IGRAs in the diagnosis of latent tuberculosis

Current latent tuberculosis infection diagnostic tests consist of the tuberculin skin test  and interferon gamma release assays (IGRAs), which will help control the tuberculosis pandemic

Clíona Ní Cheallaigh

Colm Bergin

St James’s Hospital, Dublin;

Trinity College, Dublin

Globally, tuberculosis kills more people than any other bacterial infection. There were 8.7 million new cases and 1.4 million deaths of tuberculosis in 2011. The number of drug-resistant cases of TB continues to grow, with an estimated 310,000 cases of multi-drug resistant tuberculosis, that is, tuberculosis which is resistant to the two front-line anti-tuberculous antibiotics: isoniazid and rifampicin, in 2011. Patients with multi-drug resistant tuberculosis (MDR-TB) are more costly to treat, remain infectious for longer and almost one in four cases of MDR-TB results in death.1 Recent years have seen the development of extensively drug-resistant tuberculosis (XDR-TB), a subset of multi-drug resistant tuberculosis which has acquired additional resistance to the most effective second-line TB antibiotics: fluoroquinolones and any of the three injectable agents: amikacin, capreomycin and kanamycin. Mortality rates with extensively drug-resistance tuberculosis in HIV-infected individuals have been reported to be as high as 98%.2

Tuberculosis infections are acquired when an individual inhales droplets containing M. tuberculosis, produced by coughing by another individual with active pulmonary TB disease. Three outcomes of infection are possible: (1) elimination of infection, (2) containment of the infection as latent tuberculosis infection (LTBI) or (3) development of active tuberculosis disease (Figure 1).  It is estimated that 10–20% of exposed individuals eliminate the infection.3 The majority of those who do not eliminate the infection will develop latent tuberculosis: these individuals are asymptomatic and non-infectious, carrying about 100–1000 bacteria, which are in a semi-dormant state (Figure 1). It is estimated that one third of the world’s population has latent tuberculosis. Individuals with latent tuberculosis are asymptomatic and non-infectious but are estimated to have an approximately 5–10% life-time risk of losing control of the infection and developing active TB, although a recent study from the US estimated this as a much lower risk of 0.04 cases per 100 person years.4

Those with active tuberculosis have a variety of clinical presentations. Systemic symptoms common to all sites of infection include fever, weight loss and night sweats. The most common site of disease is the lungs and symptoms of pulmonary TB include persistent cough which is usually, but not always, productive of sputum, which may be blood-stained. Extensive destruction of lung tissue may result in catastrophic haemoptysis. Extra-pulmonary disease is present in 10–42% of patients.13 The most common site of extra-pulmonary disease is the pleura, but TB disease can also affect the meninges, the kidneys, the peritoneum and bones, including the spine. 

A number of conditions increase the risk of progression from latent to active TB. HIV increases the risk of re-activation of latent tuberculosis infection to more than 10 per 100 person years.6–8  Tuberculosis is the most common opportunistic infection in HIV-infected individuals and is responsible for one third of HIV-associated deaths.9 Anti-TNF agents have revolutionised the treatment of a number of inflammatory conditions, including rheumatoid arthritis and Crohn’s disease. However, their use is associated with an increased risk of active tuberculosis, which is often disseminated and clinically difficult to diagnose.10 Other host factors that increase the risk of developing active tuberculosis and/or increase disease severity include diabetes, smoking11,12 and extremes of age. 

