Davinder P.S. Dosanjh, DPhil; Timothy S.C. Hinks, MD; John A. Innes, MD; Jonathan J. Deeks, PhD; Geoffrey Pasvol, DPhil; Sarah Hackforth, RGN; Hansa Varia, RGN; Kerry A. Millington, DPhil; Rubamalar Gunatheesan, MD; Valerie Guyot-Revol, PhD; Ajit Lalvani, DM
Note: Drs. Dosanjh and Hinks contributed equally to this work.
Acknowledgment: The authors thank the study participants; Sarah Gooding, for collection and processing of some samples; and Muhunthan Thillai, for critical appraisal and revision of the final manuscript.
Grant Support: By the Wellcome Trust (Dr. Lalvani is a Wellcome Senior Research Fellow in Clinical Science), the Sir Halley Stewart Trust (Dr. Dosanjh's studentship), a Wellcome Trust PhD Prize Studentship (Dr. Millington), and a United Kingdom Department of Health Senior Fellowship in Evidence Synthesis (Dr. Deeks).
Potential Financial Conflicts of Interest:Consultancies: A. Lalvani (Oxford Immunotec Ltd. [nonexecutive director from 2003 to 2007]). Honoraria: G. Pasvol (Transactions of the Royal Society of Tropical Medicine). Stock ownership or options (other than mutual funds): D.P.S. Dosanjh (Oxford Immunotec Ltd.), A Lalvani (Oxford Immunotec Ltd.), University of Oxford (Oxford Immunotech Ltd.). Patents received: A. Lalvani (T-cell–based diagnosis of tuberculous infection), University of Oxford (T-cell–based diagnosis of tuberculous infection). Patents pending: D.P.S. Dosanjh (T-cell–based diagnosis of tuberculous infection), A. Lalvani (T-cell–based diagnosis of tuberculous infection, University of Oxford (T-cell–based diagnosis of tuberculous infection).
Reproducible Research Statement: The analytic data set of all participants' diagnoses, tuberculin skin test results, ELISpot results, and ELISpotPLUS results can be made available to academic investigators on request by written agreement.
Requests for Single Reprints: Ajit Lalvani, DM, Tuberculosis Immunology Group, Department of Respiratory Medicine, Faculty of Medicine, Imperial College London, St. Mary's Campus, Norfolk Place, London W2 1PG, United Kingdom; e-mail, a.lalvani@imperial.ac.uk.
Current Author Addresses: Dr. Dosanjh: University of Oxford Medical School, William Osler House, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
Dr. Hinks: Allergy, Inflammation, and Repair Group, Level D, Southampton General Hospital, Southampton SO16 6YD, United Kingdom.
Dr. Innes and Ms. Hackforth: Department of Infection & Tropical Medicine, Birmingham Heartlands Hospital, Bordsley Green East, Birmingham B9 5SS, United Kingdom.
Dr. Deeks: Department of Public Health and Epidemiology, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom.
Dr. Pasvol and Ms. Varia: Department of Infection and Tropical Medicine, Northwick Park Hospital, Imperial College London, Watford Road, Harrow HA1 3UJ, United Kingdom.
Drs. Millington and Lalvani: Department of Respiratory Medicine, Imperial College London, St. Mary's Campus, Norfolk Place, London W2 1PG, United Kingdom.
Dr. Gunatheesan: University of Melbourne Medical School, Melbourne, Australia.
Dr. Guyot-Revol: Genopoietic, 1390 rue Centrale-Beynost, 01708 Miribel Cedex, France.
Author Contributions: Conception and design: J.A. Innes, A. Lalvani. Analysis and interpretation of the data: D.P.S. Dosanjh, T.S.C. Hinks, J.J. Deeks, K.A. Millington, A. Lalvani.
Drafting of the article: D.P.S. Dosanjh, T.S.C. Hinks, G. Pasvol, A. Lalvani.
Critical revision of the article for important intellectual content: D.P.S. Dosanjh, T.S.C. Hinks, J.A. Innes, J.J. Deeks, G. Pasvol, K.A. Millington, A. Lalvani.
Final approval of the article: D.P.S. Dosanjh, J.J. Deeks, G. Pasvol, V. Guyot-Revol, A. Lalvani.
