Tuberculosis Assay Research Articles

Comparison of an In-house and a Commercial RD1-based ELISPOT-IFN- Assay for the Diagnosis of Mycobacterium tuberculosis Infection

To compare a RD1-based in-house ELISPOT-interferon-gamma (IFN-gamma) assay with a commercial (T-SPOT.TB) assay for the diagnosis of Mycobacterium tuberculosis (TB) infection and the efficacy of the tuberculin skin test (TST) and ELISPOT assay in detecting latent TB infection (LTBI). Eighty-six subjects (65 household contacts of contagious TB-infected patients, 13 subjects with active or previous TB infection, and 8 with suspected TB infection) were consecutively recruited in the context of a surveillance program. Enrolled subjects underwent the Mantoux TST and two different ELISPOT-IFN-gamma assays: an in-house assay using a pool of selected M. tuberculosis peptides (MTP) and the commercial T-SPOT.TB assay. The in-house and commercial ELISPOT-IFN-gamma assays showed almost complete concordance (99%) in diagnosing acute or LTBI. When comparing the efficacy of the TST with the in-house ELISPOT assay in detecting TB infection, a small agreement was observed (k=0.344, P<0.0001): 36% of the subjects with a positive TST were ELISPOT-MTP negative and 12% with a negative TST were ELISPOT-MTP positive. Furthermore, 78% of the ELISPOT-MTP negative individuals were ELISPOT- Bacillus Calmette-Guérin (BCG) positive, most of whom had received BCG vaccination. Our in-house ELISPOT assay based on a restricted pool of highly selected peptides is equivalent to the commercial T-SPOT.TB assay, is cheaper and is probably not confounded, unlike the TST, by BCG vaccination in our setting.

The specificity of interferon-γ-based blood tests in the identification of latent tuberculosis infection

To the Editors: I have read with interest several papers describing the use of cellular interferon (IFN)-γ-based blood tests, including the editorial by Davies and Drobniewski 1. Whilst reviewing the study by Lee et al. 2, Davies and Drobniewski 1 indicate that the T-SPOT. TB assay has higher sensitivity than QuantiFERON-TB Gold, but that the QuantiFERON-TB Gold may have better specificity. I do not agree with this tentative conclusion as the study by Lee et al. 2 does not report the true specificity of the two tests, as the control group of 15- and 16-yr-old students used to examine the specificity of the assays, although healthy, had a defined risk of existing tuberculosis (TB) infection. This was acknowledged by Lee et …

Cutaneous tuberculosis: A clinico-morphological study

Background: Cutaneous tuberculosis forms a small subset of extrapulmonary tuberculosis. The present study is an attempt to observe the clinico morphological pattern seen in cases of cutaneous tuberculosis over a period of 5 years, and to correlate them with mantoux reactivity and human immunodeficiency virus (HIV) status. Methods: All cases of cutaneous tuberculosis observed among the dermatology in patients and those attending out patient department were included in the study. The basis of diagnosis was clinical, histopathological and microbiological. Intradermal mantoux test and serological test in the form of enzyme-linked immunosorbent assay (ELISA) for tuberculosis was done. HIV screening was carried out in 32 cases. CD4 counts were done in all HIV positive cases. Results: A total 0.02% patient attending the dermatology centre had cutaneous tuberculosis. The spectrum of infection included 19 (51%) cases of lupus vulgaris, 7 (19%) cases of papulonecrotic tuberculids, six cases each of tuberculosis verrucosa cutis and scrofuloderma. One case had scrofuloderma and lupus vulgaris and another both scrofuloderma and papulonecrotic tuberculide. One case of lichen scrofulosorum was seen in a seven year old boy. 11 cases revealed evidence of systemic tuberculosis. Seven cases of HIV with CD4 counts between 50–500 cells/μl were observed in this study.

