Muscle pain is present in 50% of cases. The fever typically lasts 4-9 days. WBC count can be normal or elevated. Urinalysis often contains protein and some WBCs. Serum bilirubin is usually normal, but about 10% of cases have mild elevation. Alanine aminotransferase is elevated in about 50% of cases, usually to less than five times normal. About 50% of patients experience a recurrence of fever about 1 week (range, 2-10 days) after the end of the first febrile period. Patients are more likely to demonstrate signs of hepatitis in this phase and may develop symptoms of meningitis. The most severe form of leptospirosis is called Weil’s disease and occurs in about 5% of infections. The most striking findings are a combination of hepatitis and glomerulonephritis, clinically manifested by jaundice with hematuria. Therefore, the disease is sometimes considered in the differential diagnosis of jaundice of unknown etiology. Symptoms of meningitis occasionally predominate. Laboratory findings include leukocytosis with a shift to the left. A mild normocytic-normochromic anemia usually develops by the second week. Platelet counts are normal. After jaundice develops, liver function test results are similar to those in viral hepatitis. After onset of kidney involvement, the blood urea nitrogen (BUN) level is often elevated, and hematuria is present with proteinuria. CSF examination shows normal glucose levels but increased cell count, which varies according to the severity of the case; initially, these are mainly neutrophils, but later, lymphocytes predominate. Cultures on ordinary bacterial media are negative.

    Diagnosis often requires isolating the organisms or demonstrating specific antibodies in the serum. During the first week (days 1-8), spirochetes may be found in the blood by dark-field examination in about 8% of cases and can be cultured from the blood in many more. Instead of ordinary blood cultures, one to three drops of blood are inoculated into a special culture medium (Fletcher’s), since larger quantities of blood inhibit the growth of leptospires. The CSF may be cultured toward the end of the first week. During the second week the blood results quickly become negative. During the third week (days 14-21) the spirochetes may often be recovered from the urine of patients with nephritis. Animal inoculation is the most successful method. Antibodies start to appear at about day 7 and are present in most cases by day 12. Antibodies persist for months and years after cure. A titer of 1:300 is considered diagnostic, although without a rising titer, past infection cannot be ruled out completely. If a significant titer has not developed by day 21, it is very rare for it to do so later. In summary, blood cultures during the first week and serologic tests during the second and third weeks are the diagnostic methods of choice.

  • Lyme Disease

    Lyme disease is caused by the spirochete Borrelia burgdorferi by means of several tick vectors, the principal one in the Northeast and North Central United States being the deer tick Ixodes dammini and in the Pacific Coast states, Ixodes pacificus, the Western black-legged tick (both morphologically “hard” ticks). The three major affected areas in the United States are the northeastern states (New Jersey to Connecticut), the far western states, and the upper midwestern states. However, cases have been reported elsewhere and also in Canada, Europe, and Australia.

    Ixodes dammini has a 2-year, three-form life cycle. The very young ticks (called larval stage, although the organism has a tick shape) feed on a vector organism, usually the white-foot mouse, and then are dormant until the following spring. The larval ticks are very small and have only three pairs of legs, like insects. The following year in the spring the larval tick changes to the nymph stage, which has four pairs of legs like the adult stage.

    In 50%-80% of patients, about 1 week (range, 3-68 days) after the tick bite, a reddish macular expanding lesion with central clearing (“erythema chronicum migrans”) develops on the skin at the inoculation site often followed by similar skin lesions in some other areas. This usually fades within 2-3 weeks (range, 1 day-4 weeks) and is usually accompanied by low-grade fever, weakness, fatigue, and regional lymphadenopathy. Although this characteristic skin lesion should strongly suggest Lyme disease, only 20%-30% of patients recall such a lesion. Migratory arthralgias and myalgia are frequently present. About 10% of patients develop anicteric hepatitis. In the second stage of illness, CNS (most often aseptic meningitis) or peripheral nervous system abnormalities (Bell’s palsy or Bannwarth’s polyneuritis syndrome) occur about 4 weeks (range, 2-8 weeks) after the tick bite in about 15%-20% of patients (range, 11%-35%). About 7% (range, 4%-10%) of patients develop transitory ECG abnormalities or myocardial inflammation, usually about 5 weeks after the tick bite (range, 4 days-7 months). In the third stage of illness, about 40% (range, 26%-60%) of patients develop recurrent arthritis. This is the most famous manifestation of Lyme disease and involves one or more joints, most commonly the knee, beginning about 6 weeks-6 months after the tick bite (range, 4 days-2 years).

