Although the World Health Organisation (WHO) declared tuberculosis (TB) to be a global health emergency over 15 years ago,1 TB remains as one of the leading infectious causes of morbidity in the world, accounting for 8 to 10 million cases per year.2 Despite this expanding threat to global public health, our tools for the diagnosis and prevention of TB are over one hundred years old, and they are inadequate to control the epidemic.3 However, significant advances have recently been made in the diagnosis of latent TB infection (LTBI) which promise to substantially improve TB control.4

The epidemiology of TB is distinct in that the reservoir of latently infected individuals, estimated to be a third of the world's population, is much larger than the number of active TB cases that occur.5 As a result it is becoming increasingly more evident, particularly in non-endemic countries where latently infected migrants and recently-infected contacts give rise to the bulk of the TB case-load, that effective identification and treatment of LTBI has the potential to reverse the increasing TB burden.6,7

The natural history of TB is complex and remains poorly understood. After establishing initial infection, tubercle bacilli may cause disease immediately, or they may lie dormant for many decades; evidence suggests only 5% to 10% of infected immunocompetent people go on to develop active disease over their lifetime.8 Diagnosis and treatment of LTBI is based on “targeted testing” to selectively identify persons at highest risk of progression from LTBI to active disease. This includes recently-infected individuals and all those with suppressed or immature immune systems, regardless of when they acquired infection.9

Diagnosis of LTBI has hitherto been defined as a positive tuberculin skin test (TST) in an otherwise asymptomatic person exposed to tuberculosis with no clinical or radiological evidence of active disease. The TST is based on the detection of a cutaneous delayed-typed hypersensitivity response to purified protein derivative, a poorly defined mixture of more than two hundred Mycobacterium tuberculosis proteins. However, although the TST is cheap and widely utilised, it has several drawbacks. Antigenic cross-reactivity of purified protein derivative reduces specificity in individuals previously vaccinated with BCG and/or exposed to nontuberculous mycobacteria, resulting in false-positive results.10 In addition, the TST is acknowledged to have poor sensitivity in individuals with compromised immune systems (eg, patients with HIV infection or iatrogenic immunosuppression, and children), resulting in false-negative results.11 Additional difficulties are logistic and include the need for the test to be performed by a trained health care professional and the requirement for a return visit to have the result read.

T-cell interferon-gamma release assays (IGRAs) have been developed as an alternative immunodiagnostic approach to the TST for detecting M. tuberculosis infection.9,12,13 Their development has stemmed from advances in mycobacterial genomics which identified a genomic segment (Region of Difference 1) that is deleted from all strains of BCG14 and the majority of environmental mycobacteria (except M. Kansasi, M. szulgai, M. marinum, M. flavescens, and M. gastrii).15 Two antigens encoded by this segment, early secretory antigen target-6 (ESAT-6) and culture filtrate protein 10 (CFP-10), are strong targets of Th1 T-cells in M. tuberculosis infection; their ability to elicit strong, specific, T-cell responses reduces the frequency of false-positive TST results in individuals who have previously received BCG vaccination.3,16,17 This important property has been harnessed to form the basis of the two ex vivo IGRAs that are currently available: the ELISpot, which directly counts the number of IFN-γ secreting T cells, and the whole-blood ELISA, which measures the concentration of IFN-γ secretion. Both assays have been commercially licensed with the ELISpot assay available as the T-SPOT™-TB (Oxford Immunotec, Abingdon, UK) and the ELISA as the QuantiFERON™-TB Gold In-Tube (Cellestis, Carnegie, Australia). The main similarities and differences between the two assays are summarized in Table 1.

Significant challenges exist in directly assessing whether IGRAs are superior to the TST in diagnosing LTBI as there is no available reference standard test. Studies attempting to quantify the test performance of IGRAs have therefore used surrogate markers for LTBI, including: (1) the level of contact with infectious cases, (2) active TB disease as a surrogate for LTBI and, to assess specificity, (3) IGRA-negative status in healthy individuals at low-risk of TB infection in low-prevalence settings.