The use of interferon-γ-based blood tests for the detection of latent tuberculosis infection

As recently as 3 yrs ago, a colleague complained that after 100 yrs the only way to detect tuberculosis (TB) infection was still to measure “bumps on arms”. Now, at last, we have a genuine improved alternative: the ex vivo cellular interferon (IFN)-γ-based blood tests. The crucial questions we need to ask are how good they are at detecting infection with the TB bacterium and whether they are cost-effective? Unfortunately, determining the answers to these questions is not easy. The new ex vivo cellular IFN-γ assays (CIGAs) have been developed based on the release of IFN-γ from a patient’s T-cells when exposed to mycobacterial antigens. These novel assays rely on the principle that the genes encoding the secretory proteins early secretory antigenic target (ESAT)6/culture filtrate protein (CFP)10 are absent in the bacille Calmette–Guerin (BCG) vaccine strain and most environmental mycobacteria 1. Two commercial cellular immunological assays have been developed in which IFN-γ output from the patient’s whole blood or peripheral blood mononuclear cells are measured following treatment with ESAT6/CFP10 antigens 1. Different assay formats have been developed. One method relies on the relatively straightforward ELISA detection after a fixed volume of whole blood is incubated in a tube with ESAT6/CFP10 antigens (Quantiferon Gold; Cellestis Limited, Carnegie, Australia). The other method currently available commercially is the T-Spot.TB assay (Oxford Immunotec, Oxford, UK) based on the enzyme-linked immunospot (ELISPOT) principle. The methods appear to be both sensitive and specific. These assays have the potential to change the face of the diagnosis of latent TB infection (LTBI) and active TB at an early stage in the disease process. Their use is also likely to transform the screening of new migrants and other population groups. These tests require only a single blood sample without the repeat visit required for the tuberculin skin …

Detection of mycobacterial antigen responses in lung but not blood in HIV-tuberculosis co-infected subjects

Forty-seven HIV-infected adults had broncho-alveolar lavage stimulated with purified protein derivate of Mycobacterium tuberculosis. Eighteen of 19 (95%) with tuberculosis co-infection had interferon-gamma synthetic CD4 lymphocyte responses > 1% versus three of 28 (11%) without (P < 0.0001). Lung response was unrelated to blood CD4 cell count. BAL HIV tuberculosis responses were similar in 25 HIV-uninfected tuberculosis patients. Responses in matched blood samples were often undetectable. Therefore, immunological tuberculosis assays seem less affected by HIV co-infection when lung-based.

Coexistent acute pyogenic and tubercular petrous apicitis: a diagnostic dilemma

We report the case of a 16-year-old boy who presented to us with acute otitis media, facial weakness and retro-orbital pain. Computed tomography and magnetic resonance imaging (MRI) scans of the head and temporal bone revealed otitis media with petrous apicitis. The patient responded to broad-spectrum, parenteral antibiotics, with disappearance of facial weakness and reduction in pain. One month following the completion of treatment, the patient continued to have dull retro-orbital pain and developed ear discharge. A repeat MRI of the temporal bone revealed a persistent inflammatory lesion in the petrous apex, with a nodular, ring-enhancing lesion in the cerebellum, strongly suggestive of tuberculosis. The ear discharge stained positive for acid-fast bacilli and the patient's serum enzyme-linked immunosorbent assay for tuberculosis was reactive. The patient responded well to anti-tubercular treatment and was disease free eight months following the completion of treatment.

Interferon-γ production in vitro from whole blood of foot-and-mouth disease virus (FMDV) vaccinated and infected cattle after incubation with inactivated FMDV

Studies were performed to determine whether a rapid method to detect cell mediated immune responses to foot-and-mouth disease virus (FMDV) could be used either as a diagnostic test or provide a correlate of protection in animals post-vaccination. Using protocols based on the BOVIGAM assay for tuberculosis, whole blood samples from FMDV vaccinated or control animals, before and after challenge infection, were stimulated overnight with inactivated FMDV antigen. The quantity of interferon gamma (IFN-γ) produced in the supernatants was measured using an ELISA. Specific induction of IFN-γ was detected in samples from vaccinated, infected and vaccinated-then-infected cattle. Further development of this assay may provide a useful tool for the diagnosis of FMDV immune animals, including the identification of vaccinated animals that have been subsequently infected with FMDV. In these studies, combining the results of the IFN-γ assay with virus neutralising antibody titre, in groups of vaccinated animals, provided a correlation with the capacity to control virus replication after subsequent challenge.