    Laboratory test abnormalities include elevated erythrocyte sedimentation rate in about 50% of cases. Peripheral blood WBCs are elevated in only about 10%; fluid aspirated from arthritic joints is similar to that from patients with rheumatoid arthritis. CSF in patients with meningeal or peripheral nerve symptoms usually show increased numbers of WBCs with lymphocytes predominating, normal glucose and mildly increased protein levels, oligoclonal bands similar to those of multiple sclerosis, and CSF-IgM antibody present.

    Culture can be done from biopsy of the erythema migrans (ECM) skin lesion; best results are obtained from the advancing edge of the lesion. Transport of the specimen and specimen culture in the same special BSK culture media plus incubation for several weeks if necessary has produced best results; but even so the spirochetes were isolated in less than 45% of cases (range, 5%-71%). Warthin-Starry silver stains on ECM lesion biopsy demonstrates spirochetes in less than 40% of cases. Blood cultures may be positive in the second stage of illness but only in 2%-7% of cases and therefore is not cost-effective. Culture of CSF in second-stage symptomatic patients may be positive in about 10% of patients. DNA probes with PCR amplification have been reported to have a sensitivity of 80% when performed on a biopsy of the ECM skin lesion, the same or better than the best culture results. However, thus far, DNA probe for Borrelia antigen in blood has not substantially improved serologic test results.

    Currently, the most helpful procedures are serologic tests. IgM antibody levels rise about 2-4 weeks after onset of ECM, peak about 6-8 weeks after ECM onset, and usually become nondetectable by 4-6 months after onset. However, some patients have persistent IgM levels, presumably due to continued infection or reinfection. IgG antibody levels rise about 6-8 weeks after onset of erythema migrans and peak at about 4-6 months after onset of erythema migrans, but may not peak until later or even more than a year. The highest IgG levels tend to occur when patients develop arthritis. IgG levels typically remain elevated for life. The most commonly used tests are immunofluorescent and ELISA methods. False positive results can be obtained in patients with other spirochetal diseases, such as syphilis, relapsing fever, and leptospirosis, and according to one report also in subacute bacterial endecarditis (SBE). Some of the ELISA tests attempt to adsorb out some of these antigens if they are present. Both test methods can give about the same results, although investigators generally seem to have a more favorable opinion of ELISA. In the earliest stage of the disease (ECM present 1-7 days), serologic tests are rarely positive. Later in the first stage, 3-4 weeks after onset of ECM, the tests are positive in about 40% of patients. In the second stage of illness (coincident with systemic symptoms) about 65% are positive, and in the third (arthritic) stage, about 90%-95% (range, 80%-97%) are positive. This suggests that negative serologic tests in clinical stages one and two may have to be repeated 3-4 weeks later. ELISA tests using recombinant flagellar proteins as antigen somewhat improve IgM test specificity and may increase sensitivity a little in early disease compared to ELISA tests using whole organism alone. Sensitivity of IgG antibody is significantly greater than IgM in the second and third stages of Lyme disease because disseminated (second stage) infection raises IgG more than IgM (which has already peaked or has already started to decline).

    Evaluation of different kits has shown considerable variation in sensitivity and specificity between different kits, between laboratories, and even between evaluations in the same laboratories when the same specimen was repeated later. Western blot testing is commercially available or performed with homemade reagents. This has the advantage of visually identifying which proteins are reacting to patient antibodies. Unfortunately, there still is little agreement how to interpret the Lyme Western blot test. Some of the proteins that are rather frequently detected are shared with other organisms. Some of the more specific proteins (outer coat proteins A and B) may not appear until relatively late in some patients. Nucleic acid probe testing has recently been reported, with or without PCR amplification, mostly using homemade reagents. Although results have been more sensitive than some standard ELISA or fluorescent antibody kits, DNA probes so far have not increased usable sensitivity as much as has been achieved in some other diseases. Finally, some studies have reported that some patients with Lyme disease have a reactive antinuclear body (ANA) test, usually the speckled type. One report found that the VDRL or RPR test for syphilis is usually nonreactive.

    In one report from a Lyme disease referral center, of 788 patients with positive serologic test results for Lyme disease, 23% had active Lyme disease, 20% had previous Lyme disease, and 57% were judged not to have evidence of Lyme disease.

  • Congenital Syphilis

    It has been estimated that 75% of infants born from mothers with active syphilitic infection will become infected in utero. Of those infected about one third will be symptomatic in the neonatal period and two thirds will have some symptoms within the first 2 years of life. However, it appears that the majority of patients with congenital syphilis are not diagnosed until late childhood or adolescence.