Provision of study materials or patients: J.A. Innes, G. Pasvol, S. Hackforth, H. Varia, A. Lalvani.
Statistical expertise: D.P.S. Dosanjh, J.J. Deeks.
Obtaining of funding: A Lalvani.
Administrative, technical, or logistic support: D.P.S. Dosanjh, G. Pasvol, S. Hackforth, K.A. Millington, A. Lalvani.
Collection and assembly of data: D.P.S. Dosanjh, T.S.C. Hinks, S. Hackforth, K.A. Millington, V. Guyot-Revol.
The role of new T-cell–based blood tests for tuberculosis in the diagnosis of active tuberculosis is unclear.
To compare the performance of 2 interferon-γ assays and tuberculin skin testing in adults with suspected tuberculosis.
Prospective study conducted in routine practice.
2 urban hospitals in the United Kingdom.
389 adults, predominantly of South Asian and black ethnicity, with moderate to high clinical suspicion of active tuberculosis.
Tuberculin skin testing, the enzyme-linked immunospot assay (ELISpot) incorporating early secretory antigenic target-6 and culture filtrate protein-10 (standard ELISpot), and ELISpot incorporating a novel antigen, Rv3879c (ELISpotPLUS) were performed during diagnostic assessment by independent persons who were blinded to results of the other test.
Sensitivity, specificity, predictive values, and likelihood ratios.
194 patients had a final diagnosis of active tuberculosis, of which 79% were culture-confirmed. Sensitivity for culture confirmed and highly probable tuberculosis was 89% (95% CI, 84% to 93%) with ELISpotPLUS, 85% (CI, 79% to 90%) with standard ELISpot, 79% (CI, 72% to 85%) with 15-mm threshold tuberculin skin testing, and 83% (CI, 77% to 89%) with stratified thresholds of 15 and 10 mm in vaccinated and unvaccinated patients, respectively. The ELISpotPLUS assay was more sensitive than tuberculin skin testing with 15-mm cutoff points (P = 0.01) but not with stratified cutoff points (P = 0.10). The ELISpotPLUS assay had 4% higher diagnostic sensitivity than standard ELISpot (P = 0.02). Combined sensitivity of ELISpotPLUS and tuberculin skin testing was 99% (CI, 95% to 100%), conferring a negative likelihood ratio of 0.02 (CI, 0 to 0.06) when both test results were negative.
Local standards for tuberculin skin testing differed from others used internationally. The study sample included few immunosuppressed patients.
The ELISpotPLUS assay is more sensitive than standard ELISpot and, when used in combination with tuberculin skin testing, enables rapid exclusion of active infection in patients with moderate to high pretest probability of tuberculosis.
Can new T-cell–based blood tests rule out active tuberculosis?
In this study involving 389 adults with moderate to high suspicion of active tuberculosis, the sensitivity of a new T-cell–based enzyme-linked immunoassay (ELISpotPLUS) was higher than that of 15-mm threshold tuberculin skin testing (89% vs. 79%) and similar to that of 10-mm threshold skin testing (83%) in unvaccinated patients for identifying culture-confirmed and highly probable tuberculosis. Patients with negative results on both ELISpotPLUS and skin tests had a very low likelihood of tuberculosis (negative likelihood ratio, 0.02).
The ELISpotPLUS assay, particularly when used in combination with tuberculin skin testing, can help exclude a diagnosis of active tuberculosis.
Table 1. Categorization of the Study Population
Table 2. Participant Characteristics
Study flow diagram.
ELISpot = enzyme-linked immunospot assay incorporating early secretory antigenic target-6 and culture filtrate protein-10; ELISpotPLUS = enzyme-linked immunospot assay incorporating early secretory antigenic target-6, culture filtrate protein-10, and Rv3879c; TB = tuberculosis; TST= tuberculin skin testing. *Results were indeterminate because of no achievement of positive control (11 patients), high background (1 patient), peptide contamination (1 patient), inconclusive assay (1 patient), or defective ELISpot plate (1 patient). †Results were not available because of history of TB (clinically contraindicated) (45 patients), patient did not return for reading (8 patients), result not recorded (8 patients), reason unknown (3 patients), death (1 patient), test performed elsewhere (1 patient), or patient declined consent (1 patient). ‡Tuberculin skin test results were based on a 15-mm cutoff point and considered positive if induration was ≥15 mm on the Mantoux test or grade 3 to 4 on the Heaf test regardless of bacille Calmette–Guérin vaccination status.
