Molecular detection of mycobacterium tuberculosis in environmental samples and mutational analysis of pncA gene by multiplex allele specific PCR
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Date
2016
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University of Management and Technology Lahore
Abstract
Tuberculosis (TB) has reemerged as one of the main sources of death in the most recent decade. Tuberculosis (TB), one of the basic human diseases, brought about by species from Mycobacterium tuberculosis complex, creating 3,000,000 deaths for each year around the world (WHO, 1996). Five species are included in MTBC. These are M. tuberculosis, M. canetti, M. bovis, M. microtiand M. africanum. The spread of TB is because of migration, the rise of drug resistant strains.
This studyis aimed to identify the M. tuberculosis and M. bovisin environmental samples (drinking water, sewerage water, hospital waste) and bovine milk samples using PCR method. DNA was extracted from bovine milk samples, drinking water samples (tape water), hospital waste (waste water) and sewerage samples. The primers M and S were used to generate amplicons of sizes 318 and 291 bp which represented M. tuberculosis and M. bovisrespectively. 55 samples of milk were tested and the prevalence of TB in these samples was 5.4%. 30 samples of drinking water were tested in this study and the prevalence of TB in these samples was 3.3%. 5 samples of waste water from hospitals were tested and the prevalence of TB in these samples was 40%. 10 samples of sewerage water were tested and the prevalence of TB in these samples was 40%.
The mutations of pncAgene were analyzed in these positive TB samples. Pyrazinamide (PZA) is an important first-line drug in the treatment of MDR-TB. These TB positive samples were subjected to multiplex allele specific PCR. Two mutations were targeted in this study with one being the leucine replaced by proline in the amino acid position 85 and the other being the aspartate changing to alanine in the amino acid position 12. The substitution being the as partate to alanine at amino acid 12 was more frequent as compared to substitution being the leucine replaced by proline in the amino acid position 85.
Presence of TB in these samples is a great health hazard. So control measures are required to stop the transmission of TB.
Description
Supervised by:Dr. Nouman Rasool
Keywords
Tuberculosis, M. canetti, MS Thesis