Diagnostic Performance of Pleural Fluid Adenosine Deaminase for Tuberculous Pleural Effusion in a Low-Incidence Setting.

The challenges associated with diagnosing tuberculous pleural effusion have led to the use of pleural fluid adenosine deaminase (pfADA) as a biomarker for Mycobacterium tuberculosis infection. This study retrospectively reviewed the diagnostic performance of pfADA, the pleural fluid lactate dehydrogenase (LD)/ADA ratio, and combinations of these two parameters in 1,637 episodes of pleural effusion in the low-tuberculosis (TB)-incidence setting of Auckland, Aotearoa New Zealand, from between March 2008 and November 2014. The median pfADA in 57 TB pleural effusion episodes (58.1 U/liter) was significantly higher (P < 0.001) than in 1,580 non-TB pleural effusions (11.4 U/liter). The median LD/ADA ratio in TB pleural effusion (8.2) was significantly lower (P < 0.001) than in non-TB pleural effusions (30.5). The pfADA and pleural fluid LD/ADA ratio AUCROC values (that is, receiver operating characteristic [ROC] curve analysis with determination of the ROC area under the curve) were 0.93 and 0.94, respectively. The pfADA thresholds of ≥15 and ≥30 U/liter demonstrated sensitivities of 100% (95% confidence internal = 93.7 to 100) and 93.0% (83.3 to 97.2), specificities of 62.7% (60.3 to 65.0) and 87.3% (85.6 to 88.9), positive predictive values (PPVs) of 8.8% (6.9 to 11.2) and 20.9% (16.4 to 26.4), and negative predictive values (NPVs) of 100% (99.6 to 100) and 99.7% (99.3 to 99.9), respectively. LD/ADA ratio thresholds of <25 and <15 demonstrated sensitivities of 100% (93.5 to 100) and 89.1% (78.2 to 94.9), specificities of 61.6% (59.1 to 64.0) and 84.8% (82.9 to 86.5), PPVs of 8.5% (6.6 to 10.9) and 17.3% (13.3 to 22.0), and NPVs of 100% (99.6 to 100) and 99.5% (99.0 to 99.8), respectively. A combination of pfADA ≥ 30 U/liter and an LD/ADA ratio < 15 increased the specificity and PPV to 97.8% (96.9 to 98.4) and 57.3% (46.5 to 67.5) but decreased the sensitivity to 85.5% (73.8 to 92.4). The primary value of pfADA in a low-TB-incidence setting, such as Auckland, is in utilization of its high NPV.

A case of pleural Mycobacterium tuberculosis infection with reversion of Quantiferon Gold Plus results from positive to negative.

Introduction. Mycobacterium tuberculosis (MTB) infections continue to have a high mortality and morbidity burden globally. Interferon-gamma release assays such as Quantiferon Gold Plus (QFG-Plus) aid in diagnosis of latent TB but diagnosis of pleural TB remains challenging. We present a case of active pleural MTB infection with reversion from positive to negative of IGRA result as well as negative Xpert MTB/RIF Ultra PCR result from tissues obtained from pleural biopsy. Case summary. A 52-year-old otherwise healthy male presented in August 2022 with a 2 week history of pleuritic chest pain associated with modest elevation in inflammatory markers. The patient had had a positive QFG-Plus result in 2018, however QFG-Plus during this admission was negative. Computed-tomography pulmonary angiogram and needle thoracocentesis showed an exudative left pleural effusion with predominant lymphocytes. The patient's symptoms failed to resolve with empiric antimicrobial therapy for community-acquired pneumonia. Broncho-alveolar lavage as well as biopsies of pleural tissues via video-assisted thoracoscopic surgery from the left lower lobe yielded negative results on routine microbiological culture as well as Xpert Ultra PCR. Growth of acid-fast bacilli was noted from mycobacterial cultures of pleural tissues which was identified as MTB. Conclusion. Despite significant technological advances, microbiological diagnosis of MTB infections remains challenging. We document QFG-Plus reversion during development from latent to active pleural TB. Decline in the ability of CD4+ and CD8+ T cells to produce interferon gamma in response to TB antigens (ESAT-6 and CFP-10) was likely associated with loss of host control of latent MTB. This case serves as a reminder that despite exhaustive testing with state-of-art diagnostic platforms, MTB infections can still elude discovery.

Tuberculous chylothorax in paediatric population-a case report and systematic review of the literature.

