Validation Approach (TaqMan) Real Time PCR in the Diagnosis of Tuberculosis-Related Patients (with an emphasis on TB negative) in Mashhad City

Background and Aim : Mycobacterium tuberculosis (TB) is a species of pathogenic bacteria in the family mycobacteriaceae and the causative agent of tuberculosis, remains the leading cause of death worldwide. Sometimes due to a small number of tuberculosis in some cases, in addition to smear and culture as gold standard, Real time PCR (TAq Man) is very helpful in diagnosis due to high sensitivity and specificity. The purpose of this study was to investigate the validation approach (TaqMan) Real Time PCR in the diagnosis of tuberculosis-related patients (with an emphasis on TB negative).Methods : Of all patients referred to Mashhad University of Medical Sciences hospitals referring to the similar symptoms of tuberculosis by the physician, pulmonary (lavage or phlegm) samples and extrapulmonary were taken. At first, patients' basic information was recorded for age, type, sex and type of preparation. The specimens were then evaluated for smear and culture and it was determined that the smear or culture was negative. DNA extraction was carried out to determine the molecular properties of the samples using a DNA isolation kit (Qiagen), then primers and probes were designed using the Beacon designer software and the real-time PCR method was optimized. In this research, the Real-Time PCR Taqman probe was used for IS۶۱۱۰ for diagnosis of tuberculosis. In this research, optimization of methods for use in TB diagnostic laboratories was studied.Results : The sensitivity of the real-time PCR Taqman probe designed to detect Mycobacterium tuberculosis complex was ۱۰۰%. In this study, ۲۹۴ samples were taken, ۲۰ samples were smeared and positive culture, and ۲۱۳ samples were smeared and negative culture, from ۲۳۳ samples ۴۹ samples of TaqMan positive and ۱۸۴ negative TaqMan test samples. ۶۱ samples were unculturable, of which ۱ sample was positive by performing TaqMan test.Conclusion : Classical methods are used to detect Mycobacterium tuberculosis, so in this study, Real-time PCR was described as the fastest method. According to the results of this study, the Real-Time PCR Taqman probe method can be used as a standard method for diagnosis and confirmation of tuberculosis

کلیدواژه ها:

Mycobacterium tuberculosis ، TB (Tuberculosis) and Real-time PCR Taqman probe