Long-Incubation-Time Gamma Interferon Release Assays in Response to Purified Protein Derivative, ESAT-6, and/or CFP-10 for the Diagnosis of Mycobacterium tuberculosis Infection in Children

ABSTRACTThe diagnosis of childhood active tuberculosis (aTB) and latentMycobacterium tuberculosis(M. tuberculosis) infection (LTBI) remains a challenge, and the replacement of tuberculin skin tests (TST) with commercialized gamma interferon (IFN-γ) release assays (IGRA) is not currently recommended. Two hundred sixty-six children between 1 month and 15 years of age, 214 of whom were at risk of recentM. tuberculosisinfection and 51 who were included as controls, were prospectively enrolled in our study. According to the results of a clinical evaluation, TST, chest X ray, and microbiological assessment, each children was classified as noninfected, having LTBI, or having aTB. Long-incubation-time purified protein derivative (PPD), ESAT-6, and CFP-10 IGRA were performed and evaluated for their accuracy in correctly classifying the children. Whereas both TST and PPD IGRA were suboptimal for detecting aTB, combining the CFP-10 IGRA with a TST or with a PPD IGRA allowed us to detect all the children with aTB with a specificity of 96% for children who were positive for the CFP-10 IGRA. Moreover, the combination of the CFP-10 IGRA and PPD IGRA detected 96% of children who were eventually classified as having LTBI, but a strong IFN-γ response to CFP-10 (defined as >500 pg/ml) was highly suggestive of aTB, at least among the children who were <3 years old. The use of long-incubation-time CFP-10 IGRA and PPD IGRA should help clinicians to quickly identify aTB or LTBI in young children.

Download Full-text

Evaluation of Gamma Interferon Immune Response Elicited by the Newly Constructed PstS-1(285-374):CFP10 Fusion Protein To Detect Mycobacterium tuberculosis Infection

Clinical and Vaccine Immunology ◽

10.1128/cvi.00726-13 ◽

2014 ◽

Vol 21 (4) ◽

pp. 552-560 ◽

Cited By ~ 9

Author(s):

Leonardo Silva de Araujo ◽

Fernanda Carvalho de Queiroz Mello ◽

Nidai de Bárbara Moreira da Silva ◽

Janaina Aparecida Medeiros Leung ◽

Silvia Maria Almeida Machado ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Tuberculin Skin Test ◽

Skin Test ◽

Positive Tuberculin Skin Test ◽

Gamma Interferon ◽

Preventative Treatment ◽

Mycobacterium Tuberculosis Infection ◽

Content Type ◽

Ifn Γ ◽

First Time

ABSTRACTThe PstS1 antigen is highly immunogenic, principally when combined with CFP10 during both latent and active TB infection. In the present study, a selectedpstS1gene fragment was cloned, fused with CFP10, and expressed inEscherichia coli. The product [PstS-1(285-374):CFP10] was compared to the recombinant fused RD1 (region of deletion 1) protein (ESAT-6:CFP10) in detectingMycobacterium tuberculosisinfection in 108 recent contacts of pulmonary tuberculosis (TB) cases, considering a positive tuberculin skin test (TST) to be the baseline. The release of gamma interferon (IFN-γ) in 22-h whole-blood and 5-day lymphocyte stimulation assays primed with each antigen was determined. All contacts were clinically followed for up to 1 year, and 87% of the tuberculin skin test-positive (TSTpositive) patients accepted preventative treatment. Concerning the IFN-γ response to PstS-1(285-374):CFP10 in the 22-h and 5-day assays, a slight increase in contact-TSTpositivedetection was observed (23/54 and 26/54) compared to the level seen with the RD1 protein (18/54 and 24/54) whereas in the TSTnegativegroup, similarly lower numbers (≤5/48) of responders were achieved for both antigens, except for RD1 in the 5-day assay (8/48). By combining the IFN-γ responders to both antigens in the 5-day assays, slightly higher increases in positivity were found in the TSTpositive(32/54) and TSTnegative(10/48) groups. Two of 12 untreated TSTpositivecontacts progressed to active TB and were concordantly positive in all assays, except for one contact who lacked positivity in the RD1 5-day assay. We demonstrated for the first time that PstS-1(285-374):CFP10 slightly increased contact positivity and detection of active disease progression, suggesting its potential application as a TB infection marker.

