Bronchoalveolar Lavage Enzyme-linked Immunospot for a Rapid Diagnosis of Tuberculosis

Abstract

Rationale: The rapid diagnosis of pulmonary tuberculosis (TB) is difficult when acid fast bacilli (AFB) cannot be detected in sputum smears. Objectives: Following a proof of principle study, we examined in routine clinical practice whether individuals with sputum AFB smearnegative TB can be discriminated from those with latent TB infection by local immunodiagnosis with a Mycobacterium tuberculosis-specific enzyme-linked immunospot (ELISpot) assay. Methods: Subjects suspected of having active TB who were unable to produce sputum or with AFB-negative sputum smears were prospectively enrolled at Tuberculosis Network European Trialsgroup centers in Europe. ELISpot with early-secretory-antigenic-target-6 and culture-filtrate-protein-10 peptides was performed on peripheral blood mononuclear cells (PBMCs) and bronchoalveolar lavage mononuclear cells (BALMCs). M. tuberculosis-specific nucleic acid amplification (NAAT) was performed on bronchoalveolar lavage fluid. Measurements and Main Results: Seventy-one of 347 (20.4%) patients had active TB. Out of 276 patients who had an alternative diagnosis, 127 (46.0%) were considered to be latently infected with M. tuberculosis by a positive PBMC ELISpot result. The sensitivity and specificity of BALMC ELISpot for the diagnosis of active pulmonary TB were 91 and 80%, respectively. The BALMC ELISpot (diagnostic odds ratio [OR], 40.4) was superior to PBMC ELISpot (OR, 10.0), tuberculin skin test (OR, 7.8), and M. tuberculosis specific NAAT (OR, 12.4) to diagnose sputum AFB smear-negative TB. In contrast to PBMC ELISpot and tuberculin skin test, the BALMC ELISpot was not influenced by previous history of TB. Conclusions: Bronchoalveolar lavage ELISpot is an important advancement to rapidly distinguish sputum AFB smear-negative TB from latent TB infection in routine clinical practice.Tuberculosis (TB) is among the leading causes of morbidity and mortality worldwide (1). Pulmonary TB is the major manifestation of the disease (2). Despite constant diagnostic improvements, the rapid diagnosis of pulmonary TB is still difficult in a substantial proportion of cases (3). Identification of Mycobacterium tuberculosis by culture is the diagnostic gold standard for active TB, but culture growth of M. tuberculosis may take 2 or more weeks on average (4), and its sensitivity is only approximately 80% (5).Microscopy for the identification of acid-fast bacilli (AFB) is rapid and inexpensive (6), but AFB are undetectable from the sputum smear in 85 to 90% of children (7) and in approximately 50% of adults (8) with active pulmonary TB. In these cases, the decision to initiate anti-TB treatment can be difficult, especially because sensitivity estimates for the nucleic acid amplification technique (NAAT) to detect nucleic acids of M. tuberculosis from respiratory specimen are too variable and too low to be used to exclude the diagnosis of TB (9).If combined test results are negative, immunodiagnosis by peripheral blood IFN-g release assays (IGRAs) and tuberculin skin testing (TST) may be used as rule...

Interferon-γ release assays for the diagnosis of latentMycobacterium tuberculosisinfection: a systematic review and meta-analysis

“…In nine of these, TST positivity was significantly associated with BCG vaccination as an independent predictor in multivariate analysis [47,50,55,62,67,71,72,74,77] with odd ratios ranging between 3.8 (95% CI 1.0-13.9) [50] [35] GOLETTI [36] JAFARI [37] JAFARI [38] KAMPMANN [39] KANG [40] LEE [41] MARKOVA [42] WANG [43] DETJEN [26] DHEDA [34] GOLETTI [36] KAMPMANN [39] KANG [40] MARKOVA [42] odds for test positivity with M. tuberculosis exposure gradients, or using chest radiography lesions as a surrogate of prior exposure, the IGRAs associated better with exposure than the TST, irrespective of the setting's disease burden. Furthermore, there was generally a poor agreement between IGRAs and TST results (for k statistics, see table 3).…”

“…NPV in patients with active tuberculosis 18 articles satisfied our inclusion criteria for evaluating the NPV; 13 assessed the NPV among confirmed tuberculosis cases [22,26,[34][35][36][37][38][39][40][41][42][43][44] and six assessed the prospective outcome in IGRA-negative individuals after an average of 2 yrs [19-22, 45, 46]. Among the studies in which the NPV was evaluated in patients with confirmed tuberculosis (387 tuberculosis cases with a valid T-SPOT1.TB result and 304 with valid QFT-G-IT result), the NPV varied greatly, irrespective of the IGRA used.…”

The risk of tuberculosis in transplant candidates and recipients: a TBNET consensus statement

“…New diagnostic approaches are needed that allow for a more targeted identification of patients at risk to develop TB. This may involve modifications of in vitro immunodiagnostic assays such as the use of novel stimulatory antigens [15,164,165], alternative biomarkers other than IFN-c [166][167][168][169][170][171][172], variations in incubation time [173,174], the readout system [9,12,13,175,176] or the clinical specimen instead of blood [177][178][179]. In addition, both for the detection of LTBI with a risk of reactivation and for suspected active TB, a novel approach based on a whole blood transcriptional signature could provide a biomarker system with high discriminative potential [180].…”

Diagnostic value of nucleic acid amplification tests on bronchoalveolar lavage fluid for smear-negative pulmonary tuberculosis: a meta-analysis

“…After a systematic literature search and selection, a total of nine studies with 1214 subjects were included in this meta-analysis [15][16][17][18][19][20][21][22][23]. The article selection process used in the present study is summarized in Figure 1.…”

“…There were 357 patients with SNPT and 857 controls. Seven studies performed in Asia [15][16][17][19][20][21][22] and two studies performed in Europe [18,23]. All studies provided the detailed diagnostic criteria for SNPT, including bacteriology, histopathology or clinical diagnosis, which was widely accepted in Flow chart of selection process for eligible articles studies with tuberculosis diagnosis.…”