    Congenital syphilis often gives a confusing serologic picture. Syphilitic infants usually have a positive RPR reaction. Sometimes, however, these infants have a nonreactive RPR result at birth, and the RPR responses may remain normal up to age 3 months before the titer begins rising. On the other hand, if the mother has a positive RPR reaction, even though she was adequately treated, many infants will have a positive RPR reaction due to passive transfer of maternal antibodies through the placenta. The same is true for the FTA-ABS test. However, if the VDRL or RPR titer of the infant is fourfold higher than that of the mother, this suggests congenital syphilis. Also, a fourfold or greater rising titer over the weeks or months after birth strongly suggests infection. To further confuse the picture, pregnancy is associated with an increased number of BFP reactions in the RPR, and reportedly, on occasion, even with the FTA-ABS. If the mother was adequately treated before delivery, the infant’s reactive RPR will revert to nonreactive without treatment in approximately 3-4 months. Cord blood may cause problems in neonatal syphilis testing due to possible contamination with maternal blood or false negative results caused by interference from hemoglobin or maternal infection very late in pregnancy. A modification of the FTA-ABS test that is specific for IgM antibodies is reported to detect and diagnose most cases of congenital syphilis in the newborn. However, reports indicate as many as 10% false positive results and as many as 35% false negative results. An IgM ELISA immunoassay (DCL-M) has been reported to be more sensitive and specific. A Western blot test for syphilis IgM antibody has been reported, with results also better than the FTA-IgM. However, more evaluations are needed for both the ELISA and Western blot.

  • Effect of Therapy on Test Results

    Studies before 1990 indicated that if patients with their first syphilitic infection in the primary or secondary stage were given adequate treatment, an elevated VDRL titer would decline (on the average) 2 dilutions (fourfold) by 3 months and 3 dilutions by 6 months. The rate of decline was said to be faster when the pretreatment VDRL titer was higher and more slowly when the VDRL titer was lower or when the infection was long-standing or was a reinfection. A large Canadian study published in 1991 confirmed some of these observations and disagreed with others. The Canadian study found that the first infection and the earlier stages of syphilis were more likely to respond serologically to treatment and responded faster, in agreement with previous studies. However, the Canadian study, using the RPR and current treatment recommendations, noted a slower rate of RPR decline, with a 2-tube dilution decrease (on the average) not attained until 6 months and a 3-tube decrease not achieved until 12 months. Although the rate of decrease was similar at all dilutions, the time needed to decrease to nonreactive was (as expected) much earlier with lower pretherapy titers than with high titers. In first-infection primary syphilis, if the initial titer was 1:8, the RPR (on the average) became nonreactive in 26% of patients by 3 months, in 54% by 6 months, in 70% at 1 year, and in 81% by 3 years. If the initial titer was 1:256, no patients were nonreactive at 3 months, only 6% at 1 year, and 31% at 3 years. In the early latent stage with a pretherapy titer of 1:8, 7% were nonreactive at 3 months, 31% at 1 year, and 66% at 3 years.

    In the Canadian study the FTA-ABS became nonreactive after therapy in 11% of first-infection primary stage patients at 1 year and 24% at 3 years. The FTA-ABS or the MHA-TP seldom became nonreactive when therapy began in the secondary stage and in no patients after the secondary stage.

    One study reported that HIV infection in its late stage may result in loss of FTA-ABS and MHATP reactivity in 17% of cases. In one study, BFP results were obtained using the RPR method in 4% of HIV-positive patients (all in males) versus 0.8% incidence in non-HIV patients. Another publication noted that whereas HIV infection may delay VDRL seropositivity, eventually the test does become reactive. Another noted that the term “dils” (dilutions) in a VDRL or RPR report refers to the titer by using the total volume of the diluted specimen (i.e., a titer of 1:8 may be reported as 8 dils).

  • Selection of Tests

    The selection of tests for syphilis is dictated by the clinical situation. If the patient has possible primary syphilis, a dark-field examination would be helpful, although it is becoming difficult to find laboratories with the necessary equipment and laboratorians with sufficient expertise. If the dark-field test result is negative or cannot be performed, an FTA-ABS test should be done. If the FTA-ABS result is nonreactive and clinical suspicion is strong, the physician, for practical purposes, has the option of treating the patient without a conclusive diagnosis or of repeating the FTA-ABS test in 2-3 weeks.

    If the patient has confirmed early syphilis, an RPR result should nevertheless be obtained. If the RPR result is reactive, the degree of reactivity should be titered, since a falling titer after treatment is evidence that treatment was effective.