A multiplexed and miniaturized serological tuberculosis assay identifies antigens that discriminate maximally between TB and non-TB sera

We have developed a multiplexed and miniaturized TB serological assay with the aim of identifying (combinations of) antigens that maximally discriminate between TB and non-TB patients. It features a microarray accommodating 54 TB antigens, less than 1 μl serum consumption and an indirect immunofluorescence detection protocol. With a panel of 20 TB and 80 non-TB sera we ranked combinations of TB antigens with respect to sensitivity and specificity of TB detection by means of logistic step-forward regression analysis. The highest-ranking TB antigen combination had an area-under-the-curve of the receiver–operator-characteristics (ROC) of 0.95. We also identified an antigen that on its own provided good specificity and sensitivity of TB detection (Ara 6–BSA; area-under-the-ROC curve: 0.90). These area-under-the-ROC curve values are exceptionally high for a serological TB assay. We conclude that TB antigen microarrays permit rapid identification of TB antigens that, either alone or in combination, discriminate maximally between TB and non-TB patients and that such identification provides an excellent starting point for developing point-of-care diagnostic assays.

High risk of Mycobacterium tuberculosis infection during the Hajj pilgrimage

Annually more than 2 million pilgrims from all over the world attend the Hajj in Saudi Arabia. Overcrowding during this pilgrimage leads to a high risk of transmission of airborne infectious diseases. Tuberculosis (TB) is common among hospitalized pilgrims, but the overall risk of acquiring Mycobacterium tuberculosis infection during this pilgrimage is not known. We conducted a prospective study to assess the risk of M. tuberculosis infection among Hajj pilgrims. We measured the immune response to TB antigens using a whole-blood assay (QuantiFERON TB assay) prior to departure and 3 months after return from the Hajj pilgrimage. Of 357 paired assays, 149 pilgrims were negative prior to the Hajj and 15 (10%) of these had a significant rise in immune response to TB antigens. Pilgrims may be at high risk of acquiring M. tuberculosis infection during the Hajj. This has significant public health implications for TB control in countries with large Muslim populations.

Effects of positive results for Mycobacterium avium subsp paratuberculosis as determined by microbial culture of feces or antibody ELISA on results of caudal fold tuberculin test and interferon-γ assay for tuberculosis in cattle

To determine whether cattle testing positive for Mycobacterium avium subsp paratuberculosis as determined by microbial culture of feces or antibody ELISA were more likely to have false-positive responses on the caudal fold tuberculin (CFT) test or interferon-gamma (IFN-gamma) assay for Mycobacterium bovis than cattle testing negative for M paratuberculosis. 1043 cattle from 10 herds in Michigan. Feces and blood samples for plasma were collected from cattle > or =24 months old on the day the CFT test was read. Fecal samples were submitted for microbial culture for M paratuberculosis. Plasma samples were tested for antibody against M paratuberculosis, and IFN-gamma after stimulation with purified protein derivative tuberculin from M bovis or M avium. Of 1043 cattle, 180 (17.3%) had positive CFT test results (suspects) and 8 (0.8%) had positive IFN-gamma assay results after stimulation with purified protein derivative tuberculin from M bovis. Forty-five (4.3%) and 115 (11.0%) cattle tested positive for M paratuberculosis as determined by microbial culture of feces and antibody ELISA, respectively. Cattle with positive responses for M paratuberculosis appeared to have an increased likelihood of false-positive results on the CFT test, although this association was not significant. No significant association was detected among cattle testing positive for M paratuberculosis as determined by microbial culture of feces and antibody ELISA and positive CFT test and IFN-gamma assay results for M bovis.

