Predictive utility of fluorescent microscopy for detection of drug resistance in patient of pulmonary tuberculosis

Abstract

Tuberculosis (TB) is a global health problem with 480000 new cases being reported annually, 9 percent of which exhibit extensive drug resistant TB. With 9.6 cases per lac population early detection of drug resistance at peripheral levels is must. The study was planned to use Fluorescent microscopy (FM) with Fluorescein Diacetate Ethidium Bromide (FDA/EB) stain to evaluate the viability of the mycobacteria to predict the drug resistance. This cross-sectional analytical study was carried out on all sputum smear positive patients after obtaining the Institutional Ethics Committee clearance over a period of 3 months. Samples were obtained on day 0, 3, 7 and 14. After staining with the working solution, observation was made by two independent observers for viable verses non-viable mycobacteria. FDA will stain the live bacilli and fluoresce green whereas dead bacilli lack the esterase activity and are counterstained by ethidium bromide and appear red. Data was maintained in MS Excel and analysed using tests of proportion and significance. A total of 30 participants were included based on the inclusion criteria. There was a loss of follow up from 14 patients. 4/16 (25%) patients were found be harbor drug resistant mycobacteria strains bases on decreasing ratio of viable to non-viable bacteria on followup. Results when compared with gold standard of Line Probe Assay (LiPA) was found to be highly significant with Chi square value of 11.73 and p value <0.001This study establishes the utility of fluorescent microscopy using FDA and EB to detect drug resistance in patients of pulmonary tuberculosis by evaluation of viable and non-viable at peripheral microscopic centers Fluorescent Microscopy with FDA/EB staining holds a potential to predict drug resistance in sputum smear positive patients at peripheral designated microscopic centers which will reduce financial as well as human resource burden.