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Diagnosing Latent Tuberculosis Infection

Turning Glitter to Gold

University of Texas Health Center Tyler, Texas

Elimination of tuberculosis in industrialized nations hinges on diagnosis and treatment of latent tuberculosis infection to prevent disease. The tuberculin skin test is used to diagnose latent tuberculosis infection, but it requires two visits and skilled personnel for test placement and interpretation. In addition, the tuberculin skin test does not reliably separate latent tuberculosis infection from prior immunization with Mycobacterium bovis bacille Calmette Guerin (BCG) or infection with environmental mycobacteria. A major scientific advance has been the identification of early secreted antigenic target 6 kD protein (ESAT-6) and culture filtrate protein 10 (CFP10), which are not present in BCG or in most nontuberculous mycobacteria (1, 2). These proteins are encoded within region of difference 1 (RD1) of the M. tuberculosis genome. In persons with latent tuberculosis infection, memory T cells produce interferon- in response to M. tuberculosis antigens. RD1 interferon-–based tests involve stimulating blood lymphocytes with ESAT-6 or CFP10, followed by measurement of interferon- by enzyme-linked immunoassay or detection of interferon-–producing cells by enzyme-linked immunospot assay.

In this issue of the Journal (pp. 59–64), Mori and colleagues provide the most convincing evidence to date that RD1 interferon-–based tests are substantially more specific than the tuberculin skin test in BCG-vaccinated persons without prior exposure to tuberculosis (3). Only 2% of 216 such persons had positive RD1 interferon-–based tests, whereas 65% of 113 individuals had positive tuberculin skin tests. Also in this issue (pp. 65–69), Brock and coworkers show that, in BCG-vaccinated persons, an RD1 interferon-–based test was positive in 4 (50%) of 8 close contacts of a tuberculosis patient, compared with only 2 (6%) of 32 without such contact (4).

The most critical issue in the further evaluation of RD1 interferon-––based tests is to determine their sensitivity for detection of latent tuberculosis infection. Mori and Lalvani found that the RD1 interferon-–based tests have equal or better sensitivity than the tuberculin skin test in patients with tuberculosis (3, 5). Because interferon- production and tuberculin skin test reactivity are reduced in tuberculosis, however, these findings cannot be extrapolated to persons with latent tuberculosis infection. In a tuberculosis outbreak, RD1 interferon-–based tests in healthy contacts correlated significantly better than tuberculin skin tests with extent of exposure, suggesting that the former are more sensitive for detection of latent tuberculosis infection (6). The sensitivity of the tuberculin skin test in this study, however, may have been underestimated, as tuberculin reactivity was assessed by the Heaf test, which is inferior to the Mantoux method.

Estimates of sensitivity require comparison with a gold standard, and the lack of such a standard for latent tuberculosis infection has been lamented by many authors. Nevertheless, the tuberculosis research community should recall the motto of the United States Army Corps of Engineers: "The difficult we do at once, the impossible takes a little longer." Persons who almost certainly have latent tuberculosis infection are those without prior BCG vaccination who convert their tuberculin skin test after recent exposure to a patient with infectious tuberculosis. Multicenter studies to estimate the sensitivity of RD1 interferon-–based tests in this population are of the utmost importance. However, these studies cannot determine if the RD1 interferon-–based tests are more sensitive than the tuberculin skin test. To address this issue, recent contacts of patients with tuberculosis with positive RD1 interferon-–based tests but negative tuberculin skin tests must be evaluated. These individuals may have latent tuberculosis infection or they may have transient infection or antigen exposure that resolves. Distinguishing these possibilities requires prolonged follow-up without therapy, preferably in conjunction with a parallel tuberculin-positive cohort, to identify patients that develop tuberculosis. Such a study is ethically permissible only in locations where latent tuberculosis infection is not routinely treated.

Once the sensitivities of the RD1 interferon-–based tests for latent tuberculosis infection have been established, the second priority is to determine the effect of environmental mycobacterial infection on these tests. Nontuberculous mycobacterial exposure is believed to be minimal in northern Europe, and Brock and colleagues (4) show that, among BCG-unvaccinated persons in this region, the correlation between tuberculin skin testing results and RD1 interferon-–based testing is extremely high. In regions such as the southeastern United States, however, infection with M. avium complex and M. kansasii is common and can yield false-positive tuberculin skin tests. Although small studies suggest that persons with M. avium complex pulmonary disease have negative RD1 interferon-–based tests (7), results in healthy persons with M. avium complex infection are unknown. ESAT-6 and CFP10 are present in M. kansasii and M. marinum, and some persons infected with these organisms have positive RD1 interferon-–based tests (8). Although M. avium and most nontuberculous mycobacteria tested do not have the ESAT-6 and CFP10 genes (1, 2), related genes are present in some of these organisms (9) and could cause cross-reactivity to RD1 antigens. Furthermore, because the frequency of infection with different species of environmental mycobacteria is largely unknown, it will be important to compare the specificity of RD1 interferon-–based tests and the tuberculin skin test in persons infected with nontuberculous mycobacteria but with extremely low-risk for latent tuberculosis infection. It may be possible to identify nontuberculous mycobacterial infection by immunologic reactivity to the relevant antigen preparations or by skin testing, as has been done for M. avium complex (7, 10).

Definitive evaluation of the sensitivities and specificities of the RD1 interferon-–based tests will require long-term studies. What should clinicians do in the interim? The QuantiferonTB-Gold test (Cellestis Ltd., Carnegie, Victoria, Australia) is being evaluated by the Food and Drug Administration (Dr. James Rothel, personal communication). This test uses the enzyme-linked immunoassay to detect interferon- induced by ESAT-6 and CFP10, and was used in Mori's study (3). If approved, I would favor its widespread use for diagnosis of latent tuberculosis infection. Compared with the tuberculin skin test, the Quantiferon-TB-Gold test is simple to perform, has greater specificity in BCG-vaccinated persons, and has at least equal sensitivity in patients with tuberculosis, given the caveats noted above.

To maximize the diagnostic value of RDI interferon-–based tests, many unanswered questions remain. What are the performance characteristics of the enzyme-linked immunoassay or enzyme-linked immunospot formats? Should the antigens be recombinant proteins or pools of peptides? What is the optimal time for testing after exposure to a tuberculosis patient? These and other questions are important, but determination of test sensitivity is the central issue that must be answered to turn the glitter of these new tests to gold.

FOOTNOTES

Conflict of Interest Statement: P.F.B. does not have a financial relationship with a commercial entity that has an interest in the subject of this manuscript.

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