Download Full-text

Spontaneous Phthiocerol Dimycocerosate-Deficient Variants of Mycobacterium tuberculosis Are Susceptible to Gamma Interferon-Mediated Immunity

Infection and Immunity ◽

10.1128/iai.00097-11 ◽

2011 ◽

Vol 79 (7) ◽

pp. 2829-2838 ◽

Cited By ~ 44

Author(s):

Meghan A. Kirksey ◽

Anna D. Tischler ◽

Roxane Siméone ◽

Katherine B. Hisert ◽

Swapna Uplekar ◽

...

Keyword(s):

Immune Response ◽

Mycobacterium Tuberculosis ◽

Point Mutation ◽

Chronic Phase ◽

Gamma Interferon ◽

Wild Type ◽

Content Type ◽

Virulence Attenuation ◽

Ifn Γ ◽

Phthiocerol Dimycocerosate

ABSTRACTOnset of the adaptive immune response in mice infected withMycobacterium tuberculosisis accompanied by slowing of bacterial replication and establishment of a chronic infection. Stabilization of bacterial numbers during the chronic phase of infection is dependent on the activity of the gamma interferon (IFN-γ)-inducible nitric oxide synthase (NOS2). Previously, we described a differential signature-tagged mutagenesis screen designed to identifyM. tuberculosis“counterimmune” mechanisms and reported the isolation of three mutants in the H37Rv strain background containing transposon insertions in therv0072,rv0405, andrv2958cgenes. These mutants were impaired for replication and virulence in NOS2−/−mice but were growth-proficient and virulent in IFN-γ−/−mice, suggesting that the disrupted genes were required for bacterial resistance to an IFN-γ-dependent immune mechanism other than NOS2. Here, we report that the attenuation of these strains is attributable to an underlying transposon-independent deficiency in biosynthesis of phthiocerol dimycocerosate (PDIM), a cell wall lipid that is required for full virulence in mice. We performed whole-genome resequencing of a PDIM-deficient clone and identified a spontaneous point mutation in the putative polyketide synthase PpsD that results in a G44C amino acid substitution. We demonstrate by complementation with the wild-typeppsDgene and reversion of theppsDgene to the wild-type sequence that theppsD(G44C) point mutation is responsible for PDIM deficiency, virulence attenuation in NOS2−/−and wild-type C57BL/6 mice, and a growth advantagein vitroin liquid culture. We conclude that PDIM biosynthesis is required forM. tuberculosisresistance to an IFN-γ-mediated immune response that is independent of NOS2.

Download Full-text

Evaluation of Gamma Interferon (IFN-γ)-Induced Protein 10 Responses for Detection of Cattle Infected with Mycobacterium bovis: Comparisons to IFN-γ Responses

Clinical and Vaccine Immunology ◽

10.1128/cvi.05657-11 ◽

2012 ◽

Vol 19 (3) ◽

pp. 346-351 ◽

Cited By ~ 24

Author(s):

W. R. Waters ◽

T. C. Thacker ◽

B. J. Nonnecke ◽

M. V. Palmer ◽

I. Schiller ◽

...