Susceptibilidad in vitro a los medicamentos anti-tuberculosos de aislados de cepas del complejo Mycobacterium tuberculosis obtenidos a partir de lobos marinos

Mycobacteria strains belonging to the Mycobacterium tuberculosis complex were isolated from seals found in the South Atlantic. The animals were received in Mundo Marino installations and treated for Mycobacterium tuberculosis complex by conventional therapy of intensive care and enriched food supply; however, in all cases treatment failed. Necropsies of all animals revealed extensive lesions compatible with tuberculosis involving lungs, liver, spleen and lymphatic nodes. Classical biochemical methods as well as molecular techniques using the IS6110 probes were performed for mycobacterial identification. Furthermore, the LCx M. tuberculosis assay (Abbott Laboratories) identified all strains as Mycobacterium tuberculosis complex members. The in vitro susceptibility pattern was examined in mycobacterial strains isolated from seven seals and in 3 reference strains--BCG, H37Rv (M. tuberculosis) and AN5 (Mycobacterium bovis)--to 4 medications--isoniazid, rifampin, streptomycin and ethambutol. Minimal inhibitory drug concentrations were determined by the Mycobacterial Growth Indicator Tube (BD Argentina) method and a microdilution and colorimetric assay using 3-(4-5 dimethyltiazol-2)-2,5 diphenyltetrazolium bromide. All the isolates and the reference strains BCG and AN5 were inhibited by MIC values similar to those of H37Rv with good agreement obtained by both techniques. These findings suggest that a therapeutic regimen aimed to seals diagnosed with tuberculosis play an important role in the prevention of tuberculosis transmission from infected animals to humans that are in routine contact with them.

Rapid dot sputum and serum assay in pulmonary tuberculosis

A rapid direct sputum (Sp.) and/or antibody assay, based on immunoblotting and enzyme immunoassay is described. The test can detect mycobacterial antigens or antibodies in clinical specimens from pulmonary tuberculosis (TB) patients. In this study, 87 sputa, 87 sera and 40 paired sputa and sera were utilized from smear-positive and smear-negative, culture-positive patients; 59 sputa, 37 sera and 22 paired sputa and sera from nontuberculosis respiratory disease patients and 68 sera from healthy controls. The antigen detection in sputum by dot assay has 86.1% sensitivity on active tuberculosis patients, 92.9% specificity, 91.6% positive predictive value (PPV), 88.2% negative predictive value (NPV) and 10.3% error. The antibody assay has 83.6% sensitivity, 95.4% specificity, 94.4% positive predictive value, 85.6% negative predictive value and 11% error. The test performed on paired sputum and serum (Sr.) samples has a sensitivity of 93.3%, which rose to 96.1% on smear-positive and culture-positive patients, but the specificity decreased to 83% in sputum, whereas in serum it was 92%. The results of the assay, combined with clinical and radiological data, could form the basis for starting an earlier course of treatment for tuberculosis.

Differentiation between intestinal tuberculosis and Crohn’s disease in endoscopic biopsy specimens by polymerase chain reaction

OBJECTIVES: It is difficult to differentiate intestinal tuberculosis from Crohn’s disease because of similar clinical, pathological, radiological, and endoscopic findings. The purpose of this study was to investigate the value of polymerase chain reaction (PCR) assay in the differentiation intestinal tuberculosis from Crohn’s disease, and compare the histopathological features of endoscopic biopsy of the two disorders. METHODS: A total of 39 endoscopic biopsy specimens from patients with intestinal tuberculosis and 30 specimens from patients with Crohn’s disease were subjected to pathological analysis retrospectively, Ziehl-Neelsen stain, and PCR assay. RESULTS: Except for granuloma with caseation and confluence, which was the characteristic of intestinal tuberculosis, other pathological features of intestinal tuberculosis and Crohn’s disease were very similar or were difficult to find in endoscopic biopsy specimens. The positivity rate by PCR in 39 intestinal tuberculosis specimens was 64.1% (25/39), but was zero by PCR in 30 Crohn’s disease specimens. Moreover, in the tissues of intestinal tuberculosis with granulomas similar to those of Crohn’s disease, there were 71.4% (10/14) positive by PCR, and there were 61.1% (11/18) positive in intestinal tuberculosis tissues without granulomas. CONCLUSIONS: Biopsy is of limited diagnostic value in the differentiation intestinal tuberculosis from Crohn’s disease, and PCR is valuable in the differentiation between intestinal tuberculosis and Crohn’s disease.

