Development of DNA Aptamers to Visualize Release of Mycobacterial Membrane-Derived Extracellular Vesicles in Infected Macrophages

Extracellular vesicles (EVs) have emerged into a novel vaccine platform, a biomarker and a nano-carrier for approved drugs. Their accurate detection and visualization are central to their utility in varied biomedical fields. Owing to the limitations of fluorescent dyes and antibodies, here, we describe DNA aptamer as a promising tool for visualizing mycobacterial EVs in vitro. Employing SELEX from a large DNA aptamer library, we identified a best-performing aptamer that is highly specific and binds at nanomolar affinity to EVs derived from three diverse mycobacterial strains (pathogenic, attenuated and avirulent). Confocal microscopy revealed that this aptamer was not only bound to in vitro-enriched mycobacterial EVs but also detected EVs that were internalized by THP-1 macrophages and released by infecting mycobacteria. To the best of our knowledge, this is the first study that detects EVs released by mycobacteria during infection in host macrophages. Within 4 h, most released mycobacterial EVs spread to other parts of the host cell. We predict that this tool will soon hold huge potential in not only delineating mycobacterial EVs-driven pathogenic functions but also in harboring immense propensity to act as a non-invasive diagnostic tool against tuberculosis in general, and extra-pulmonary tuberculosis in particular.

Accurate Integrated System to detect Pulmonary and Extra Pulmonary Tuberculosis using Machine Learning Algorithms

Several studies have been reported the use of machine learning algorithms in the detection of Tuberculosis, but studies that discuss the detection of both types of TB, i.e., Pulmonary and Extra Pulmonary Tuberculosis, using machine learning algorithms are lacking. Therefore, an integrated system based on machine learning models has been proposed in this paper to assist doctors and radiologists in interpreting patients’ data to detect of PTB and EPTB. Three basic machine learning algorithms, Decision Tree, Naïve Bayes, SVM, have been used to predict and compare their performance. The clinical data and the image data are used as input to the models and these datasets have been collected from various hospitals of Jalandhar, Punjab, India. The dataset used to train the model comprises 200 patients’ data containing 90 PTB patients, 67 EPTB patients, and 43 patients having NO TB. The validation dataset contains 49 patients, which exhibited the best accuracy of 95% for classifying PTB and EPTB using Decision Tree, a machine learning algorithm.

COMPARISON OF CLINICO-SOCIAL PROFILE OF PULMONARY AND EXTRA PULMONARY TUBERCULOSIS IN TERTIARY CARE HOSPITAL IN A RURALAREA: A RETROSPECTIVE STUDY

BACKGROUND AND OBJECTIVES: The burden of tuberculosis (TB) in India is the highest accounting for 26% of the global incidence. A total of 1.4 million people died from TB in 2019 (including 208 000 people with HIV). India accounts for a fourth of the global burden of TB and 29% of global mortality. Therefore, we carried out this study to compare demographic, lifestyle and clinical characteristic between pulmonary TB (PTB) and extrapulmonary TB (EPTB). MATERIALS AND METHODOLOGY:Aretrospective analysis was carried of 348 patients diagnosed in DOTS centre, Pravara Rural Hospital, Loni. Characteristics of demographic and clinical characteristics were obtained from medical case records. RESULTS:Among the 348 cases, 71.3% were PTB and 28.7% were EPTB including, pleural (36%), meningeal (27%) and lymphatic (20%) cases. The male to female ratio in PTB and EPTB are 1.99 and 1.22 respectively. EPTB was more common at younger age (<25 years). Tobacco addiction (10.9%), diabetes mellitus (4.03%), HIV positivity (12.1%) and history of contact with Tb patients (17.7%) were more likely to be associated with PTB. CONCLUSION:Increased awareness of the risk factors may facilitate early case nding and better management outcomes for these patients.

Evolving Stroke in Tuberculous Meningitis

Tuberculous meningitis (TBM) is the most severe form of extra-pulmonary tuberculosis which carries high mortality with 100% mortality without treatment. A neurological complication of TBM includes hydrocephalus, brain abscess and stroke. In this report, we would like to illustrate a case of stroke in a patient with TBM. In this case, a 37-year old man initially presented with fever for 1 week associated with severe headache and occasional vomiting. Computed tomography (CT) of the brain showed leptomeningeal enhancement and lumbar puncture findings consistent with infective in nature. His MARAIS score was 13 and was treated as tuberculous meningitis with anti-tuberculous therapy. While in the ward, he developed right-sided body weakness with evolving CT brain findings. His condition then stabilized with anti-tuberculous treatment which consists of isoniazid, rifampicin, pyrazinamide and streptomycin. Dexamethasone was also initiated. On follow up, his condition further improves and is functionally independent. In conclusion, tuberculous meningitis is an aggressive disease with high morbidity. Stroke can occur as a result of TBM. Timely initiation of treatment is important in improving the outcome of the patients.