    If the patient has possible, equivocal, or late syphilis, an FTA-ABS (or MHA) and STS test should be done. If the FTA-ABS (or MHA) test result is reactive (2+ or more), the diagnosis is probable syphilis. If the FTA-ABS (or MHA) test result is weakly reactive (borderline or 1+ reactive), the test should be repeated in 1 month. If it is still weakly reactive, the diagnosis is possible or probable syphilis, depending on the clinical picture. The RPR results are useful mainly as additional evidence in equivocal cases.

    If a routing screening RPR result is found to be positive in a person with no history or clinical evidence of syphilis, a confirmatory test should be done. If the confirmatory test result is negative, the patient should be screened for diseases known to cause a high incidence of BFP reactions. In this respect, a weakly positive RPR result may be due only to an acute BFP etiology, and the RPR result should be negative in 2-3 months. If the confirmatory test result is positive, past or present syphilis is a strong probability. Nevertheless, since even the confirmatory tests may occasionally produce a false positive (or false negative) reaction, in certain patients it may be necessary to repeat the confirmatory test.

  • Sensitivity and Specificity of Syphilis Tests

    Studies have been done in which duplicate samples from known syphilitic patients in various stages of their disease and also from normal persons were sent to various laboratories. Besides this, many reports have appeared from laboratories all over the world comparing one test with another in various clinical stages of syphilis, in nonsyphilitic diseases, and in apparently normal persons. These results are summarized in Table 15-3.

    Comparison of serologic tests for syphilis

    Table 15-3 Comparison of serologic tests for syphilis (approximate percentage reported reactive)*

    Table 15-3 demonstrates considerable variation in results. Several factors must be involved besides the inherent sensitivity and specificity of the individual tests themselves:

    1. Antibiotic treatment may cause some previously reactive syphilitic patients to become nonreactive.

    2. Some clinically normal persons may have unsuspected subclinical syphilis.

    3. True BFP reactions, either acute or chronic, must be taken into account.

    4. There is obvious variation in technique and ability between laboratories. Some laboratories introduce their own modifications into standard techniques.

    5. The time of appearance differs for the various antibodies. In general, the FTA-ABS test result becomes positive in significant numbers of patients in the middle or end of the primary stage, followed by the MHA and RPCF, and then the VDRL. All of these procedures usually give positive results in the secondary stage, and also probably in the early latent stage.

  • Specific Treponema Pallidum Tests

    Treponema pallidum immobilization (TPI) test. Syphilitic spirochetes can grow in rabbits. Nelson devised a Treponema pallidum immobilization (TPI) test in which syphilitic spirochetes are incubated with the patient’s serum. If specific antisyphilitic antibody is present, it will attack the spirochetes and immobilize them, causing them to stop moving when viewed under the microscope. This involves an antibody that is different from reagin and is specific against pathogenic Treponema spirochetes. Besides syphilis, other Treponema spirochetal diseases such as yaws may give positive reactions. The main disadvantages of this test are that it requires working with live spirochetes and use of an animal colony, is difficult to perform accurately, and is expensive. The TPI test has been replaced by the FTA-ABS and the microhemagglutination (MHA) tests (to be discussed later), and it is now very difficult, almost impossible, to obtain a TPI study in the United States.

    Reiter protein complement fixation (RPCF) test. The TPI test is done using the Nichol strain of pathogenic spirochetes. It was discovered that a certain nonpathogenic Treponema spirochete called the “Reiter strain” could be cultured more easily and cheaply on artificial media. Antigen prepared from this organism was adapted to a CF technique, and the result was the Reiter protein complement fixation (RPCF) test.

    The Reiter antibody is different from the Treponema-immobilizing antibody of the TPI test. Apparently, the nonpathologic Reiter and the pathologic Nichol spirochete share a common protein antigen, and it is this protein that is used in the RPCF test. In addition, the Nichol organism has a specific antigen that results in the immobilizing antibody response of the TPI assay. Several investigators have found the RPCF test to be almost as sensitive and specific as the TPI test, although others are less enthusiastic about its sensitivity in late syphilis. The Reiter antibody also appears at a different time than does the TPI antibody. The disadvantages of the RPCF are those inherent in all CF tests. The RPCF test is practically never used in the United States.

    Fluorescent treponemal antibody (FTA) test. In the FTA procedure, dead Nichol strain spirochetes are fixed to slides; these spirochete-containing slides can be prepared in the laboratory or obtained ready for use from commercial sources. The patient’s serum is placed on the slide with the spirochetes and allowed to incubate with the organisms. Antispirochete antibody in the patient serum coats the surface of the spirochetes. The serum is then washed off and replaced by antibodies against human gamma globulin that are tagged with a fluorescent dye. Since human antibodies against syphilis produced after the first month of infection are gamma globulins, the antibodies against human gamma globulin, with the fluorescein dye attached, will attack and adhere to any patient antitreponemal antibody that coats the spirochetes on the slide. The spirochetes will then appear fluorescent when viewed with an ultraviolet microscope.