Use of recombinant proteins in antibody tests for bovine tuberculosis

Tuberculosis (TB) in cattle remains a major zoonotic and economic problem in many countries. Since the standard diagnostic assay, the intradermal test (IDT) with bovine PPD tuberculin, has less than optimal accuracy in all situations, other diagnostic methods such as serological assays have been investigated. Because of fundamental concerns for the low sensitivity and specificity of previous ELISA protocols, a profiling ELISA with nine purified, recombinant proteins of TB complex mycobacteria, was employed on samples from four groups of cattle: (a) naturally Mycobacterium avium-exposed and experimentally Mycobacterium bovis-infected, (b) officially-certified TB-free herds, (c) exposed to M. bovis in two field TB outbreaks and scored as bovine reactors in the γ-IFN assay for bovine TB, (d) paratuberculosis (para TB)-infected. The described ELISA proved to be highly specific. In fact, the antibody (Ab) response could be consistently detected in 3 out of 3 endotracheally-infected calves and in 1 out of 3 contact-infected calves. There was also a very low prevalence of low-titered, non-specific Ab responses in paraTB-infected animals. As for the animals exposed to field TB outbreaks, 16 out of 28 γ-IFN positive cattle were also Ab-positive; importantly, 7 out of 12 γ-IFN positive, IDT-negative cattle showed Ab responses to TB proteins. In general, the profile of the Ab response varied among animals; the reaction to single recombinant antigens was sometimes transient and fluctuating, whereas the panel of antigens on the whole was indeed more effective in Ab detection.

LCx Mycobacterium tuberculosis assay is valuable with respiratory specimens, but provides little help in the diagnosis of extrapulmonary tuberculosis

BACKGROUND. Commercial nucleic acid amplification tests, designed for the detection of Mycobacterium tuberculosis DNA/RNA in respiratory samples, are often applied also in nonrespiratory specimens in order to verify the diagnosis of extrapulmonary tuberculosis.AIM. To evaluate the value of the Abbott LCx Mycobacterium tuberculosis assay for the diagnosis of pulmonary and extrapulmonary tuberculosis based on routine clinical laboratory results.METHODS. The assay was used to analyse 350 respiratory and 826 nonrespiratory specimens from 961 patients, of whom 3.6% had culture-proven tuberculosis. The results obtained by the LCx assay were compared with the records on mycobacterial isolates of the national reference laboratory and, in the case of positive findings, with clinical data.RESULTS. In comparison with culture, the sensitivity, specificity and positive/negative predictive value of the assay on respiratory specimens were 87.5%, 99.7%, 93.3% and 99.4%, respectively. With nonrespiratory specimens, the overall sensitivity, specificity and positive/negative predictive value of the LCx assay were 73.3%, 98.0%, 40.7% and 99.5%, respectively. When clinical and histological data were also included, the positive predictive value of LCx with nonrespiratory specimens was 45.8%.CONCLUSION. Critical interpretation of the nucleic acid amplification results obtained from nonrespiratory specimens is necessary in both laboratory and clinical settings.