Keyword(s):

Mycobacterium Bovis ◽

Similar Pattern ◽

Gamma Interferon ◽

Content Type ◽

Purified Protein Derivative ◽

Testing Protocol ◽

Archived Samples ◽

Ifn Γ ◽

Highly Correlated ◽

Mycobacterial Antigens

ABSTRACTGamma interferon (IFN-γ)-induced protein 10 (IP-10) has recently shown promise as a diagnostic biomarker ofMycobacterium tuberculosisinfection of humans. The aim of the current study was to compare IP-10 and IFN-γ responses uponMycobacterium bovisinfection in cattle by using archived samples from two aerosol inoculation studies. In the first study (104CFUM. bovisby aerosol,n= 7),M. bovispurified protein derivative (PPDb)-specific IP-10 and IFN-γ gene expression was detected as early as 29 days after challenge. PPDb-specific IP-10 and IFN-γ mRNA responses followed a similar pattern of expression over the course of this study and were highly correlated (r= 0.87). In the second study (105CFUM. bovisby aerosol,n= 5), IP-10 and IFN-γ (protein) responses to mycobacterial antigens were compared following challenge. IFN-γ responses to mycobacterial antigens were detected at 29 days after challenge and were sustained during the remainder of the study. IFN-γ responses to mycobacterial antigens exceeded corresponding responses in nonstimulated cultures. IP-10 responses to mycobacterial antigens exceeded preinfection responses at 7, 29, and 63 days after challenge. In contrast to IFN-γ responses, IP-10 responses to mycobacterial antigens generally did not exceed the respective responses in nonstimulated cultures. IP-10 responses to medium alone and to mycobacterial antigens followed a similar pattern of response. Correlations between IP-10 and IFN-γ (protein) responses were modest (r≈ 0.50 to 0.65). Taken together, these findings do not support the use of IP-10 protein as a biomarker for bovine tuberculosis using the current testing protocol and reagents; however, mRNA-based assays may be considered for further analysis.

Download Full-text

Incubation of Whole Blood at 39°C Augments Gamma Interferon (IFN-γ)-Induced Protein 10 and IFN-γ Responses to Mycobacterium tuberculosis Antigens

Clinical and Vaccine Immunology ◽

10.1128/cvi.00051-11 ◽

2011 ◽

Vol 18 (7) ◽

pp. 1150-1156 ◽

Cited By ~ 13

Author(s):

Martine G. Aabye ◽

Pernille Ravn ◽

Isik S. Johansen ◽

Jesper Eugen-Olsen ◽

Morten Ruhwald

Keyword(s):

Mycobacterium Tuberculosis ◽

T Cell ◽

Whole Blood ◽

Incubation Temperature ◽

Gamma Interferon ◽

Content Type ◽

Interleukin 7 ◽

Ifn Γ

ABSTRACTA rarely challenged dogma in cell-mediated immune (CMI) assays is the incubation temperature, 37°C. Fever augments proinflammatory immune responsesin vivo, and the aim of this study was to explore whether incubation at fever-range temperature could increase antigen-specific biomarker responses. We compared CMI responses following incubation of whole blood at 37°C and 39°C. Whole blood was obtained from (i) 34 healthy subjects whose blood was incubated with TB10.4 antigen, present in theMycobacterium bovisbacillus Calmette-Guérin vaccine and many environmental mycobacteria; (ii) 8 TB patients and 8 controls incubated withMycobacterium tuberculosis-specific antigens in the QuantiFERON-TB Gold test (QFT-IT); and (iii) from both groups incubated with a T cell mitogen. T cell responses (gamma interferon [IFN-γ]) and responses from antigen-presenting cells (IFN-γ-induced protein 10 [IP-10]) were determined. We further evaluated the effect of adding interleukin-7 (IL-7) and blocking IL-10 during incubation. In TB patients, IFN-γ and IP-10 levels were increased 4.1- and 3.4-fold, respectively, at 39°C incubation (P< 0.001). Similar results were seen after mitogen stimulation. In subjects responding to TB10.4, the effects were less pronounced and significant only for IP-10. Incubation at 39°C increased IP-10 and IFN-γ responsiveness to both antigens and mitogen in persons with baseline or initial low responses. Adding IL-7 and blocking IL-10 augmented the effects in synergy with fever-range temperature. Incubation at fever-range temperature vividly increases CMI responsiveness to antigen stimulationin vitroin tuberculosis patients and may increase the sensitivity of CMI assays.

