Xpert MTB/RIF Assay for Diagnosis of Pleural Tuberculosis

Abstract

We prospectively investigated the diagnostic utility of the Xpert MTB/RIF (Mycobacterium tuberculosis/ rifampin [RIF] resistance) assay in 20 cases with confirmed tuberculous pleural effusion. The sensitivity and specificity of the Xpert assay in pleural fluid were 25% and 100%, respectively. All cases positive by the Xpert assay were also positive by pleural fluid culture.The detection of Mycobacterium tuberculosis in sputum samples from suspected pulmonary tuberculosis (TB) patients has been significantly improved by the Xpert MTB/RIF (Mycobacterium tuberculosis/rifampin [RIF] resistance) assay (Xpert; Cepheid, Sunnyvale, CA). This automated system employs real-time PCR and molecular beacon probes to determine the presence of M. tuberculosis complex DNA, as well as rpoB gene mutations conferring rifampin resistance, rapidly and with high accuracy (1, 2, 7). The assay seems predestined for use with other specimens than sputum, such as cerebrospinal fluid or effusions from various sites, but data are still scarce. Pleural TB is the second most frequent form of extrapulmonary TB and the most frequent cause of exudative pleural effusions in areas with a high prevalence of HIV infection. Patients present at all ages with uni-or bilateral pleural effusion and acute to subacute onset of chest pain, fever, weight loss, breathlessness, and cough. The diagnostic workup includes pleural fluid aspiration and pleural biopsy, as pleural fluid smear and culture are often negative due to the paucibacillary nature of pleural TB (9). We investigated the utility of the novel Xpert MTB/ RIF assay for diagnosing pleural TB from pleural fluid.Our prospective series included 25 consecutive patients referred to Tygerberg Hospital, a tertiary medical institution in Cape Town, South Africa, with an undiagnosed pleural effusion and high clinical suspicion of pleural TB for sputum collection, pleural biopsy, and fluid aspiration. Sputum was collected from patients able to produce samples spontaneously and processed with routine methods for smear and culture. Abrams needle biopsy specimens were conducted and the specimens analyzed as published previously (4); specimens were considered positive when granulomatous inflammation with or without necrosis and/or acid-fast bacilli (AFB) were found by Ziehl-Neelsen (ZN) microscopy or when M. tuberculosis was cultured from them. A minimum of 160 ml pleural fluid was collected from each patient and split for two mycobacterial cultures and one Xpert measurement. The samples for culture were centrifuged for 20 min at 3,000 ϫ g and 4°C, and the pellets were decontaminated for 20 min with an equal volume of 2.5% sodium hydroxide-N-acetyl-L-cysteine (NaOH-NALC; Merck, Cape Town, South Africa) and neutralized with sterile phosphate-buffered saline (PBS, pH 6.8) added to a total volume of 40 ml. A second centrifugation step followed (3,000 ϫ g at 4°C for 20 min). The remaining pellets were resuspended in PBS, the bacterial concentrations were titrated to 0.5 McFarland standard, and 0.5 ml of this mixture ...

Performance of Xpert MTB/RIF on pleural tissue for the diagnosis of pleural tuberculosis: Table 1–

“…Culture, however, is limited in its ability to detect EPTB and the comparison to only cultureconfirmed cases is likely to overestimate the sensitivity of Xpert. Nonetheless, in our study Xpert missed all cases with positive cultures on tissue.The low sensitivity in pleural fluid observed in this study has been described in previous studies [5][6][7]10]. Explanations for the limited sensitivity of Xpert in pleural fluid and tissue samples could relate to PCR inhibitors or insufficient sample volume in this paucibacillary disease.…”

“…The low sensitivity in pleural fluid observed in this study has been described in previous studies [5][6][7]10]. Explanations for the limited sensitivity of Xpert in pleural fluid and tissue samples could relate to PCR inhibitors or insufficient sample volume in this paucibacillary disease.…”

“…Given the limitations of available tests for the diagnosis of pleural TB, several studies have evaluated the performance of Xpert using pleural fluid as a sample type. Overall, these studies show limited accuracy with sensitivity averaging around 44% [5][6][7]. However, the preferred specimen for the diagnosis of pleural TB is pleural tissue.…”

“…Pleural fluid was centrifuged (1370 6g for 15 min) and the concentrated sediment, resuspended in 1 mL of the original supernatant, was used for Xpert [5]. Pleural tissue was finely ground and re-suspended in 1 mL of sterile saline [8].…”

NAAT: A New Ray of Hope in the Early Diagnosis of EPTB

“…If fresh sample is used then a slightly higher sensitivity and a lower specificity is observed than frozen samples. On the other hand, lower sensitivities have been observed for pleural, peritoneal, cerebrospinal, pericardial, and synovial fluid samples [8,9].…”