    Unfortunately, fluorescent work is not as simple as this description or the recent literature would imply. Many technical problems remain. These tests at present are not suitable for mass screening, although they are less time consuming than the RPCF test and easier than the TPI test. Many substances give varying degrees of natural fluorescence, and it is sometimes difficult to decide whether a preparation is actually positive or not. There may be nonspecific antigen-antibody binding of the cross-reacting type, as well as specific reaction. When the animal anti-human globulin antibody is conjugated with fluorescein, not all of the fluorescein binds to it. Any remaining free fluorescein may nonspecifically stain various proteins, including the spirochetes, when the tagged mixture is added to the patient’s serum.

    Because of the problem of nonspecific fluorescence, the FTA underwent modification to become the FTA test with absorption (FTA-ABS). Reiter Treponema antigen is used to absorb nonspecific cross-reacting antibodies out of the patient’s serum. Antibody to T. pallidum is not absorbed out by this technique. The absorbed patient serum replaces nonabsorbed patient serum in the standard FTA procedure.

    The FTA-ABS is a well-established test. It has relatively good sensitivity in primary syphilis (except in very early disease) and is reported to be even more sensitive than the TPI in tertiary syphilis. If the patient is treated adequately in the primary or secondary stage, the FTA-ABS response will usually return to nonreactive state, but after the secondary stage it usually will not become nonreactive in spite of therapy. It is said to be at least as specific as the TPI, possibly even more so.

    Drawbacks. Weak reactions may cause interpretation problems. Official recommendations are that equivocal or 1+ reactive specimens should be repeated and the 1+ reclassified as borderline if the repeat test result is nonreactive and called 1+ reactive if reactivity remains 1+. This is important because some false positive and false negative results may occur in the FTA-ABS due to laboratory technical factors. Some studies have revealed a 5%-10% variance between laboratories. This is much more likely to happen in weakly (1+) reactive specimens than with specimens having reactivity graded 2+ to 4+.

    No laboratory test is free from the possibility of nonhuman error, and the FTA-ABS is no exception. Occasional false positive FTA-ABS results have been reported in persons with hyperglobulinemia due to macroglobulins and in patients with antinuclear antibodies. In addition, atypical fluorescent patterns (“beaded”) that could be misinterpreted as reactive have occurred in some patients with systemic lupus erythematosis. Cross-reaction may occur in other spirochetal diseases such as Borrelia (Lyme disease), leptospirosis, and relapsing fever. Problems of nonspecific fluorescence mentioned earlier have been reduced but not entirely eliminated. Occasional false negatives may occur even with the FTA-ABS test. Several reports suggest that concurrent infection by the human immunodeficiency virus type 1 (HIV-1) can sometimes delay the development of a positive VDRL or FTA-ABS reaction.

    Microhemagglutination (MHA-TP) test. An MHA test is available using formalin-treated RBCs coated with Nichol strain T. pallidum material. Patient serum is preabsorbed with Reiter Treponema reagent in the same manner as the FTA-ABS technique. Antibody to T. pallidum prevents normal RBC agglutination when the test is performed. About 1%-2% of sera contain nonspecific Forssman-type antibodies, so that reactive sera must be retested with nonsensitized control RBCs.

    The MHA test is not as sensitive in primary syphilis as the FTA-ABS test, although it is reactive in more than 50% of patients. It seems equally as sensitive as the FTA-ABS test in secondary and possibly in late syphilis. Compared to FTA-ABS results, various studies have shown 90%-98% overall correlation. Our laboratory has performed a comparison of this type and found that nearly 85% of the disagreements represented either nonreactive or 1+ reactive MHA results and nonreactive or 1+ FTA-ABS results. Therefore, most disagreements seem to occur at low reactivity levels in which the accuracy of either test result is open to some question. This being the case, there is reason to conclude that the MHA could be substituted for the FTA-ABS, except possibly in primary syphilis (in which case an FTA-ABS test could be done if the MHA test results were nonreactive). The MHA test is much easier to perform than the FTA-ABS test and is less expensive.

    More recently, several enzyme-linked immunosorbent assays (ELISA) have been reported, with preliminary results similar to those of the FTA-ABS. However, more independent evaluations are needed. Several research centers have also developed Western blot methods. Several investigators have published nucleic acid probe tests amplified by polymerase chain reaction (PCR). Again, initial very good results must be verified by others.