Background: Chylothorax as part of the clinical spectrum of tuberculosis (TB) is a rare entity, especially among children. However, it is crucial for clinicians to be able to identify, correlate, and diagnose chylothorax as it poses significant morbidity to patients. Case Description: We report on a paediatric case of pleural TB complicated with complex chylothorax, and systematically reviewing the literature for cases of tuberculous chylothorax among children particularly looking at the (I) demographic, (II) clinical presentations, (III) radiological findings, and (IV) investigations among this cohort of patients. The systematic review complied to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. We used three separate databases, namely PubMed, Ovid, and Scopus to search for articles with terms "chylothorax", "tuberculosis", and "children". We included article reporting on: (I) children aged below 18; (II) infected by Mycobacterium TB, and (III) written in English only. We reviewed the publications from the inception up to August 2022. After an extensive search, 7 articles were reviewed. We included 10 patients from all the reports. Most common symptoms reported were intermittent fever (n=6) and chronic cough (n=5). Other symptoms include shortness of breath and chest discomfort due to fluid accumulation and/or effusion. Typical TB symptoms like night sweat, loss of weight, and loss of appetite were present in some of the reported patients. Chest radiography was the most used diagnostic imaging modality, in which all of the reported cases noted presence of pleural effusion, with some cases proceeded with computer tomography (CT) of thorax to aid in the diagnosis. Most of the patients had lymphadenopathies and all patients underwent pleural tapping for analysis and symptomatic relief. From the pleural fluid milky appearance and biochemistry correlation, the diagnosis of chylothorax was established in all 10 patients. Every patient was tested positive for TB infection. Conclusions: Most of the cases showed similar clinical presentation, radiological findings, and laboratory investigations particularly the classic analysis findings of pleural fluid. Even though TB is not a common cause of chylothorax, clinicians should have raised suspicion of it especially after gathering full history and correlating it with the other findings.

Diagnosis of pleural tuberculosis by multi-targeted loop-mediated isothermal amplification assay based on SYBR Green I reaction: comparison with GeneXpert® MTB/RIF assay.

BACKGROUND: Diagnosis of pleural tuberculosis (TB) is tedious owing to its close resemblance with malignant pleural effusion and sparse bacterial load in clinical specimens. There is an immediate need to design a rapid and dependable diagnostic test to prevent unnecessary morbidity/mortality. RESEARCH DESIGN AND METHODS: A multi-targeted loop-mediated isothermal amplification (MT-LAMP) was deliberated using mpt64 and IS6110 to diagnose pleural TB within pleural fluids/biopsies. MT-LAMP products were analyzed by gel-based and visual detection methods, viz. SYBR Green I, SYBR Green I+deoxyuridine triphosphate uracil-N-glycosylase (dUTP-UNG), and dry methyl green reactions. RESULTS: In a pilot study, while assessing pleural TB/non-TB control subjects (n = 40), both SYBR Green I+dUTP-UNG/gel-based MT-LAMP assays exhibited better sensitivity/specificity than SYBR Green I and dry methyl green MT-LAMP. Since it is facile to work with SYBR Green I+dUTP-UNG than gel-based MT-LAMP, we validated the performance of SYBR Green I+dUTP-UNG in a higher number of specimens (n = 97), which revealed somewhat higher sensitivity (85.2 vs. 81.5%) and specificity (97.7 vs. 90.7%) than SYBR Green I MT-LAMP. Furthermore, the sensitivity attained by SYBR Green I+dUTP-UNG MT-LAMP was significantly higher (p < 0.001) than GeneXpert. CONCLUSIONS: Our SYBR Green I+dUTP-UNG MT-LAMP is a simple and reliable method to diagnose pleural TB, which may translate into a point-of-care test.

Insight into diagnosis of pleural tuberculosis with special focus on nucleic acid amplification tests.