Study flow diagram, stratified by final diagnosis and then by test result.
These are the same data shown in Figure 1 but displayed in a different manner to allow scrutiny of the entire raw data set. ELISpot = enzyme-linked immunospot assay incorporating early secretory antigenic target-6 and culture filtrate protein-10; ELISpotPLUS = enzyme-linked immunospot assay incorporating early secretory antigenic target-6, culture filtrate protein-10, and Rv3879c; TB = tuberculosis; TST= tuberculin skin testing. *Results were not available because of history of TB (clinically contraindicated) (45 patients), patient did not return for reading (8 patients), result not recorded (8 patients), reason unknown (3 patients), death (1 patient), test performed elsewhere (1 patient), or patient declined consent (1 patient). †Tuberculin skin test results were based on a 15-mm cutoff point and considered positive if induration was ≥15 mm on the Mantoux test or grade 3 to 4 on the Heaf test regardless of bacille Calmette–Guérin vaccination status. ‡Results were indeterminate because of no achievement of positive control (11 patients), high background (1 patient), peptide contamination (1 patient), inconclusive assay (1 patient), or defective ELISpot plate (1 patient).
Table 3. Final Diagnoses of Study Participants
Table 4. Accuracy for the Diagnosis of Active Tuberculosis
Likelihood ratios, sensitivities, and specificities of tests used in combination, using ELISpot or ELISpotPLUS first.
Data are for patients in whom results on both tests were available (n = 265). Except where stated, values are likelihood ratios with 95% CIs. Tuberculin skin test thresholds for positivity were induration ≥15 mm on the Mantoux test or grade 3 to 4 on the Heaf test. ELISpot = enzyme-linked immunospot incorporating early secretory antigenic target-6 and culture filtrate protein-10; ELISpotPLUS = enzyme-linked immunospot incorporating early secretory antigenic target-6, culture filtrate protein-10, and Rv3879c; TB = tuberculosis; TST = tuberculin skin testing. Top. ELISpot followed by TST. *Combined sensitivity of 1 or more positive results from tests used in combination, 97% (CI, 93% to 99%). †Combined specificity for a double-negative result from tests used in combination, 62% (CI, 53% to 71%). Bottom. ELISpotPLUS followed by TST. ‡Combined sensitivity of 1 or more positive results from tests used in combination, 99% (CI, 95% to 100%). §Combined specificity for a double-negative result from tests used in combination, 61% (CI, 52% to 70%).
Likelihood ratios, sensitivities, and specificities of tests used in combination for diagnostic evaluation, using tuberculin skin testing (TST) as the first test.
Data are for patients in whom results on both tests were available (n = 265). Except where stated, numbers are likelihood ratios with 95% CIs. Tuberculin skin test thresholds for positivity were induration ≥15 mm on the Mantoux test or grade 3 to 4 on the Heaf test. ELISpot = enzyme-linked immunospot incorporating early secretory antigenic target-6 and culture filtrate protein-10; ELISpotPLUS = enzyme-linked immunospot incorporating early secretory antigenic target-6, culture filtrate protein-10, and Rv3879c; TB = tuberculosis. Top. TST followed by ELISpot. *Combined sensitivity of 1 or more positive results from tests used in combination, 97% (CI, 93% to 99%). †Combined specificity for a double-negative result from tests used in combination, 62% (CI, 53% to 71%). Bottom. TST followed by ELISpotPLUS. ‡Combined sensitivity of 1 or more positive results from tests used in combination, 99% (CI, 95% to 100%). §Combined specificity for a double-negative result from tests used in combination, 61% (CI, 52% to 70%).
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Liu Hong
Institute of Digestive Diseases, Xijing Hospital, Fourth Military Medical University
March 15, 2008
The application strategy of tuberculin skin testing, ELISpotPLUS and ELISpot
To the editor:
We read with interest the article by Dosanjh and colleagues (1). They clearly show that ELISpotPLUS is a sensitive and clinically useful diagnostic test for evaluation of patients with suspected tuberculosis. However, they don¡¯t show the cost effectiveness of ELISpotPLUS versus tuberculin skin testing and ELISpot. Clinicians will need to know which assay is recommended for wide clinical application. Could the authors comment on the application strategy of the three assays, including separate application and combined application?
