Manoj Kumar Panigrahi

Pulmonary cryptococcosis with cryptococcal meningitis in an immunocompetent host.

Cryptococcosis is a systemic fungal infection associated with significant morbidity and mortality. It predominantly affects people with immunosuppresion and human immunodeficiency virus infection. Extrapulmonary dissemination is rare in immunocompetent hosts. We present here a case of disseminated cryptococcosis in an immunocompetent patient who presented with an unusually large pulmonary mass and meningitis and successfully managed with medical therapy.

Pulmonary Mucormycosis Presenting as Nonresolving Pneumonia in a Patient With Diabetes Mellitus

Zygomycosis refers to a group of disorders caused by filamentous fungi in the class Zygomycetes. This class is subdivided into 2 orders, Mucorales and Entomophthorales, both implicated in human disease. The majority of human infections are caused by fungi in the Mucorales order; hence, the terms

A case of systemic melioidosis: unravelling the etiology of chronic unexplained fever with multiple presentations

Melioidosis, caused by the environmental saprophyte, Burkholderia pseudomallei, is an important public health problem in Southeast Asia and Northern Australia. It is being increasingly reported from other parts, including India, China, and North and South America expanding the endemic zone of the disease. We report a case of systemic melioidosis in a 58-year-old diabetic, occupationally-unexposed male patient, who presented with chronic fever, sepsis, pneumonia, pleural effusion and subcutaneous abscess, was undiagnosed for long, misidentified as Pseudomonas aeruginosa infection elsewhere, but was saved due to correct identification of the etiologic agent and timely institution of appropriate therapy at our institute. A strong clinical and microbiological suspicion for melioidosis should be considered in the differential diagnosis of acute pyrexia of unknown origin, acute respiratory distress syndrome and acute onset of sepsis, especially in the tropics.

MET Amplification and Response to MET Inhibitors in Stage IV Lung Adenocarcinoma

Background: Non-small-cell lung cancers with MET amplification may respond to c-MET inhibitors. Methods: We examined lung adenocarcinoma patients for mutations and amplification status of epidermal growth factor receptor (EGFR), anaplastic lymphoma kinase (ALK), ROS, MET. The clinical characteristics of patients with MET amplification and their responses to MET inhibitor therapy were studied. Results: Of the 76 patients analyzed, 5 were positive for c-MET gene amplification and 4 cases showed an intermediate result. For 12 patients who were EGFR positive, a c-MET analysis on secondary biopsy tissue was performed following disease progression. All 5 c-MET-positive patients were men. The age range in the study was 34-83 years. 4 of the 5 patients were started on crizotinib. 2 of these cases were positive following tyrosine kinase inhibitor therapy. 3 patients showed a response. 1 patient showed no response and was later found to have a concurrent T790M mutation. Conclusions: There are 2 categories of MET gene amplification in lung cancer patients, de novo and that secondary to TKI therapy. These patients can benefit from MET inhibitor therapy. Dual mechanisms of resistance, EGFR T790M mutation and c-MET amplification after TKI therapy, may suggest a poor prognosis.

Detection of false positive mutations in BRCA gene by next generation sequencing.

BRCA1 and BRCA2 genes are implicated in 20–25% of hereditary breast and ovarian cancers. New age sequencing platforms have revolutionized massively parallel sequencing in clinical practice by providing cost effective, rapid, and sensitive sequencing. This study critically evaluates the false positives in multiplex panels and suggests the need for careful analysis. We employed multiplex PCR based BRCA1 and BRCA2 community Panel with ion torrent PGM machine for evaluation of these mutations. Out of all 41samples analyzed for BRCA1 and BRCA2 five were found with 950_951 insA(Asn319fs) at Chr13:32906565 position and one sample with 1032_1033 insA(Asn346fs) at Chr13:32906647, both being frame-shift mutations in BRCA2 gene. 950_951 insA(Asn319fs) mutation is reported as pathogenic allele in NCBI dbSNP. On examination of IGV for all these samples, it was seen that both mutations had ‘A’ nucleotide insertion at 950, and 1032 position in exon 10 of BRCA2 gene. Sanger Sequencing did not confirm these insertions. Next-generation sequencing shows great promise by allowing rapid mutational analysis of multiple genes in human cancer but our results indicate the need for careful sequence analysis to avoid false positive results.

Quantification of DNA Extracted from Formalin Fixed Paraffin-Embeded Tissue Comparison of Three Techniques: Effect on PCR Efficiency

INTRODUCTION Mutation detection from Formalin Fixed Paraffin-Embedding (FFPE) tissue in molecular lab became a necessary tool for defining potential targeted drug. Accurate quantification of DNA extracted from FFPE tissue is necessary for downstream applications like Polymerase Chain Reaction (PCR), sequencing etc. AIM To check and define which method for FFPE DNA quantification is suitable for downstream processes. MATERIALS AND METHODS In this experimental experience study Biorad Smartspec Plus spectrophotomery, Qubit Fluorometer, and Qiagen Rotorgene qPCR was used to compare 20 FFPE DNA quantification in Rajiv Gandhi Cancer Institute and Research Centre, in 2015 and quantified amount of DNA used for PCR reaction. RESULTS The average concentration of DNA extracted from FFPE tissue measured using the spectrophotometer was much higher than the concentration measured using the Qubit Fluorometer and qPCR. CONCLUSION Results varied depending upon the technique used. A fluorometric analysis may be more suitable for quantification of DNA samples extracted from FFPE tissue compared with spectrophotometric analysis. But qPCR is the best technique because it details DNA quantity along with quality of amplifiable DNA from FFPE tissue.