The rapid diagnosis of smear‐negative pulmonary tuberculosis: A cost‐effectiveness analysis

The prompt diagnosis of smear-negative pulmonary tuberculosis (PTB) is a clinical challenge. It may be achieved by a number of tests which have varying accuracies, costs and degrees of invasiveness. The objective of this study was to compare the cost-effectiveness of clinical judgement (empirical), the Roche Cobas amplicor assay for Mycobacterium tuberculosis (amplicor), acid-fast staining of bronchoalveolar lavage specimens (BAL), nucleic acid amplification tests of bronchoalveloar lavage specimens for M. tuberculosis (BAL + NAA), computed tomography (CT) and amplicor assay followed by BAL. The range of predictive values of the various strategies were derived from published data and a new study of 441 consecutive adult patients with suspected smear-negative PTB prospectively stratified into three pretest risk groups: low, intermediate and high. The cost-effectiveness was evaluated with a decision tree model (DATA software). The incidence of PTB was 5.7% (4% culture positive) for the whole group, 95% in the high-risk group, 0.9% in the low-risk group and 3.4% in the intermediate-risk group. The sensitivity of the empirical approach was 49% and of the amplicor assay was 44%. Patient outcomes were expressed as life expectancy for the base case of a 58-year-old man with a pretest probability of 5.7%. At this low pretest risk the differences in life expectancies between tests was < 0.1 years and the empirical approach incurred the lowest cost. Sensitivity analysis at increasing pretest risks showed better life expectancies (approximately 1 years) for CT scan and test combinations than empirical and amplicor for additional costs of US$243-US$309. Bronchoalveolar lavage had the worst overall cost-effectiveness. We conclude that the pretest risk of active PTB was a key determinant of test utility; that the AMPLICOR assay was comparable to clinical judgement; that BAL was the least useful test; and that with increasing risks, CT scan and test combinations performed better. Further studies are needed to better define patients with intermediate risk for PTB and to directly compare the cost-effectiveness of more sensitive nucleic acid amplification tests such as the enhanced Gen Probe, CT scan and test combinations/sequences in these patients.

The rapid diagnosis of smear-negative pulmonary tuberculosis: A cost-effectiveness analysis

Objective The prompt diagnosis of smear-negative pulmonary tuberculosis (PTB) is a clinical challenge. It may be achieved by a number of tests which have varying accuracies, costs and degrees of invasiveness. The objective of this study was to compare the cost-effectiveness of clinical judgement (empirical), the Roche Cobas amplicor assay for Mycobacterium tuberculosis (amplicor), acid-fast staining of bronchoalveolar lavage specimens (BAL), nucleic acid amplification tests of bronchoalveloar lavage specimens for M. tuberculosis (BAL + NAA), computed tomography (CT) and amplicor assay followed by BAL. Methodology The range of predictive values of the various strategies were derived from published data and a new study of 441 consecutive adult patients with suspected smear-negative PTB prospectively stratified into three pretest risk groups: low, intermediate and high. The cost-effectiveness was evaluated with a decision tree model (DATATM software). Results The incidence of PTB was 5.7% (4% culture positive) for the whole group, 95% in the high-risk group, 0.9% in the low-risk group and 3.4% in the intermediate-risk group. The sensitivity of the empirical approach was 49% and of the amplicor assay was 44%. Patient outcomes were expressed as life expectancy for the base case of a 58-year-old man with a pretest probability of 5.7%. At this low pretest risk the differences in life expectancies between tests was < 0.1 years and the empirical approach incurred the lowest cost. Sensitivity analysis at increasing pretest risks showed better life expectancies (~1 years) for CT scan and test combinations than empirical and amplicor for additional costs of US$243–US$309. Bronchoalveolar lavage had the worst overall cost-effectiveness. Conclusions We conclude that the pretest risk of active PTB was a key determinant of test utility; that the AMPLICOR assay was comparable to clinical judgement; that BAL was the least useful test; and that with increasing risks, CT scan and test combinations performed better. Further studies are needed to better define patients with intermediate risk for PTB and to directly compare the cost-effectiveness of more sensitive nucleic acid amplification tests such as the enhanced Gen Probe, CT scan and test combinations/sequences in these patients.

The ligase chain reaction as a primary screening tool for the detection of culture positive tuberculosis

BACKGROUNDThe ligase chain reaction Mycobacterium tuberculosis assay uses ligase chain reaction technology to detect tuberculous DNA sequences in clinical specimens. A study was undertaken to determine its sensitivity and specificity...