Varied presentation of tuberculosis adenopathy: an immunological and diagnostic contemplation

Tuberculous lymphadenitis is the most common presentation of extra pulmonary tuberculosis EPTB. Clinical and radiological presentation of disease may vary and differential diagnosis includes both benign and malignant conditions such as lymphomas and sarcoidosis. We present two case reports of tuberculous lymphadenitis with varied presentation. In case report Ⅰ, patient had undergone splenectomy due to traumatic splenic rupture had classical symptoms of tuberculosis, chest X-ray, CT thorax showed mediastinal and cervical lymphadenopathy. FNAC showed a picture of granulomatous lesion. In case report ⅠⅠ patient came with complaints of abdominal pain in hypochondriac region. CT and USG abdomen showed multiple enlarged retroperitoneal lymph nodes which led to a provisional diagnosis of lymphoma after which lymph node biopsy showed a granulomatous picture. This case report stresses the importance of early and quick diagnosis of tuberculosis in immunocompromised patients and highlights the risk of misdiagnosis of tuberculous lymphadenitis with other benign and malignant causes of lymphadenopathy and the importance of humoral mediated immunity in tuberculosis.

ROLE OF GENE XPERT AND LIQUID CULTURE IN DIAGNOSIS OF EXTRA PULMONAR TUBERCULOSIS

INTRODUCTION: Extrapulmonary Tuberculosis (EPTB) accounts for 15- 25% of all TB cases. It is more difficult to diagnose than Pulmonary tuberculosis and often requires invasive procedures to obtain tissue and or fluid samples. Histology is time-consuming and establishing a diagnosis of TB with high specificity remains difficult. Tissue smear microscopy after special staining is often negative. Tissue culture often leads to considerable delays compromising patient care and outcomes. AIMS AND OBJECTIVES:1. To diagnose Extra Pulmonary Tuberculosis by Gene Xpert(Xpert MTB/Rif assay or CBNAAT) and Liquid Cultures. 2. To evaluate the Sensitivity and Specificity of Gene Xpert in Extra Pulmonary Tuberculosis in comparison with Liquid Culture MGIT960 system. MATERIALS AND METHODS: This retrospective cross-sectional study was carried out by reviewing all suspected extra pulmonary tuberculosis samples of 430 patients attending OPD at Institute of Respiratory Diseases, Jaipur from April 2020 to March 2021.The extrapulmonary samples (pleural fluid,CSF,pus,BAL,Ascitic fluid,Synovial fluid,Gastric aspirate,Liver aspirate) were subjected to GeneXpert and Liquid culture MGIT960 system. RESULTS: Of the 430 Extra Pulmonary Samples, The Sensitivity and Specificity of CBNAAT was 79.77% and 95.30% respectively in comparison with Liquid Culture. Out of the 430 Samples CBNAAT was Positive in 87 samples of which 71(81.60%) were Rifampicin sensitive and 16(18.39%) were Rifampicin Resistant.Out of the 430 Samples,Liquid cultures was Positive in 89 samples. CONCLUSION: Gene Xpert has a notable advantage of detecting tuberculosis within two hours which is acceptable to all clinicians to institute early treatment.CBNAAT is one of the rapid diagnostic tests available in the country and it should be routinely used under the public and private health sector effectively to detect early tuberculosis in Extra Pulmonary Samples.

A STUDY ON EXTRA PULMONARY TUBERCULOSIS: SOCIO ECONOMIC CLASS DISTRIBUTION

Background:Extra-pulmonary tuberculosis can affect the lymph nodes, pleura, bones, joints, genitourinary tract, nervous system (meningitis, tuberculoma), abdominal tuberculosis (intestines, mesentery, solid organs), skin. Materials and Methods: It is a record based, observational prospective study which assesses the presentation and evaluation of the socio-economic distribution of extrapulmonary tuberculosis and management of it in a tertiary care hospital. Result: In present study, out of 50 cases 52% patients were from low socio-economic class, 44% patients were from middle socioeconomic class and 4% were from high socio-economic class. Conclusion:The frequency of extrapulmonary TB in this study was highest in low socio-economic class followed by middle and high socio-economic class.