INTRODUCTION: Pleural tuberculosis (TB) is the archetype of extrapulmonary TB (EPTB), which mainly affects the pleural space and leads to exudative pleural effusion. Diagnosis of pleural TB is a difficult task predominantly due to atypical clinical presentations and sparse bacillary load in clinical specimens. AREA COVERED: We reviewed the current literature on the globally existing conventional/latest modalities for diagnosing pleural TB. Bacteriological examination (smear/culture), tuberculin skin testing/interferon-γ release assays, biochemical testing, imaging and histopathological/cytological examination are the main modalities. Moreover, nucleic acid amplification tests (NAATs), i.e. loop-mediated isothermal amplification, PCR/multiplex-PCR, nested-PCR, real-time PCR and GeneXpert® MTB/RIF are being utilized. Currently, GeneXpert Ultra, Truenat MTBTM, detection of circulating Mycobacterium tuberculosis (Mtb) cell-free DNA by NAATs, aptamer-linked immobilized sorbent assay and immuno-PCR (I-PCR) have also been exploited. EXPERT OPINION: Routine tests are not adequate for effective pleural TB diagnosis. The latest molecular/immunological tests as discussed above, and the other tools, i.e. real-time I-PCR/nanoparticle-based I-PCR and identification of Mtb biomarkers within urinary/serum extracellular vesicles being utilized for pulmonary TB and other EPTB types may also be explored to diagnose pleural TB. Reliable diagnosis and early therapy would reduce the serious complications associated with pleural TB, i.e. TB empyema, pleural fibrosis, etc.

Epidemiology of Adult Pleural Disease in the United States.

BACKGROUND: Comprehensive US epidemiologic data for adult pleural disease are not available. RESEARCH QUESTION: What are the epidemiologic measures related to adult pleural disease in the United States? STUDY DESIGN AND METHODS: Retrospective cohort study using Healthcare Utilization Project databases (2007-2016). Adults (≥ 18 years of age) with malignant pleural mesothelioma, malignant pleural effusion, nonmalignant pleural effusion, empyema, primary and secondary spontaneous pneumothorax, iatrogenic pneumothorax, and pleural TB were studied. RESULTS: In 2016, ED treat-and-discharge (T&D) visits totaled 42,215, accounting for charges of $286.7 million. In 2016, a total of 361,270 hospitalizations occurred, resulting in national costs of $10.1 billion. A total of 64,174 readmissions contributed $1.16 billion in additional national costs. Nonmalignant pleural effusion constituted 85.5% of ED T&D visits, 63.5% of hospitalizations, and 66.3% of 30-day readmissions. Contemporary sex distribution (male to female ratio) in primary spontaneous pneumothorax (2.1:1) differs from older estimates (6.2:1). Decadal analyses of annual hospitalization rates/100,000 adult population (2007 vs 2016) showed a significant (P < .001) decrease for malignant pleural mesothelioma (1.3 vs 1.09, respectively), malignant pleural effusion (33.4 vs 31.9, respectively), iatrogenic pneumothorax (17.9 vs 13.9, respectively), and pleural TB (0.20 vs 0.09, respectively) and an increase for empyema (8.1 vs 11.1, respectively) and nonmalignant pleural effusion (78.1 vs 100.1, respectively). Empyema hospitalizations have high costs per case ($38,591) and length of stay (13.8 days). The mean proportion of readmissions attributed to a pleural cause varied widely: malignant pleural mesothelioma, 49%; malignant pleural effusion, 45%; nonmalignant pleural effusion, 31%; empyema, 27%; primary spontaneous pneumothorax, 27%; secondary spontaneous pneumothorax, 27%; and iatrogenic pneumothorax, 20%. Secondary spontaneous pneumothorax had the shortest time to readmission in 2016 (10.3 days, 95% CI, 8.8-11.8 days). INTERPRETATION: Significant epidemiologic trends and changes in various pleural diseases were observed. The analysis identifies multiple opportunities for improvement in management of pleural diseases.

Hiding in plain sight: Diagnosing pleural tuberculosis using lung ultrasound.

INTRODUCTION: Diagnosing pleural tuberculosis can be difficult in patients with ambiguous presentation, especially in resource-limited health centres. Thus, lung ultrasound had been studied as a novel method in helping clinicians to diagnose this condition. CASE PRESENTATION: A 48-year-old woman presented with worsening dyspnoea and orthopnoea for one week. She had also experienced weight loss, minimal dry cough and right-sided pleuritic chest pain for several weeks. A chest radiograph showed a right lower zone pleural effusion with no apparent lung consolidation. Lung ultrasound showed a right apical consolidation and right lower zone septated pleural effusion. Pleural fluid investigations showed exudative features of mixed lymphocytic, mesothelial and neutrophilic cellular components. Tuberculin skin test was strongly positive. She was subsequently treated for pleural tuberculosis. One month after treatment, her symptoms had improved considerably. DISCUSSION: Lung ultrasound has been found to be more effective than chest radiograph in detecting consolidation and diagnosing pneumonia. The portability and efficacy of today's ultrasound machines, including the handheld types, show that lung ultrasound is a practical, reliable and valuable diagnostic tool in managing pulmonary conditions including tuberculosis, provided that the operators are adequately trained. CONCLUSION: Lung ultrasound in tuberculosis is the next frontier for clinicians and researchers.