Relationship Between Estimated Pretest Probability and Accuracy of Automated Mycobacterium tuberculosis Assay in Smear-Negative Pulmonary Tuberculosis

The AMPLICOR assay (Roche; Branchburg, NJ), a rapid direct amplification test for Mycobacterium tuberculosis, has only been licensed for use in smear-positive respiratory specimens. However, many patients with pulmonary tuberculosis (PTB) have smear-negative disease. The clinical utility of this test in patients with smear-negative PTB is unknown. To evaluate the effect of pretest probability of PTB estimated by chest physicians on the accuracy of the AMPLICOR assay in patients with smear-negative PTB. A prospective study of consecutive patients suspected of having smear-negative PTB. Two chest physicians estimated the pretest probability of active disease (high, intermediate, and low categories). Respiratory specimens were examined with radiometric broth medium cultures and with the AMPLICOR assay for M tuberculosis. The decision on a final diagnosis of PTB was blinded to the AMPLICOR results. Active PTB was diagnosed in 25 of 441 patients (5.7%). The AMPLICOR assay had an overall sensitivity of 44% and a specificity of 99%. Results of the assay were negative in seven patients with culture-negative PTB. The proportions of patients in the high, intermediate, and low pretest groups were 4.5%, 19.7%, and 75.7%, respectively. The incidence of PTB for each group was 95%, 3.4%, and 0.9%, respectively. The sensitivities of the AMPLICOR assay in the three groups of patients were 47%, 33%, and 33%, respectively, while the specificities were 100%, 98%, and 99%, respectively. In patients suspected of having smear-negative PTB, the following conclusions were drawn: (1) the incidence of active PTB was low; (2) pretest estimates accurately discriminated between patients with high and low risk of PTB; (3) the risk of PTB was overestimated in the intermediate group; and (4) the utility of the AMPLICOR assay in the intermediate-risk group may be limited by the overestimation of disease prevalence and low test sensitivity. Further studies are needed on the role of the AMPLICOR assay in better selected patients with an intermediate risk of having smear-negative PTB.

Detection of Mycobacterium tuberculosis in Paraffin-Embedded Pleural Biopsy Specimens by Commercial Ribosomal RNA and DNA Amplification Kits

To evaluate the utility of two gene amplification systems in historical paraffin-embedded pleural biopsy (PEB) tissues from patients with pleural tuberculosis, and to compare the results to those obtained with conventional histologic and microbiological methods. A retrospective study. Seventy-four formalin-fixed PEB tissues collected and stored over 12 years (1984 through 1995) were retrieved. Gene amplifications were performed in 57 tissues from patients with diagnoses of pleural tuberculosis and in 17 from patients with carcinoma as controls, using the first version of the Amplified Mycobacterium tuberculosis Direct Test (AMTDT; Gen-Probe; San Diego, CA) and the LCx Mycobacterium tuberculosis Assay (LCxMTB; Abbott Laboratories; Abbott Park, IL). The sensitivities of the AMTDT and LCxMTB were 52.6% and 63.2%, respectively (p = not statistically significant). The specificity of both tests was 100%. Twenty tissue samples (35.1%) were positive by both systems, and 10 tissues (17.5%) were positive only by the AMTDT, while 16 tissues (28.1%) were positive only by the LCxMTB. Both tests gave negative results for 11 specimens (19.3%). When both tests were used, a positive diagnosis was achieved in 80.7% of the samples. Diagnosis of 73.7% of patient conditions had previously been made by smear examination of pleural biopsy and sputum, pleural liquid, or biopsy culture. The overall diagnostic yield with both culture and amplification techniques was 96.5% (55 of 57 patients) for pleural tuberculosis, with amplification techniques adding 22.8% of the diagnoses. Amplification techniques are useful in archival PEB tissues, providing additional diagnoses beyond culturing, although the sensitivity should be improved, possibly by standardizing protocols.