Rapid Diagnosis of Smear-negative Tuberculosis by Bronchoalveolar Lavage Enzyme-linked Immunospot

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Abstract
Rationale: In a large proportion of patients with active pulmonary tuberculosis (pTB), acid-fast bacilli smear results for sputum and bronchial secretions are negative. Detectable growth of Mycobacterium tuberculosis (MTB) in cultures takes several weeks and MTB-specific DNA amplification results on sputum and bronchial secretions are variable in these patients. Objective: We investigated whether a rapid diagnosis of pTB can be established by enumeration of MTB-specific mononuclear cells from bronchoalveolar lavage (BAL) fluid in routine clinical practice. Methods: Patients presenting to a tertiary hospital with medical histories and pulmonary infiltrates compatible with tuberculosis, and negative acid-fast bacilli smear results (three) from sputum, were prospectively enrolled in this study. An MTB-specific enzyme-linked immunospot assay (ELISPOT [T-SPOT.TB; Oxford Immunotec, Abingdon, UK]) with early antigenic target-6 (ESAT-6) and culture filtrate protein-10 (CFP-10) peptides was performed on peripheral blood mononuclear cells (PBMCs) and mononuclear cells from the BAL fluid (BALMCs). Measurements and Main Results: Of 37 patients, 12 were found to have smear-negative pTB and 25 were found to have an alternative diagnosis. Patients with tuberculosis had a median number of 17 ESAT-6–specific cells and 24.5 CFP-10–specific cells per 200,000 PBMCs and 37.5 ESAT-6–specific cells and 49.5 CFP-10–specific cells per 200,000 cells in the BAL fluid. Control patients had a median of 1 ESAT-6–specific cell and 1 CFP-10–specific cell per 200,000 PBMCs and no ESAT-6– and CFP-10–specific cells per 200,000 cells in the BAL fluid (p < 0.0001). All patients with TB but none of the control subjects had more than 5 spot-forming cells per 200,000 BALMCs with either peptide in the BAL fluid ELISPOT. Conclusion: Smear-negative pulmonary tuberculosis can be diagnosed rapidly by identification of MTB-specific cells in the BAL fluid.

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