Clinical characteristics that portend a positive Xpert Ultra test result in patients with pleural tuberculosis.

BACKGROUND: The performance of Xpert-MTB/RIF, including the newer Xpert Ultra test, for the diagnosis of pleural TB is poor (~28 - 38%). There are no data on patient characteristics that portend a positive Xpert-Ultra test in pleural fluid. These characteristics could be useful for selecting patients for Xpert-Ultra testing, thus maximising benefits of a positive test, while minimising cost. OBJECTIVES: To determine the clinical, radiological, microbiological and biochemical characteristics associated with Xpert-Ultra positivity in patients with suspected pleural tuberculosis. METHODS: We performed a subgroup analysis of a prospective observational cohort (N=165 patients with suspected pleural TB) evaluating same-day diagnostic tools for pleural tuberculosis. Forty-nine patients with confirmed pleural tuberculosis (culture and/or histology) were included in the final analysis. RESULTS: Of the 49 participants, 17 (35%) were female and 9 (18.4%) were HIV-infected. In the multivariate analysis including demographic, radiological and pleural fluid test characteristics, there were no independent predictors of Xpert-Ultra positivity. However, when pleural fluid test results were excluded, and when only rapidly ascertainable pre-test factors (demographic and radiologic variables) were considered, the multivariable analysis showed that only a chest X-ray (CXR) suggestive of active TB (cavities, consolidation and hilar lymphadenopathy) was associated with Xpert-Ultra positivity (p=0.021). Notably, only 22% (n=11/49) of the participants had a CXR suggestive of active TB and of these, 73% (n=8/11) had a positive Xpert-Ultra result. CONCLUSION: CXR features suggestive of active TB are significantly associated with a positive Xpert-Ultra test result on pleural fluid. These data inform clinical practice in resource-poor settings.

Comparative evaluation of GeneXpert MTB/RIF and multiplex PCR targeting mpb64 and IS6110 for the diagnosis of pleural TB.

AIM: Diagnosis of pleural TB poses serious challenges due to paucibacillary nature of specimens and there is an urgent need to devise a reliable diagnostic test. METHODS: We compared GeneXpert Mycobacterium tuberculosis/rifampin assay and the multiplex PCR (M-PCR) targeting mpb64 (Rv1980c) and IS6110 in pleural fluids (n = 78) of pleural TB patients and non-TB controls. RESULTS: The sensitivities of 89.6 and 33.3%, and specificities of 96.7 and 100%, were observed with M-PCR and Xpert assay, respectively. CONCLUSION: M-PCR showed superiority over Xpert assay and may facilitate an efficient diagnosis of pleural TB.

Use of lateral flow assays to determine IP-10 and CCL4 levels in pleural effusions and whole blood for TB diagnosis.

One of the key problems in combating TB is the lack of fast and accurate diagnostic tests that are affordable and easy to use in resource-limited settings. We have used a field-friendly up-converting phosphor (UCP) reporter technology in a lateral flow (LF) based test for the diagnosis of respiratory infections. In this study we analysed samples obtained from patients presenting with symptoms suggestive of TB but prior to confirmation by microbiology in The Gambia. Following clinical and microbiological evaluation they were classified as either having TB or other respiratory disorder (ORD). Analysis of blood was performed for those with pulmonary TB and pleural fluid for those with pleural TB. UCP-LF test for detection and quantitation of IP-10 and CCL4 were used being the two chemokine markers that have been shown to increase in active TB disease. UCP-LF test accurately determined concentrations of both markers as compared to ELISA and multiplex cytokine array. However, only IP-10 could discriminate between TB and ORD, and this was significantly enhanced by analysing the site of infection (pleural fluid), which showed 92% correct classification. Future work will assess the use of multiple markers to increase diagnostic accuracy.

Expansion and productive HIV-1 infection of Foxp3 positive CD4 T cells at pleural sites of HIV/TB co-infection.

