Latent Infection

Latent tuberculosis infection (LTBI) is a state of persistent immune response to stimulation by Mycobacterium Tuberculosis antigens without evidence of active TB. Tools for reliable identification of patients with LTBI are required for a rational use of preventive therapy. Nowadays, the most applied diagnostic tools for LTBI are the in vivo Tuberculin Skin Test (TST) and the ex vivo Interferon-γ Release Assays (IGRAs). While useful, these tests are unable to determine the likelihood of progressing to active TB.

TST reveals the body’s hypersensitivity to the presence of TB bacillus proteins, a state acquired in most cases after initial infection. The assay consists of an intradermal injection of 0.1 ml of tuberculin purified protein derivative (PPD) into the inner surface of the forearm. Reading should be made 48–96 hours later by measuring the skin induration caused by the reaction. Although very simple, this test requires standardization, training and appropriate supervision. Bacillus Calmette-Guerin (BGC) vaccination can also cause hypersensitivity, leading to false positive results. Conversely, IGRAs detect circulating T-cells responsive to specific Mycobacterium Tuberculosis antigens, which are absent after BCG vaccination as well as after exposure to most non-tuberculosis Mycobacteria [16]. However, a recent meta-analysis comparing the accuracy of IGRAs vs. TST for the diagnosis of LTBI concluded that IGRAs may be better only in immune competent children aged >5 years residing in high-income areas. Even in these subjects, IGRAs sensitivity was only 67–86%, indicating that neither test may rule out LTBI nor confirm with certainty a true negative diagnosis [17]. The IGRAS currently in use primarily elicit a CD4+ response, but evidence supports the important contribution of CD8+ T-cells in host defense against TB. A new generation of QFT-GIT, the QuantiFERON-TB Gold Plus (QFT-Plus), contains peptides able to stimulate IFN-γ production by both CD4+ and CD8+ T-cells. Early data suggest that this test offers higher sensitivity and specificity, particularly in patients at the highest risk for TB-infection and in immunocompromised patients [18, 19]. However, larger studies are needed to confirm the accuracy of the QFT Plus [20].

Per current WHO recommendations [21, 22] and other international guidelines [23, 24] either TST or IGRAs can be used to identify candidates for LTBI treatment in high and middle-income countries. The TST is more affordable and is therefore preferred in low-income regions, even if is less specific than IGRAs in vaccinated patients [17]. TST should be also used in children, patients with immunodeficiency, people who live with patients infected with TB and health care workers [25, 26].

Active Tuberculosis

TB diagnosis is still based on the observance of clinical symptoms (unexplained weight loss, decreased appetite, night sweats, fever, fatigue, coughing for >2 weeks, hemoptysis, chest pain, etc.); radiographic findings; and microbiological testing. Chest X-ray is the most sensitive (>95%) test for the diagnosis of pulmonary TB in immune-competent patients and, for this reason, is probably the best strategy for screening for TB.

Bacteriological procedures are the most specific and the only accepted diagnostic test to confirm a diagnosis of TB. These include:

1. Bacilloscopy: microscopic assessment of sputum for Mycobacterium TB is the initially recommended test due to its low cost and simplicity. However, the role of Bacilloscopy is limited due to moderate sensitivity (60–70%) and possible false positives due to identification of atypical Mycobacteria (specificity 95–98%) [5].
2. Culture: sputum culture (especially liquid media) is the most sensitive bacteriological method available and considered the diagnostic gold standard.
3. Mycobateria specie identification, necessary to confirm TB vs infection by other Mycobacteria.
4. Drug susceptibility test or Antibiogram: in vitro testing of anti-TB drug sensitivity is critical for selecting an appropriate TB regimen but takes several weeks. Novel molecular methods that can provide results in hours have recently become available.