Download Full-text

Early chest X-ray in persons with presumptive tuberculosis increases Xpert® MTB/RIF diagnostic yield and efficiency

Public Health Action ◽

10.5588/pha.19.0052 ◽

2020 ◽

Vol 10 (1) ◽

pp. 17-20

Author(s):

Z. Nadiah ◽

R. C. Koesoemadinata ◽

S. M. McAllister ◽

G. Putriyani ◽

L. Chaidir ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Diagnostic Yield ◽

Content Type ◽

X Ray ◽

Acid Fast Bacilli ◽

Number Needed To Screen ◽

Sputum Microscopy ◽

Chest X Ray

Adult presumptive tuberculosis (TB) patients (n = 1690) were screened for TB using a questionnaire, chest X-ray (CXR) and sputum microscopy for acid-fast bacilli (AFB); <named-content content-type="genus-species">Mycobacterium tuberculosis</named-content> culture was performed for 74% of the patients and Xpert® MTB/RIF was done for 17.2%. Among patients recruited, 943 (55.8%) were diagnosed with TB, of whom 870 (92.3%) were bacteriologically confirmed and 73 (7.7%) were clinically diagnosed on the basis of CXR. Using CXR prior to culture or Xpert testing reduces the number needed to screen from 7.6 to 5.0. Using CXR to triage for culture or Xpert testing reduces the number of missed cases and increases the efficiency of culture and Xpert testing.

Download Full-text

Mycobacterium bovis BCG-Specific Th17 Cells Confer Partial Protection against Mycobacterium tuberculosis Infection in the Absence of Gamma Interferon

Infection and Immunity ◽

10.1128/iai.01392-09 ◽

2010 ◽

Vol 78 (10) ◽

pp. 4187-4194 ◽

Cited By ~ 90

Author(s):

Teresa M. Wozniak ◽

Bernadette M. Saunders ◽

Anthony A. Ryan ◽

Warwick J. Britton

Keyword(s):

T Cells ◽

Mycobacterium Tuberculosis ◽

Th17 Cells ◽

Protective Immunity ◽

Bacterial Load ◽

Gamma Interferon ◽

Interleukin 17 ◽

Mycobacterium Tuberculosis Infection ◽

Significant Prolongation ◽

Ifn Γ

ABSTRACT Protective immunity against tuberculosis (TB) requires the integrated response of a network of lymphocytes. Both gamma interferon (IFN-γ)- and interleukin 17 (IL-17)-secreting CD4+ T cells have been identified in subjects with latent TB infection and during experimental Mycobacterium tuberculosis infection, but the contribution of Th17 cells to protective immunity is unclear. To examine their protective effects in vivo, we transferred mycobacterium-specific IL-17- and IFN-γ-secreting CD4+ T cells isolated from M. tuberculosis BCG-immunized IL-12p40−/− and IFN-γ−/− or wild-type mice, respectively, into M. tuberculosis-infected IL-12p40−/− or RAG−/− mice. In the absence of IL-12 and IL-23, neither IL-17-secreting (Th17) nor IFN-γ-secreting (Th1) BCG-specific T cells expanded or provided protection against M. tuberculosis. In RAG−/− recipients with an intact IL-12/IL-23 axis, both Th17 and Th1 cells were activated and induced significant protection against M. tuberculosis. The reduction in the bacterial load following transfer of IFN-γ−/− Th17 cells was associated with significant prolongation of survival compared to recipients of naïve IFN-γ−/− T cells. This effect was at the cost of an increased inflammatory infiltrate characterized by an excess of neutrophils. Therefore, Th17 cells can provide IFN-γ-independent protection against M. tuberculosis, and this effect may contribute to the early control of M. tuberculosis infection.

Download Full-text

Evaluation of Gamma Interferon and Antibody Tuberculosis Tests in Alpacas

Clinical and Vaccine Immunology ◽

10.1128/cvi.00405-12 ◽

2012 ◽

Vol 19 (10) ◽

pp. 1677-1683 ◽

Cited By ~ 16

Author(s):

Shelley Rhodes ◽

Tom Holder ◽

Derek Clifford ◽

Ian Dexter ◽

Jacky Brewer ◽

...