This study is based on the participants predominantly in South Asian and black ethnicity, so the results may not be applicable for all patients with suspected tuberculosis. Will incorporation of Rv3879c improve sensitivity over standard ELISpot for all patients? Could the authors comment on the potential effect of epigenetic inheritance on the roles of Rv3879c, which is regulated by a special molecular network? So it is indispensable to ask whether the molecular network involve other important antigens for the clinical diagnostic evaluation of suspected tuberculosis.
References:
1. Dosanjh DP, Hinks TS, Innes JA, et al. Improved Diagnostic Evaluation of Suspected Tuberculosis. Ann Intern Med. 2008 Mar 4;148(5):325-36.
None declared
David M. Brett-Major
Division of Infectious Diseases, National Naval Medical Center, Bethesda MD
March 17, 2008
No Title
No Comment
Ivan Landires
Institut Pasteur, UP Pathogénomique Mycobacterienne Integrée, Paris, France
April 2, 2008
Can IFN Gamma Release Assays Differentiate Between Active Tuberculosis and Sarcoidosis ?
Dear Editor,
We have read with great interest the recent article by Dosanjh and colleagues (1) who found that the ELISpotPLUS assay was a clinically useful diagnostic test for evaluation of patients with suspected tuberculosis. This assay uses an additional region of difference 1"“encoded antigen, Rv3879c, alongside early secretory antigenic target-6 (ESAT-6) and culture filtrate protein-10, which are all part of the recently identified, novel ESX-1 secretion system of Mycobacterium tuberculosis (2).
As mentioned in the editorial (3), the use of these tests for the diagnosis of active tuberculosis is based on the logic that one must have tuberculous infection in order to have tuberculosis disease. But, is the ELISpotPLUS assay able to establish a differential diagnosis between active tuberculosis and sarcoidosis? M. tuberculosis has been proposed as a candidate trigger for the development of sarcoidosis and several mycobacterial antigens have been recovered from sarcoidal tissue (4). Recent studies have detected Th-1 immune responses to M. tuberculosis ESAT -6 using the ELISpot assay in sarcoidosis patients (5). A diagnosis of sarcoidosis is firm when chest radiographic evidence is accompanied by compatible biopsy and characteristic clinical features, with all other causes of granulomas ruled out (4). For that reason it will be important to evaluate ELISpotPLUS responses to ESX-1 peptides in sarcoidosis patients and compare these findings with ELIspotPLUS results from patients with active tuberculosis. Overall, we suggest that this issue merits further investigation.
References.
1. Dosanjh DP, Hinks TS, Innes JA, Deeks JJ, Pasvol G, Hackforth S, et al. Improved diagnostic evaluation of suspected tuberculosis. Ann Intern Med. 2008; 148: 325-36.
2. Brodin P, Rosenkrands I, Andersen P, Cole ST, and Brosch R. ESAT-6 proteins: protective antigens and virulence factors? Trends Microbiol. 2004; 12:500-508.
3. Menzies D. Using tests for latent tuberculous infection to diagnose active tuberculosis: can we eat our cake and have it too? Ann Intern Med. 2008; 148: 325-36.
4. Iannuzzi MC, Rybicki BA, Teirstein AS. Sarcoidosis. N Engl J Med. 2007; 357: 2153-65.
5. Drake WP, Dhason MS, Nadaf M, Shepherd BE, Vadivelu S, Hajizadeh R, et al. Cellular recognition of Mycobacterium tuberculosis ESAT-6 and KatG peptides in systemic sarcoidosis. Infect Immun. 2007; 75: 527-30.
Dosanjh DP, Hinks TS, Innes JA, et al. Improved Diagnostic Evaluation of Suspected Tuberculosis. Ann Intern Med. 2008;148:325–336. doi: https://doi.org/10.7326/0003-4819-148-5-200803040-00003
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© 2019
Published: Ann Intern Med. 2008;148(5):325-336.
DOI: 10.7326/0003-4819-148-5-200803040-00003
Infectious Disease, Mycobacterial Infections, Prevention/Screening.
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