Drug-resistant TB

The immediate future for the management of TB will involve the use of rapid molecular microbiological techniques [27]. Susceptibility drug tests can be either phenotypic or genotypic. Phenotypic DST, widely used in developing countries, is based on assessing bacterial growth with fixed concentrations of first (e.g., INH, RIF, ethambutol [EMB], and streptomycin [SM]) or secondary-line anti-TB drugs. This method is simple and cost-effective; however, it leads to considerable delays and is unreliable for EMB and other drugs. Genotypic methods take advantage of specific mutations that are associated with response to particular drugs [28]. Molecular methods can be performed directly on sputum or other samples, and can provide results (including drug susceptibility) in 24 to 48 hours. The most used methods are:

1. GenXpert Mycobacterium Tuberculosis/RIFAMPICIN (GENXpert MTB/RIF), is a nucleic acid amplification assay conducted in sputum that simultaneously tests for presence of Mycobacterium Tuberculosis DNA and RIF resistance. This is a simple and reproducible test with a 90% sensitivity (reaching 98% in patients with positive bacilloscopy but dropping to 70% in smear negative disease) and 99% specificity. The assay is automated and does not require laboratory infrastructure. The identification of RIF resistance is often considered a marker of MDR-TB (in 95% of cases) with INH resistance [29]. WHO has recently recommended Gene-Xpert as an initial diagnosis test for patients with HIV suspected of having TB or for patients who are considered at risk for RIF resistance or MDR-TB. Similar assays currently under development include: the GeneXpert Omni, also intended for TB testing combined with RIF resistance assessment but uses a cheaper and more portable platform, potentially allowing for bedside testing and the Xpert MTB/RIF Ultra, a more sensitive and specific assessment which may improve detection of TB in smear-negative patients.
2. GenoType, also known as line probe assay (LPA), a test that can detect RIF or INH resistance within 48 hours [29, 30]. Additionally, the test is very accurate for detecting resistance to fluoroquinolones and second line injectable [31]. The assay is based on a conventional polymerase chain reaction, thus, requires access to a biomolecular laboratory.

Classification of TB Drugs (Table 1)

The current classification divides anti-TB drugs in 4 groups: A, B, C, D [36]. This classification is specifically designed to incorporate the treatment of RIF-resistant or MDR-TB cases [37].

• – Group A includes fluoroquinolones (high dose of Levofloxacin, Moxifloxacin, and Gatifloxacin). These are considered core drugs due to their bactericidal and sterilizing activity, as well as good safety profile.
• – Group B includes injectable drugs (Streptomycin, Amikacin, Kanamycin and Capreomycin), which have quality bactericidal activity but poorer safety profile than group A medications.
• – Group C is composed by Ethionamide, Prothionamide, Cycloserine/Terizidone, Clofazimine and Linezolid. These drugs are recommended as core second-line medicines for MDR-TB given growing evidence about their efficacy and tolerability.
• – Group D includes non-core MDR-TB drugs and is divided in 3 subgroups: D1: high dose INH, EMB, Pirazinamide; D2: Bedaquiline and Delamanid; and D3: Meropenem, Para-Aminosalicylic Acid, Imipenem-Cilastatin, Amoxicillin-Clavulanate, and Clarithromycin.

New trends in TB

After 50 years of minimal anti-TB drug development, a promising pipeline has recently emerged through the repurposing of old drugs and discovery of new compounds. Combinations of new and existing drugs are being assessed to shorten the duration of therapy and to effectively treat cases of MDR TB. There has also been progress in development of drugs that are active against dormant populations of Mycobacterium tuberculosis.

Bedaquiline is the first new anti-TB drug approved by the Food and Drug Administration (FDA) in 50 years to be used as part of combination treatment with MDR-TB for adults when other alternatives are not available [38]. It targets both active and dormant bacilli [39, 40], and therefore has the profile required of a core drug. A randomized phase II trial reported faster sputum-culture negative conversion in the patients receiving Bedaquiline (hazard ratio: 11.8, 95% confidence interval: 2.3–61.3) [41]. Cure rates were 58% for patients who received Bedaquiline compared to 32% for controls (p = 0.003) [42]. However, another trial showed a small increase in all-cause mortality among patients in the Bedaquiline arm [43]. Common adverse reactions associated with Bedaquiline were an increased QTc interval and cross-resistance with Clofazimine [44, 45].

Delamanid, approved by the European Medicine Agency, is a metronidazole derivate with bactericidal and sterilizing activity [46, 47]. Unlike Bedaquiline, Delamanid does not show cross-resistance with other anti-TB drugs. A recent study showed that when added to standard-of-care chemotherapy in patients with MDR TB, Delamanid was associated with a significantly higher proportion of sputum culture conversion at two months of treatment initiation (45% versus 29%) [48, 49]. QTc prolongation was also observed in Delamanid-treated patients [50, 51].