Keyword(s):

Sensitivity And Specificity ◽

Gamma Interferon ◽

Enzyme Linked Immunosorbent Assay ◽

Content Type ◽

South American Camelids ◽

Purified Protein Derivative ◽

History Of ◽

Similar Range ◽

Ifn Γ ◽

Antibody Tests

ABSTRACTWe describe the performance of cell-based and antibody blood tests for the antemortem diagnosis of tuberculosis (TB) in South American camelids (SAC). The sensitivity and specificity of the gamma interferon (IFN-γ) release assay, two lateral flow rapid antibody tests (Stat-Pak and Dual Path Platform [DPP]), and two enzyme-linked immunosorbent assay (ELISA)-based antibody tests (Idexx and Enferplex) were determined using diseased alpacas fromMycobacterium bovisculture-confirmed breakdown herds and TB-free alpacas from geographical areas with no history of bovine TB, respectively. Our results show that while the sensitivities of the IFN-γ and antibody tests were similar (range of 57.7% to 66.7%), the specificity of the IFN-γ test (89.1%) was lower than those of any of the antibody tests (range of 96.4% to 97.4%). This lower specificity of the IFN-γ test was at least in part due to undisclosedMycobacterium microtiinfection in the TB-free cohort, which stimulates a positive purified protein derivative (PPD) response. The sensitivity of infection detection could be increased by combining two antibody tests, but even the use of all four antibody tests failed to detect all diseased alpacas. These antibody-negative alpacas were IFN-γ positive. We found that the maximum sensitivity could be achieved only by the combination of the IFN-γ test with two antibody tests in a “test package,” although this resulted in decreased specificity. The data from this evaluation of tests with defined sensitivity and specificity provide potential options for antemortem screening of SAC for TB in herd breakdown situations and could also find application in movement testing and tracing investigations.

Download Full-text

Tuberculin-Purified Protein Derivative-, MPT-64-, and ESAT-6-Stimulated Gamma Interferon Responses in Medical Students before and after Mycobacterium bovis BCG Vaccination and in Patients with Tuberculosis

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.6.6.934-937.1999 ◽

1999 ◽

Vol 6 (6) ◽

pp. 934-937 ◽

Cited By ~ 66

Author(s):

P. D. R. Johnson ◽

R. L. Stuart ◽

M. L. Grayson ◽

D. Olden ◽

A. Clancy ◽

...

Keyword(s):

Medical Students ◽

Venous Blood ◽

Gamma Interferon ◽

Active Tuberculosis ◽

Skin Tests ◽

Purified Protein Derivative ◽

Before And After ◽

Specific Proteins ◽

Ifn Γ ◽

Interferon Responses

ABSTRACT QuantiFERON-TB (QIFN) (CSL Limited) is a whole-blood assay for the recognition of infection withMycobacterium tuberculosis. QIFN measures gamma interferon (IFN-γ) production when purified protein derivatives (PPDs) of mycobacteria are incubated with venous blood samples. The specificity of QIFN in medical students before and after BCG immunization was assessed, and sensitivity in patients with tuberculosis was assessed. Antigens were PPD derived from M. tuberculosis and twoM. tuberculosis-specific proteins, ESAT-6 and MPT-64. Of 60 medical students, all of whom had 0-mm tuberculin skin tests (TSTs) at study entry, 58 (97%) were initially classified as negative forM. tuberculosis infection by PPD QIFN. Five months after BCG immunization, 7 of 54 students (13%) had a TST result of ≥10 mm and 11 of 54 students (20%) tested positive by PPD QIFN. ESAT-6- and MPT-64-stimulated IFN-γ responses in the medical students were negative prior to and after BCG immunization. For patients with active tuberculosis, 12 of 19 (63%) were positive by PPD QIFN, 11 of 19 (58%) were positive by ESAT-6 QIFN, and 0 of 12 were positive by MPT-64 QIFN. In conclusion, PPD QIFN was negative in 97% of a low-risk population who had not received BCG and who had negative TSTs. The specificities of both the TST and PPD QIFN were reduced following BCG immunization. PPD QIFN and ESAT-6 QIFN were of similar and moderate sensitivity in patients with active tuberculosis, but ESAT-6 QIFN is likely to be more specific because it is not influenced by past BCG exposure.