Correctly identifying and treating LTBI in high-risk individuals with impaired cell-mediated immunity is a central concern for clinicians. Groups at particular risk of progressing from LTBI to active TB disease are HIV-positive individuals and individuals with immune-mediated inflammatory diseases (such as rheumatoid arthritis, Crohn's disease, and ankylosing spondylitis), who are iatrogenically immunosuppressed or treated with TNFα blockade.68 Unfortunately, the TST is well-recognised to have poor sensitivity in these immunocompromised populations; hence, there is a need to evaluate the diagnostic performance of the IGRAs in these populations.

As the IGRAs begin to be more widely used outside the research setting, it is important to consider the reliability of the blood tests in routine clinical use. Both IGRAs are known to suffer from indeterminate results.3 These most commonly arise due to a failed positive control, which usually reflects underlying cellular immune suppression. Data from a wide range of studies suggest that indeterminate results are not infrequent with the ELISA, occurring in between 5 and 40% of cases with the second generation ELISA, although they may be less common with the newer in-tube ELISA.45,49,55,90,91 Indeterminate ELISA results seem to be associated with young (<5 years) and old age (>80 years) and immunosuppression, either due to HIV infection or immunosuppressant medication.91 Conversely, indeterminate results are rarer with the ELISpot, occurring in 0 to 5.4% of tests undertaken.91

Preventive treatment for IGRA-positive contacts will only confer clinical benefit if they are at increased risk of progression to active TB compared with IGRA-negative contacts. Only very recently has clinical outcome of this nature begun to emerge from longitudinal studies. To date, 6 studies have been published assessing the predictive value of IGRAs.92–97 A small study from a low-burden region investigated the use of the ELISA in 601 contacts, and found that a significantly higher percentage of untreated household contacts with a positive ELISA progressed to TB disease as compared to contacts with a 5–mm positive TST, with all 6 incident cases ELISA-positive at recruitment.92 More recently, Aichelberg et al explored the prognostic power of a positive ELISA (QuantiFERON-Gold In-tube) result in HIV-positive individuals without active TB at recruitment.97 The investigators found that over a median follow-up period of 19 months, 3/37 individuals with a positive ELISA at baseline went on to develop active TB, whereas 0/738 with a negative ELISA subsequently developed TB.97

A study from Turkey with 908 child contacts and 15 incident cases found that contacts with a positive ELISpot had a significantly increased risk of developing active TB as compared to contacts with a negative ELISpot.93 TB incidence in ELISpot-positive contacts was similar to that in TST-positive contacts, although the ELISpot predicted these from fewer contacts tested.93 However, as per national guidelines, a large percentage of the children in this study received isoniazid chemoprophylaxis, which is likely to have resulted in an underestimation of the incidence rate ratios.

A longitudinal study conducted in a high-prevalence setting in Gambia followed up 2348 household contacts for 2 years.94 The study found that 11/649 ELISpot-positive individuals developed active disease as compared to 14/843 TST-positive individuals; 10/1087 ELISpot-negative and 11/1387 TST-negative individuals developed TB.94 It was concluded that either test could be used in the initial screen for LTBI in contacts, although neither the TST nor the ELISpot predicted subsequent progression to TB disease. Reasons for this lack of prognostic power of both the TST and ELISpot remain unclear but may partly relate to the highly endemic setting in which de novo community transmission outside the household-setting accounts for a substantial proportion of incident TB cases.98

Longitudinal clinical outcome studies have provided the preliminary evidence base that supports the use of IGRAs to target preventive therapy for recent IGRA contacts, and this will enable prevention of a similar number of cases as when using the TST, but necessitates treatment of significantly fewer contacts.

Over the last few years, the IGRAs have gained regulatory approval in the US and Europe and as their evidence base has increased, a number of national guidelines have been rewritten to recommend their use in the diagnosis of LTBI. In Europe and Canada, guidelines advise that the IGRA should be used in 2 situations99,100:

• 1.

