Detection of Mycobacterium tuberculosis Complex by a laboratory making PCR Method

Tato, V. Detection of Mycobacterıum tuberculosis Complex by A Laboratory-Made Polymerase Chain Reaction (PCR) Method. Hacettepe University Graduate School of Health Sciences, PhD Thesis in Microbiology, Ankara, 2024. The aim of this study is to demonstrate the effectiveness of the real-time PCR method, developed by targeting the senX3-regX3 gene system and using specific primers, in detecting MTBC pathogen. For this purpose, two different primer-probe sets were designed targeting the senX3-regX3 gene region, which is a specific and well-conserved region for MTBC. In this study, 160 samples obtained by preparing eight serial dilutions of pure cultures of 20 MTBC isolates isolated from sputum samples of patients diagnosed as tuberculosis at Hacettepe University Hospitals Central Laboratory were used. With the addition of twenty negative control group isolates, the total number of samples became 180. Two different laboratory-made real-time PCR tests developed with primer and probe sets designed for MTBC senX3 and regX3 target gene regions were applied to the samples. The samples were simultaneously inoculated on Löwenstein-Jensen (LJ) medium as the gold standard culture method and the results were compared. As a result of the study, the sensitivity of the laboratory-made ('in-house') real-time PCR test targeting the senX3 region was found to be 95.8% and the specificity was found to be 95%. The sensitivity and specificity for the laboratory-made real-time PCR test targeting the regX3 gene region, were found to be 97.5% and 90%, respectively. As a result of this study, promising results were obtained with the senX3-regX3 gene region selected as the target for the laboratory-made real-time PCR diagnostic test developed for the detection of MTBK. It is thought that this region may be a good target region for the development of different molecular diagnostic methods.