Linezolid may be also considered an “essential” drug. Two small randomized clinical trials [52, 53] and various meta-analyses [54, 55, 50] have shown its effectiveness for treating MDR-TB, especially XDR-TB. However, there are two downsides of Linezolid: its cost and toxicity profile (hematological disorders and polyneuropathies) when administered for more than 6–8 weeks [56]. Sutezolid is a Linezolid analogue which has shown greater anti-mycobacterial activity in clinical trials and is active against non-replicating Mycobacterium Tuberculosis both in vitro and in vivo [57]. Recent studies showed highly additive effects for combination of this drug with rifamycins [58]. In addition, phase I trials showed sputum early bactericidal activity in patients with TB and no hematological toxic effects [59].

Regimens for new cases of TB

WHO currently recommends a 6-month treatment regimen for TB; combining four effective medications, three of which should be “essential” drugs (INH, RIF, Z), two with good bactericidal activity (INH, RIF), and another two with good sterilizing capacity (RIF, Z). EMB has the role of an “accompanying” drug; supporting the core drugs to avoid resistance selection. EMB is not included in the regimen to eliminate bacilli and, in theory, it is no longer necessary after the bacteriological conversion is achieved [60]. Thus, patients are treated with INH, RIF, pyrazinamide and EMB for the first two months (intensive phase), followed by INH and RIF for the remaining 4 months (continuation phase) [34]. Importantly, if a core drug cannot be used because of documented resistance or toxicity, it should be replaced by another drug with a similar efficacy (i.e., bactericidal and sterilizing properties). Similarly, an accompanying drug should be replaced by another drug with an equivalent action. Patients with HIV infection and those with certain forms of extrapulmonary TB (meningeal, disseminated) may have their treatment extended to 9–12 months [66, 61].

WHO Recommendations for Drug Resistant TB

Following the current WHO recommendations, supported by a multi-center study of 1,200 patients, a stepwise approach using one of the newer shorter regimens is recommended for MDR-TB [62]. This approach is likely to improve safety, tolerability and adherence, improving the patients’ outcomes [67]. The specific regimen may be chosen on a standardized or empirical basis and then switched to individualized therapy after results of DST become available [36]. Drugs used in the treatment of MDR and XDR-TB are less efficacious and less well tolerated when used for prolonged periods, leading to a high risk of toxic effects. The treatment in these cases requires combination regimens.

• – MDR-TB cases not resistant to and/or never treated with second-line anti-TB drugs: WHO recommends a new short regimen consisting of 4–6 months of kanamycin, high dose Koxifloxacin, Prothionamide, Clofazimine, Pyrazinamide, high dose INH and EMB followed by five months of high dose Moxifloxacin, Clofazimine, Pyrazinamide and EMB. This shorter regimen (9–12 months) has produced excellent outcomes in recent studies leading to relapse-free success in near 90% of patients treated, [63, 64] much higher than the global success of the conventional MDR-TB regimen (52%). The strongest risk factor for an unfavorable outcome is a high level of resistance to fluoroquinolone, particularly when compounded by initial Pyrazinamide resistance [71].
• – Multidrug resistance TB or rifampicin-resistant: MDR-TB treatment is recommended for all patients with RIF-resistant tuberculosis, regardless of whether isoniazid resistance is confirmed.
• – When the shorter regimens are not applicable, treatment should include at least five effective drugs during the intensive period. These drugs may include Pyrazinamide and four second line core drugs (one from group A; one from group B; and at least two from group C). If this regimen cannot be followed due to problems with tolerance, contraindication or drug availability, then the total of five drugs must be complemented by a group D2 or D3 drug. This regimen must be strengthened with high-dose INH and/or EMB.
• – If INH resistance is absent or unknown + RIF resistance, all patients (child or adult) must follow the previous regimens to which INH is added, but it should not be counted among the four active drugs.
• – Surgical interventions in patients with MDR-TB: elective partial lung resection (lobectomy or wedge resection) may be used alongside a recommended MDR-TB regimen.