Comparative Study of Polymerase Chain Reaction (Pcr)and Loop-Mediated Isothermal Amplification (Lamp) for Direct Detection of my cobacterium Tuberculosis in Sputum

Abstract

The risk of spread of infection and emmergence of drug-resistant strain has created the need for a rapid, sensitive and specific diagnostic test for tuberculosis. In addition,clinically suspicious cases that do not give positive result in conventional laboratory test need the development and evaluation of new diagnostic technique, which can identify the etiological agent in rapid way. The study was carried out from October 2006 to November 2007 based at German Nepal Tuberculosis Project, Mycobacterial Research Laboratory, Anandaban Hospital and Everest International Clinic and Research Center in collaboration with Central Department of Microbiology (CDMTU) and Osaka Perfectural Institute of Public Health, Japan. A total of 106 (53 fluorochrome staining positive and 53 fluorochrome staining negative) sputum samples were collected in this study. Out of 53fluorochrome staining positive samples, all the samples 53 (100%) were positive on culture on Ogawa medium and 51 (96.22%) were found to be positive in both PCR and LAMP. Two (3.77%) PCR and LAMP negative samples were positive in culture. Similarly, of 53 fluorochrome staining negative samples, 4 (7.54%) samples were positive and 45 (84.90%) were negative in all culture, PCR and LAMP. There was 1(1.88%) sample which was positive by LAMP and PCR but culture negative and 3(5.66%) samples were positive only by PCR but negative by culture and LAMP. While comparing the microscopy results with culture as gold standard, the sensitivity,specificity, positive and negative predictive values were 92.98%, 100%, 100% and92.45% respectively, however, these values for PCR were 96.49%, 91.83%, 93.22%and 95.74% respectively with reference to culture. Similarly, the sensitivity,specificity, positive and negative predictive values for LAMP with reference to culture were 96.49%, 97.95%, 98.21% and 96% respectively, however, with reference to PCR, these values were 94.91%, 100%, 100% and 94% respectively. This study showed that PCR and LAMP could be a possible diagnostic tool for the confirmation of the smear negative cases that show clinical symptoms of TB. Due to its easy operation without sophisticated equipment, LAMP will be simple enough to use in small scale hospitals, primary care facilities, and clinical laboratories in developing countries if the remaining issues suc has sample preparation, nucleic acid extraction and cross-contamination controls are addressed.