Bilateral Spontaneous Pneumothorax in Chronic Silicosis: A Case Report

Silicosis is an occupational lung disease caused by inhalation of crystalline silica. People working in occupations like sandblasting, surface drilling, tunneling, silica flour milling, ceramic making, and so forth are predisposed to develop silicosis. Crystalline forms of silica are more fibrogenic than the amorphous forms, highlighting the importance of the physical form in pathogenesis. Lung biopsy is rarely performed for the diagnosis of silicosis as it can easily be detected by occupational history and radiological features. Patients with silicosis can develop complications like tuberculosis, lung cancer, progressive massive fibrosis, cor pulmonale, broncholithiasis, or tracheobronchial compression by lymph nodes. Pleural involvement in silicosis is rare. Spontaneous pneumothorax is a pleural complication that can develop in such patients. Usually in silicosis pneumothorax is unilateral. We hereby report the lung biopsy findings and discuss the mechanism of pneumothorax development in a case of chronic silicosis who, later on died during the course of the disease.

NPM1 mutation analysis in Acute Myeloid Leukemia: Comparison of three techniques Sanger Sequencing, Pyrosequencing, and Real Time PCR

Objective: Nucleophosmin-1 (NPM1) mutations have prognostic importance in acute myeloid leukemia (AML) patients with intermediate-risk karyotype at diagnosis. Approximately 30% of newly diagnosed cytogenetically normal AML (CN-AML) patients harbor the NPM1 mutation in India. In this study we compared the efficiency of three molecular techniques in detecting NPM1 mutation in peripheral blood and bone marrow samples. Materials and Methods: In a single-center cohort we analyzed 165 CN-AML bone marrow/peripheral blood samples for NPM1 mutation analysis. About 30% of the CN-AML samples revealed NPM1 mutations. For the detection, three methods were compared: Sanger sequencing, pyrosequencing, and real-time polymerase chain reaction (PCR). Results: NPM1 exon 12 mutations were observed in 52 (31.51%) of all CN-AML cases. The sensitivity of Sanger sequencing, pyrosequencing, and real-time PCR was 80%, 90%, and 95%, whereas specificity was 95%, 100%, and 100%, respectively. The minimum limit of mutation detection was 20%-30% for Sanger sequencing, 1%-5% for pyrosequencing, and 0.1%-1% for real-time PCR. Conclusion: The sequencing method, which is the reference method, has the lowest sensitivity and is sometimes difficult to interpret. Real-time PCR is a highly sensitive method for mutation detection but is limited for specific mutation types. In our study, pyrosequencing emerged as the most suitable technique for the detection of NPM1 mutations on the basis of its easy interpretation and less time-consuming processes than Sanger sequencing.

ROS1 rearrangement and response to crizotinib in Stage IV non-small cell lung cancer

Background: The frequency of ROS1 rearrangement in non-small cell lung cancers has been reported from 1.6% to 2.3%. Materials and Methods: We examined 105 lung adenocarcinoma patients for ROS1 rearrangement which were negative for EGFR and anaplastic lymphoma kinase. Clinical characteristics of ROS1 rearranged patients and their responses to crizotinib therapy were studied. Results: Of the 105 patients, three cases were positive for ROS1 rearrangement by fluorescence in situ hybridization analysis. All of them showed heterogeneous pattern. All the 3 ROS1-positive patients were females in their forties and started on crizotinib. All of them responded to treatment. One of them developed resistance after 3 months. Another one showed marked systemic response but central nervous system lesions progressed. The third case is doing well till date with inactive lesions on positron emission tomography scan. Conclusions: The frequency of ROS1 rearrangement is low in non-small cell lung carcinoma, but their diagnosis offers patients an opportunity to receive highly effective targeted therapies.

Actively caseating endobronchial tuberculosis successfully treated with intermittent chemotherapy without corticosteroid: a report of 2 cases

Tuberculous infection of the tracheobronchial tree confirmed by microbiological or histopathological evidence with or without parenchymal involvement is known as endobronchial tuberculosis. Chronic cough is the predominant symptom. Expectorated sputum examination for acid fast bacilli is often negative leading to delay in diagnosis. Therefore, bronchoscopy is crucial for early diagnosis and evaluation of the extent of disease. Bronchostenosis is a significant complication of endobronchial tuberculosis that may be present at the time of diagnosis or develops during the course of treatment. Previously, corticosteroids have been used along with antitubercular therapy to prevent or reduce the extent of bronchostenosis; however, their role is debatable as bronchostenosis often develops despite the use of corticosteroids. Furthermore, the duration of treatment varied from 6-9 months of daily therapy in previous series and little is known about efficacy of intermittent antituberculous therapy. Here we report two cases of actively caseating endobronchial tuberculosis successfully managed with six months of intermittent oral antitubercular therapy without corticosteroids.