BACKGROUND: CD4 T-cells expressing Foxp3 are expanded systemically during active tuberculosis (TB) regardless of HIV-1 co-infection. Foxp3+ CD4 T cells are targets of HIV-1 infection. However, expansion of HIV-1 infected Foxp3+ CD4 T cells at sites of HIV/TB co-infection, and whether they contribute to promotion of HIV-1 viral activity is not known. METHODS: Pleural fluid mononuclear cells (PFMC) from HIV/TB co-infected patients with pleural TB were characterized by immune-staining and FACS analysis for surface markers CD4, CD127, CCR5, CXCR4, HLA-DR and intracellular expression of Foxp3, HIVp24, IFN-γ and Bcl-2. Whole PFMC and bead separated CD4+CD25+CD127- T cells were assessed for HIV-1 LTR strong stop (SS) DNA by real-time PCR, which represents viral DNA post cell entry and initiation of reverse transcription. RESULTS: High numbers of HIV-1 p24 positive Foxp3+ and Foxp3+CD127- CD4 T cells were identified in PFMC from HIV/TB co-infected subjects. CD4+Foxp3+CD127- T cells displayed high expression of the cellular activation marker, HLA-DR. Further, expression of the HIV-1 co-receptors, CCR5 and CXCR4, were higher on CD4+Foxp3+T cells compared to CD4+Foxp3- T cells. Purified CD4+CD25+CD127- T cells isolated from PFMC of HIV/TB co-infected patients, were over 90% CD4+Foxp3+T cells, and exhibited higher HIV-1 SS DNA as compared to whole PFMC, and as compared to CD4+CD25+CD127- T cells from an HIV-infected subject with pleural mesothelioma. HIV-1 p24+ Foxp3+ CD4+T cells from HIV/TB patients higher in Bcl-2 expression as compared to both HIV-1 p24+ Foxp3- CD4 T cells, and Foxp3+ CD4+T cells without HIV-p24 expression. CONCLUSION: Foxp3+ CD4 T cells in PFMC from HIV/TB co-infected subjects are predisposed to productive HIV-1 infection and have survival advantage as compared to Foxp3 negative CD4 T cells.

Comparison between Diagnostic Yield of Pleural Fluid ELISPOT and Thoracoscopic Guided Pleural Biopsy in Undiagnosed Pleural Effusion

Context: Despite recent advances in the available diagnostic modalities, diagnosis of pleural tuberculosis remains a challenge because of the low yield of conventional methods. Pleural biopsy is the gold standard for confirmation of diagnosis, which is invasive and cumbersome. The concentration of mycobacterial peptide-specific activated lymphocytes at the site of infection can be utilized as the basis for using IGRA (interferon-gamma release assays) based evaluation of undiagnosed exudative pleural effusions.  Aim: To evaluate the performance of IGRA (Enzyme-linked Immunospot (ELISPOT) in pleural fluid for the diagnosis of pleural tuberculosis in histopathologically confirmed cases. Settings and Design: A prospective observational study compared the utility of ELISPOT with thoracoscopy guided pleural biopsies for the diagnosis of tubercular pleural effusions. Methods and Material: Forty-two consecutive cases of undiagnosed pleural effusions were enrolled and subjected to thoracoscopy guided pleural biopsy. Thirteen patients were confirmed to have tuberculosis, 27 had malignancy, and 2 had normal pleura. A total of 1x103 pleural fluid mononuclear cells (PFMCs) were cultured in the presence of early secretory antigenic target-6 (ESAT-6) and culture filtrate protein-10 (CFP-10) for 24 hours. The individual spots were then counted using an automated analyzer ELISPOT reader system.  Results: The number of spots developed in the pleural fluid was significantly higher in tubercular pleural effusions as compared to non-tubercular effusions (CFP-10:154.76±14.61 vs 49.24±8.9; ESAT-6: 150.3±17.27 v/s 45.34±8.23, p<0.001). At a cut-off value of more than 67 spots taken as positive for tuberculosis, the sensitivity of the test was 100% (95% CI 75.29% to 100.00%), specificity was 96.5% (95 % CI 82.24% to 99.91%), positive predictive value was 92.86% (95 % CI 65.45% to 98.89%) and negative predictive value was 100%.  Conclusions: ELISPOT can be a useful non-invasive test for the evaluation of undiagnosed pleural effusions and making a diagnosis of pleural tuberculosis with confidence.