Download Full-text

Clinical Significance of Interleukin-2/Gamma Interferon Ratios in Mycobacterium tuberculosis-Specific T-Cell Signatures

Clinical and Vaccine Immunology ◽

10.1128/cvi.05013-11 ◽

2011 ◽

Vol 18 (8) ◽

pp. 1395-1396 ◽

Cited By ~ 18

Author(s):

Franziska Suter-Riniker ◽

Antonia Berger ◽

Désirée Mayor ◽

Pascal Bittel ◽

Patricia Iseli ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

T Cell ◽

Simultaneous Determination ◽

Clinical Significance ◽

Interleukin 2 ◽

Gamma Interferon ◽

Content Type ◽

Ifn Γ

ABSTRACTThe simultaneous determination of interleukin-2 (IL-2) and gamma interferon (IFN-γ) in QuantiFERON-TB test plasma supernatants permitted the detection of shifts inMycobacterium tuberculosis-specific T-cell signatures. A subset of the 84 subjects tested revealed a significantly elevated IL-2/IFN-γ ratio, which may be a marker for the successful elimination ofM. tuberculosisinfection.

Download Full-text

Diagnosis of Active Tuberculosis in China Using an In-House Gamma Interferon Enzyme-Linked Immunospot Assay

Clinical and Vaccine Immunology ◽

10.1128/cvi.00044-09 ◽

2009 ◽

Vol 16 (6) ◽

pp. 879-884 ◽

Cited By ~ 39

Author(s):

Xinchun Chen ◽

Qianting Yang ◽

Mingxia Zhang ◽

Michael Graner ◽

Xiuyun Zhu ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Elispot Assay ◽

Lung Diseases ◽

Gamma Interferon ◽

Active Tuberculosis ◽

Skin Tests ◽

Healthy Controls ◽

Specificity And Sensitivity ◽

Different Populations ◽

Ifn Γ

ABSTRACT Gamma interferon (IFN-γ) release assays have been proven to be useful in the diagnosis of Mycobacterium tuberculosis infection. Nevertheless, their specificity and sensitivity vary among the different populations studied. Here, we evaluate the value of an in-house IFN-γ enzyme-linked immunospot (ELISPOT) assay in the diagnosis of active tuberculosis (TB) in Shenzhen, China, where the prevalence of tuberculosis is severe and Mycobacterium bovis BCG vaccination is mandatory at birth. A total of 305 patients with active tuberculosis, 18 patients with nontuberculosis lung diseases, and 202 healthy controls were recruited in this study. Among them, 156 individuals were simultaneously tested for IFN-γ responses by the commercial QuantiFERON-TB Gold in-tube (QFT-IT) assay. Tuberculin skin tests (TST) were performed with 202 healthy controls. The overall sensitivities of the ELISPOT and QFT-IT assays for active tuberculosis were 83.60% and 80.85%, respectively; the specificities were 76.6% and 73.26%, respectively. The IFN-γ ELISPOT responses, but not those of the TST, were significantly correlated with TB exposure (r = −0.6040, P < 0.0001). The sensitivities of the ELISPOT assay varied for patients with different forms of tuberculosis, with the highest sensitivity for patients with sputum-positive pulmonary tuberculosis (89.89%) and the lowest for those with tuberculous meningitis (62.5%). In conclusion, the IFN-γ ELISPOT assay is a useful adjunct to current tests for diagnosis of active TB in China. The ELISPOT assay is more accurate than TST in identifying TB infections.

Download Full-text