  As a confirmatory test in individuals who have already tested positive with the TST

• 2.

  As a direct replacement for the TST in those individuals in whom the TST is likely to be unreliable (immunocompromised individuals)

Conversely, in the US and Japan, guidelines recommend that the IGRA should completely replace the TST as the test of choice for LTBI in all individuals.101

Economic considerations played a role in the eventual recommendations made by different countries. Although the IGRAs are more expensive than the TST,3 health economic analyses have found that they are cost effective as they reduce the number of individuals needing unnecessary chemoprophylaxis and clinical and laboratory monitoring while on drug therapy.102,103 One such economic analysis, conducted by the United Kingdom National Institute of Health and Clinical Excellence (NICE), concluded that a two-step TST and confirmatory IGRA approach would be most cost-effective strategy, thereby forming the basis of the recommendation made by NICE and subsequently adopted by most other European countries.99

A consequence of the European/Canadian recommendations is that, potentially, immunocompetent individuals with a negative TST who may have been IGRA-positive will not be identified as having LTBI and hence, left untreated. On the other hand, it could be argued that the US/Japanese guidelines imply potential overtreatment of individuals who are IGRA-positive/TST-negative because the risk of progression to active TB in this subgroup is yet to be quantified. This highlights the urgent need for more longitudinal data to quantify the predictive value of positive IGRA results and, in particular, the prognosis of contacts with discordant IGRA and TST results.

Interferon-gamma release assays are recognised as the 100-year upgrade of the TST for diagnosing LTBI.4 However, it is important to realise that they are ultimately a work in progress with an ever-expanding and evolving clinical and research evidence base. As a result, their advantages and limitations are becoming clearer. Aside from the inability for IGRAs to differentiate between active and latent TB, longitudinal studies involving serial IGRA testing have shown that these tests cannot be used for treatment monitoring or as a test of cure.3 However, there is promising recent evidence that simultaneous profiling of IL-2 and IFN-γ secretion at the single T-cell level correlates with therapeutic response and may be useful in disease monitoring.104,105

While existing IGRAs have higher diagnostic sensitivity than the TST, ongoing research indicates that next-generation assays will have higher sensitivity. One promising approach used for both the ELISA and ELISpot has been to include additional antigens.106 Recent studies have shown that incorporating novel antigens, Rv2645 for the in-tube ELISA107 and RV3879c for the ELISpotPLUS,44 to ESAT-6 and CFP-10 significantly improves sensitivity without reducing specificity.

Current studies are also exploring whether measuring additional, alternative downstream chemokines secreted by IFN-γ-activated macrophages, such as inducible protein 10 (IP-10), monocyte chemoattractant protein (MCP-2) and monocyte inducible protein (MIG), can provide higher sensitivity than the currently available assays which measure only IFN-γ.108 A recent case-control study found that combining IFN-γ and IP-10 significantly increased sensitivity as compared to using the in-tube IFN-γ ELISA alone, without adversely affecting specificity.109 Although these are encouraging results, there is a need for prospective, longitudinal, studies in a range of patient groups (immunocompromised, children, etc.) to establish the accuracy and predictive value of these new IFN-γ and IP-10 tests.

IGRAs are an important new class of diagnostic tool that have the potential to supersede the TST and revolutionise the diagnosis of LTBI. In the few years since their development, the evidence base supporting their use has expanded so rapidly that they now form an integral part of many national guidelines from low-prevalence countries. However, residual uncertainty remains in the differing recommendations framed in these guidelines which reflects the need for large, longitudinal studies to further define the prognostic value of positive IGRA results for development of active TB disease, particularly in high-risk groups and subgroups with discordant IGRA and TST results.

Professor Lalvani is the inventor of patents underpinning T cell-based diagnosis. The Lalvani ELISpot was commercialised by an Oxford University spin-out company (T-SPOT.TB®, Oxford Immunotec Ltd, Abingdon, UK) in which Oxford University and Professor Lalvani have a minority share of equity.