Diagnosing tuberculosis 

Improved diagnostics would facilitate quicker identification of individuals with active and latent infection, and are needed to control the TB pandemic.13 The gold standard method of detection of active tuberculosis remains to be the culture of mycobacteria, which is slow and inaccurate. The recently developed GeneXpert PCR method is quicker, but also has limited sensitivity in smear-negative and extra-pulmonary cases.14 

Latent TB infection represents a reservoir from which active disease and subsequent transmission can propagate, particularly when the immune system is compromised. Isoniazid preventative treatment of HIV-infected individuals with positive tuberculin skin tests (TST) has been shown to reduce the risk of developing active TB by 60%,21 and observational studies have shown an additive benefit when it is used alongside antiretroviral treatment.22–24 It is estimated that rates of LTBI in the community need to be less than 1% to allow TB elimination.25 However, latent tuberculosis diagnosis is particularly challenging. Development of new LTBI diagnostics is hampered by a lack of understanding of latent tuberculosis and, particularly, by the lack of a gold standard method of LTBI diagnosis to use as a comparator. Ideally, a LTBI diagnostic test would actually identify the small proportion of individuals who will fail to contain the infection and progress to active tuberculosis.

LTBI diagnostics

Current LTBI diagnostic tests consist of the tuberculin skin test (TST) and interferon gamma release assays (IGRAs). The tuberculin skin test was developed over 100 years ago and consists of the intradermal injection of purified mycobacterial protein derivatives (PPD) and the measurement, 48–72 hours later, of resulting skin induration, which is thought to represent the cell-mediated immune reaction to PPD. Until recently, the TST was the only test available to diagnose LTBI. However, the TST has impaired sensitivity in immunocompromised individuals including those infected with HIV26,27 and in patients with severe active disease. Perhaps the biggest disadvantage to the TST is that it requires a return visit at 48–72 hours: a step with which as few as 30% of patients comply.28 The TST can also be falsely positive in the setting of recent BCG vaccination. The positive predictive value of the TST for subsequent development of active tuberculosis is low, but it has been used successfully to guide LTBI treatment with isoniazid.

Interferon gamma release assays (IGRAs) are recently developed blood tests that measure in vitro responses to mycobacterial RD1 antigens, which are unique to mycobacteria in the Mycobacterium tuberculosis complex (Mycobacterium tuberculosis, Mycobacterium bovis, Mycobacterium africanum) and Mycobacterium kansasii and are not present in M. bovis BCG.29 There are three commercially available assays: the Quantiferon 3G In Tube (QFT-IT) assay (Qiagen), which utilises an ELISA technique to measure the amount of interferon gamma secreted in response to ESAT-6, CFP-10 and TB 7.7; the recently released Quantiferon Plus which uses ESAT-6 and CFP-10 of varying lengths to stimulate CD4+ and CD8+ T-cells; and the T.SPOT-TB (Oxford Immunotec, Abingdon, UK), which uses an ELISPOT to quantify the number of cells producing interferon gamma in response to ESAT-6 and CFP-10. 

Advantages of IGRAs over TSTs include lack of cross-reactivity to BCG and atypical mycobacteria, an internal control for false negative results (although clinically this control may have limited sensitivity), and the need for only one visit to test.  In addition, it has been reported that the effect of immunocompromise including HIV infection on IGRA sensitivity may be less marked than that on the TST, although sensitivity of both IGRAs does appear to be impaired in advanced HIV.30

A comparison of sensitivity and specificity for tests for LTBI is impossible in the absence of a gold standard comparator. The sensitivity and specificity of the tests for diagnosis of active tuberculosis has been used to evaluate the tests, but the utility of this approach is questionable given the differences between active and latent tuberculosis. Only measuring their positive and negative predictive values for the subsequent development of active tuberculosis can truly assess the value of the tests. Unfortunately, positive test results on TST and/or IGRA are only weakly predictive of the subsequent development of active tuberculosis.31,32

The negative predictive value of both IGRAs and TST is acceptable in low TB prevalence countries but is less accurate in high TB prevalence settings. In addition, the clinical significance of the fact IGRA results often vary over time, with positive results changing to negative and vice-versa, especially in those with borderline results,33 is poorly understood and difficult to interpret. The recent development of the QuantiFERON Gold Plus, which purports to stimulate CD8+ in addition to CD4+ T-cells, may offer improved sensitivity in the diagnosis of active tuberculosis and be less affected by immunodeficiency but its positive and negative predictive value for patients at risk of developing active tuberculosis remains to be established.

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